CHM/BIOS-372

What does metabolic pathway mean?

Elements of Biochemistry II

Final Exam Study Guide

5/1/2017

Introduction to Metabolism

Course Objectives

∙ Develop an understanding of major metabolic pathway details ∙ Learn the major ways in which metabolic pathways are regulated ∙ Focus on how metabolic pathways are involved in specific aspects of  physiology

What is true about metabolic pathways?

Chemical Reactions in Your Bodies – Fig. 16.1

∙ Complicated diagram shows how reaction pathways interrelate ∙ There is a fair amount of coordinate control

∙ Ex: “Epigenometric reprogramming…” etc. shows that this material is  ongoing and relevant because this article was posted Jan 17, 2017 Section Goals

∙ Introduce concepts and terminology we will encounter  

How do phototrophs and chemotrophs differ in where they get their energy?

If you want to learn more check out What was the nullification crisis in 1832-1837?

regularly in our study of metabolism

∙ To begin our examination of thermodynamics as a  

major player in control of metabolism

Metabolic Pathways/Terms 

A. Terms and Concepts – Fig. 16.2

a. Phototrophs – plants etc. that get energy  

from the sun

b. Chemotrophs – rest of life etc. that gets  

energy from chemical molecules (ingestion)

c. Anabolism – adding up

d. Catabolism – breaking apart Don't forget about the age old question of Who is vladimir lenin?

e. Fig. 16.2 – Biosynthesis is when the body takes simple  

molecules and uses them to make complex metabolites (a  

substance formed in or necessary for metabolism)

B. Ways of Grouping Reactions – Fig. 16.3

a. Pathways for specific goals in metabolism

b. Fig. 16.3 – shows various pathways of proteins, carbohydrates,  and lipids through our body and how they relate to glycolysis,  pyruvate, acetyl-CoA, the citric acid cycle, and oxidative  

phosphorylation

C. Control Mechanisms for Metabolic Pathways

a. Regulation of pathways – diet helps regulate pathways

b. Irreversibility of Pathways – need alternative pathways for back-reactions

c. Committed Steps – makes decision about resources in our bodies (Ex: Incoming AA,  should we break them down or keep them whole?)

d. Isolation by Location

i. Ex: Livers carry out specific reactions as well as other tissues having unique  functions

ii. Ex: Same thing with cells – cytoplasm and membrane surfaces deal with the  catabolism of fatty acids while mitochondria degrades them If you want to learn more check out What are the economic variables of the development of russia?

D. Chemical Logic – Fig. 16.5

a. Nucleophile to electrophile

i. ^Fig. 16.5 shows examples of nucleophilic forms

ii. Nucleophile – a chemical species that donates an electron pair to an  If you want to learn more check out What model shows the flow of memory between 3 different types of memory?

electrophile to form a chemical bond in relation to a reaction – All molecules or  ions with a free pair of electrons or at least one pi bond can act as nucleophiles  – Because nucleophiles donate electrons, they are by definition Lewis Bases

iii. Electrophile – a reagent attracted to  

electrons – They are positive or neutral  

species having vacant orbitals that are  

attracted to an electron rich center – It  

participates in a chemical reaction by  We also discuss several other topics like What is the meaning of solar flux?

accepting an electron pair in order to bind  

to a nucleophile

b. Group Transfer Reactions Include – Fig. 16.7

i. Larger structures are typically transferred  

in anabolic and catabolic reactions

ii. Common types are acyl, phosphoryl, and  

glycosyl group transfers

iii. Ex: Fig. 16.7 – glucose and ATP combine  

with a phosphate group to make a  

trigonal bipyramid intermediate and then  

form Glucose-6-phosphate and ADP (the  

phosphate group is a group transfer)

c. Oxidation-Reduction Reactions

i. Oxidation/Catabolic – breaking down - reactions are accompanied by reduction  of an electron carrier

a. H3C(CH2)3OH → H3C(CH2)2CH=O (+ CO2 as a byproduct)

ii. Reduction/Anabolic – making up - reactions need lots of electrons from electron  carriers

a. H3CCOO- → H3C(CH2)nCOO

d. Eliminations, Isomerizations, Rearrangements

i. Elimination of water Don't forget about the age old question of What is the meaning of slum?

ii. Keto- to Enol- conversions

iii. Rearrangements are less common

e. Make and Break Carbon-Carbon Bonds. All Types of Reactions

i. Ex: Breakdown of glucose to carbon dioxide involves separating all 6 carbons  from each other to produce 6 CO2

E. Techniques for Studying Metabolism - Fig. 16.17

a. Metabolic Inhibitors and a little intuition

b. Genetic Defects – phenylketonuria is a well-known example in which patients are unable  to process pheylalanine

c. Genetic manipulation to generate  

blocks in a pathway – pick and  

choose through careful  

consideration

d. Isotopes (one example in next  

paragraph)

e. Isolated Organelles – Ex: Can do a  

lot with just a mitochondria

f. Metabolomics, etc.

g. Fig. 16.17 – using radioisotopes to  

figure out how heme is made using  

radioactive isotopes to find the  

individual building blocks

Thermodynamics in Biological Systems 

A. Basic Termodynamics

a. Biological conditions!!  

Remember that the body is in  

aqueous solution and  

physiological pH is 7

b. Find formulas necessary for thermodynamic calculations in chapter 3, p. 58 in textbook  (same as above tho)

B. Steady State or Death!! – Fig. 16.31

a. Equilibrium is in a closed system, while biological systems are open!

b. Therefore, biological systems are never in equilibrium, they are in steady state c. Ex: Photos of a river will look the same from one day to another but we know the water  molecules are not the same

d. Ex: Same with pathways in our bodies –

individual molecules are changing but levels stay  

relatively the same

C. Coupling Reactions – Fig. 16.23

a. The ending dG value of -16.7kJmol-1 is still  

negative (favorable) even though the endergonic  

half reaction requires energy and is unfavorable

b. Enzyme Coupled Reactions Fig. 16.32

i. Have a test tube with aqueous buffer

ii. Energy barrier for overall reaction is too  

high so we know that it could possibly be  

coupled

iii. Enzymes create  

mechanisms or  

environments in which  

that reaction can take  

place (some sort of kinase)  

iv. Allows you to obtain  

intermediates more easily

v. Enzymes make  

intermediates more  

favorable

ATP, Energy, and Thermodynamics 

A. ATP Structure – Fig. 16.22

a. Sometimes ATP is written as AT~P to denote  

that there is so much energy within the  

phosphoanhydride bonds

b. ATP is high energy – how favorable hydrolytic E  

is

c. Provides a source of energy (“energy  

currency”)

d. Why are ATP Phosphoanhydride Bonds “High  

Energy”?

i. Charge repulsion

1. ~3 negative charges in close  

proximity that repel each other

2. Taking one away takes away one negative charge which means less  

repulsion/more stability/more favorable

ii. Resonance Stabilization – Fig. 16.24

1. When the phosphoanhydride bonds  

break, each compnonent has 2  

resonance structure that stabilize the  

break

iii. Solvation Energy

1. Packing water around the  

ATP/ADP/AMP

2. Deals with free energy

iv. Entropy

1. Body likes 2 (ADP + Pi) or 3 (AMP + 2Pi) pieces better than one piece  (ATP)

e. Other “High Energy Molecules” – Table 16.3

i. Phosphoenolpyruvate and 1,3  

Bisphosphoglycerate can be more  

favorable than ATP pathways  

combined

ii. Phosphocreatine (like creatine  

powder) is an intermediate that can  

store energy

iii. The point is that ATP is in the middle  

of the list

f. Source of Phosphoenolpyruvate Free Energy of Hydrolysis - Fig. 16.25 i. Hydrolytic reactions  

is more favorable  

than the hydrolysis  

of ATP

ii. The reaction is  

driven by the ability  

to interconvert enol  

to keto because keto  

is more stable  

(favorable)

iii. Provides very  

favorable hydrolysis

iv. Can be coupled to  

the synthesis of ATP

B. Use of The Free Energy – Fig. 16.27

a. ATP is in the middle – this is why it is used as  

“currency”

b. ATP is relatively stable in solution which is good  

because we know it will be around for reactions

c. Specific Uses for ATP

i. ATP + NDP → ADP + NTP

ii. Phosphorylation of glucose and other  

intermediates as well as Amino Acid activation to  

amino acyl-tRNA in Fig. 16.29

iii. Activation of a Fatty Acid

iv. Enzyme in question links MP to carboxylic acid  

carbon of fatty acid and becomes hydrolyzed

v. CoA exchanges AMP

vi. In cells, ATP is always active

vii. Hydrolyzing both phosphoanhydride bonds from  

ATP gives us 2x the energy which we can  

calculate because getting ADP and Amp is  

through enzymes that convert  

phosphoanhydride bonds

viii. When we are adding up E we are using equivalence of 1 phosphoanhydride  bond from ATP so we can add it up in our heads that way

d. Synthesis of ATP - Fig. 16.23

i. This coupled  

reaction is  

favorable because  

of free E

ii. Some molecules  

have enough free E  

to provide for the  

synthesis of ATP

e. Energy for a Person

i. Simplistic summary bc of different activities:

ii. Need amt of  

ATP =~ body  

weight

iii. Production of  

ATP under  

strenuous  

activity is  

much higher  

bc of survival  

etc.

f. Maintaining ATP Availability

i. ATP concnetrations is maintained because we need it so much in pathways,  brain functions, being alive in general, phosphorylation of glugocse to Glucose 6-phosphate etc.

1. 2ADP → AMP + ATP (catalyzed by adenosine kinase)

ii. Cells in body monitor the amounts/concnetrations of ATP, ADP, and AMP iii. E charge kept at about 0.9 (measure of phosphoanhydride bonds) 1. E Charge = {[ATP] + ½ [ADP]}/{[ATP] +[ADP] + [AMP]}

iv. 0.9 means 90% in cell so we can make more if needed for when you see a spider  etc. You have enough ATP for any activity that has to happen

Oxidation-Reduction and Energy 

∙ Glucose and other small molecules are the source of energy

∙ As they are degraded to small molecules, energy is harvested to produce ATP ∙ Most of the reactions are not transfer reactions that produce ATP directly, but rather reactions that  result in reduced electron carriers (NAD and FAD)

∙ Can come in multiple forms

∙ Usually directly provides E for ATP synthesis

∙ Reduced electron carriers contribute their energy to the synthesis of ATP

∙ The available energy in a single oxidation step is often much greater that that needed for ATP  synthesis (details in Ch22)

∙ We can obtain energy values for reactions based on evaluating the electron potential difference  between reduced and oxidized molecules

∙ dG = -nFdE can be rearranged to yield the NERST equation

A. Electron Carriers Employed

a. NADH is the most commonly used electron carrier in catabolic (destroying) pathways  and, like ATP, is maintained at very high concentrations

b. Unlike ATP which is used in synthesis and degradation, biosynthetic pathways typically  employ NADPH as the electron donor during synthesis

Glycolysis

Section Goals

∙ To develop an understanding of the glycolytic pathway and catabolic metabolism of sugars for  energy

∙ To use this commonly studied pathway as a system to study how chemistry controls metabolism and  how metabolic reactions are really organic chemistry reactions catalyzed by enzymes The Glycolytic Pathway – Fig. 17.1

∙ Glucose with 2 molecules of ADP and  

NAD+ each to make Fructose-1,6-

bisphosphate (F-1,6-BP)

∙ Get 2 pyruvate

∙ Citric acid cycle, oxidative  

phosphorylation etc. etc.

∙ On the sides there are two possible  

scenarios when there isn’t oxygen of  

there is limited oxygen (anaerobic)

∙ Fermentation mechanisms to  

regenerate NAD+

A. An Overview of The Reactions –

Fig. 17.3

a. Think about what is most  

critical for the core of the  

pathway

b. Can think of as two portions

c. Start out with glucose and commit 2 ATP’s to jumpstart the pathways

d. Need ATP around or the pathway won’t even start

e. The top part is a preparatory pat of the pathway but at the bottom is where we really  harvest energy

f. Each 6 carbon sugar gives us two 3-carbon pieces

g. Later we will talk about how the electrons from NADH are used to make ATP (?) h. Use glycolytic pathway to pull all of the pieces together

B. Hexokinase – P. 597

a. First part of the reaction is catalyzed by  

kexokinase

b. Is part of a family of enzymes

c. “Kinase” refers to an enzyme that can add a  

phosphate group to another metabolite using ATP  

as a source of phosphate

d. “Hexo” refers to 6 carbons

e. Glucose is not the only substrate for some  

versions of hexokinase

f. Very straightforward reaction

g. Use of magnesium, commonly found with ATP in  

the body (activates it), can use other divalent  

metal ions but magnesium is usually there

h. Product is phosphate groups on 6 carbon of  

glucose sugar

i. Often we draw out glucose is hemiacetal form bc  

typically in this form but can also be in an open  

conformation

j. Thermodynamic issues – Table 17-1

i. Shown in textbook in a different location

ii. dG value based on a study done in a rat heart  

muscle (naught)

iii. If you put in other variables like amounts of  

ATP present in the body, becomes much  

more favorable

iv. DO NOT MEMORIZE SPECIFIC Dg VALUES – just know that it is favorable v. This step is not limited to first step in glycolysis – used for several purposes like  from a metabolism perspective is activates glucose but it also is phosphorylated  immediately within the cell bc this trap it there – by putting a phosphate on  glucose you’re trapping it in the cell – most cells take up glucose in proportion  to amount in bloodstream bc passive transportation (high  

conc to low conc) and this is a way to accumulate glucose in  

cells so that you have the energy

vi. Ex: Jelly donut with lots of sugars, no complex  

carbohydrates, makes it to bloodstream, levels rise, cell will

immediately phosphorylate the glucose. If they don’t want more, they’re not  going to have as many or any transporter structure on cell surface

C. Glucokinase in liver

a. Enzyme catalyses the exact same  

reaction except in the liver – sometimes  

referred to as hexokinase 4

b. Reaction is different b/c glucokinase uses  

only glucose, has a Km of 10mM, and  

liver supplies glucose during low blood  

glucose

c. Glucokinase waits for glucose conc to be  

high to phosphorylate

d. Important bc don’t need to trap glucose in cells bc its in the liver which supplies glucose  to the liver (liver is responsible for sending glucose to body when you haven’t just eaten  a jelly donut)

D. Phosphoglucose isomerase – Fig. 17.6

a. Need to transform glucose to fructose

b. Opening ring and making it into a ketofructose when you  

close the ring back up

c. Keto group comes at position 2

d. Position 6 is still phosphorylated

e. Carbon 1 is not in the ring (drawn above)

f. Carbon 2 is the ketocaron so we have the carboxyl grop and  

the CH2OH

g. Conditions – dG slightly unfarovable in regular conditions  

but becomes more  

faorable in the actual rat  

heart conditions but its  

still a guestimation

h. Enzyme constantly interconverts the sugars

i. Need to be adding in glucose and taking out fructose –

relative concentrations are essentially identical

E. Discussion – what does it mean?

a. Given Keq for phosphoglucose isomerase reaction of 0.5  

(approx.) and a steady state value of 0.2 G6P, what is  

steady state concentration of F6P?

b. Answer is 0.1 which makes sense b/c a Keq of 0.5 means it  

is only turning half of the G6P into F6P and half of the G6P  

concentration of 0.2 is 0.1

F. Phosphofructokinase – P.600

a. Under standard conditions of rat heart it is more favorable  

so it is quite a favorable reactions

b. Adds another phosphate group to F6P to make Fructose

1,6-bisphosphate (FBP)

c. This is the committed reaction for glycolysis! And it is crucial for pathway to be  regulated

i. Tells you that every FBP is going to be used for glycolysis

G. Aldolase – Fig. 17.9

a. Reactions are  

reversible

b. Standard free energy  

for this is  

unfavorable and in  

vivo is slightly  

favorable

c. Open up ring to get  

reaction to occur

i. Diagram is color coded

ii. Ketocarbon is crucial to link sugar to enzyme (lycine)

d. Preserved carbon 2 as ketocarbon

e. Preserved position of phisphate as well

f. Product 1 is Glyceraldehyde-3-phosphate  

(GAP) (red)

g. Product 2 is dihydroxyacetone phosphate  

(DHAP)

h. Now have two unique carbon fragments – will ultimately interconvert each one H. Discussion continued

a. Keq’ = [DHAP][GAP]/[FBP] = 2x10^-4 for aldolase – is there a concentrations of FBP  where the Gap concentration would be equal? Note that DHAP is about 2x[GAP] b. Answer is about .1 mM

c. What looks like and unfavorable reactions is actually favorable bc of concentrations  d. Functional situation is different than the thermodynamics

I. Triosephosphate isomerase – Fig. 17.3 P. 603

a. Makes two 3-

carbon sugars  

interconvert b/w  

each other

b. Phosphates end  

up being in the same position even though they were on opposite ends before c. Taking ketocarbon and moving around  

electrons

d. GAP gets processes farther

e. Concentrations of DHAP are higher even  

though can be turned into GAP

f. End up having molecules flowing through pathway with net processing of the glucose J. GAP Dehydrogenase (GADPH)– Fig. 17.13

a. Finally harvesting some energy

b. Oxidizing the molecule  

(GAP)

c. Using NAD+ as an electron  

acceptor (gets reduced) this  

is where we get NADH

d. Not surprising that you can  

oxidize aldehyde to  

carboxylic acid

e. Can add phosphate onto carboxylic acid carbon without the use of ATP, can use GADPH! f. Thioester is reactive so can be replaced by incoming phosphate group

g. 1,3-BPG is a high energy molecule that can be used to synthesize ATP

h. Have transferred electrons into electron carrier NAD3H

i. With PGK (phosphoglycerate kinase) the dG is favorable but without it is unfavorable.  There is fairly good reason to believe that the enzymes are somehow near each other  and fed directly between enzymes – not free-flowing

j. Is a dehydrogenase

k. The dG is close to 0 which is important because a lot of reactions in the pathway are  close to 0 which has an impact when we talk about the overall reactions

K. Phosphoglycerate Kinase – Fig. 17.16

a. Reaction that makes our first ATP

b. Transfers phosphate groyp to ADP  

c. Hydrolysis of this  

molecule (hydrolytic  

release) makes it a  

favorable reaction

d. Table 1 says dG is  

favorable but that the  

reaction by itself is  

much more favorable

e. Have made first  

phosphate (have made 2 technically bc one glucose makes two 3-carbon pieces f. At this point we have paid back the cost of activating glucose

g. And have made saving of making NADH

h. Name seems like is the reverse reaction but in fact names the product (in vivo can  operate in both directions tho)

i. Estimated dG is close to 0 in vivo

L. Phosphoglycerate mutase – Fig. 17.18

a. Note that 2,3-bisphosphoglycerate can  

be made at low levels and serves specific  

other functions in RBC

b. Net reaction is 3 to 2

c. dG is close to 0 (again) in  

vivo

d. Reaction is reversible – operates at essentially  

equilibrium values

e. “Mutase” = moving around/changing location of  

phosphate group

M. Enolase – Fig. 17.21

a. Don’t have to know electron movements, just the  

substrate (2PG) and product (PEP)

b. Reaction removes water, creating a double bond

c. Operating at  

equilibrium (dG  

close to 0, again)

d. By the last two  

reactions we  

produce PEP which is a good donor of a phosphate  

group to make ATP

e. “Enolase” = removing water and creating double bond

N. Pyruvate Kinase – Fig. 17.22

a. Named for the reverse reaction that  

doesn’t happen favorably

i. dG in Table 17.1 shows  

reverse reaction is hard

ii. Has a Magnesium

iii. Ends up giving us ATP

iv. Two 3-carbon fragments =  

net yield 2 pyruvates = 2  

ATPs at this step

v. Even in vivo we can estimate  

that standard free energy is still favorable

Glycolysis Summary

∙ Glucose + 2NAD+ +2ADP +2Pi → 2NADH + 2 Pyruvate + 2ATP + 2H2O + 4H+ ∙ Will discuss what happens when we don’t have enough oxygen (anaerobic)

Regulation of Glycolysis 

A. General Considerations in Regulation

a. Enzyme activity can be altered,  

Table 17.2

b. Think about thermodynamics etc.  

with each reaction and how they  

are/can be in flux

c. End up with situation in which each  

reaction operates at dG=0  

(equilibrium) values so you can’t moderate activity at these steps

d. Can only control pathways in which the reations have a very favorable dG value in vivo:  kexokinase, PFk, PK

i. Hexokinase – high levels of G6P inhibit it, step is controlled not to control  glycolysis (bc not committed)

ii. PK – inhibited by ATP, activated by AMP, PEP,FBP, and low levels of ATP in  muscles particularly – also not controlled for glycolysis but for the relationship  between glcose synthesis vs. degradation (need to inhibit for synthesis)

iii. PFK – main place to regulate glycolytic pathway

1. ATP is a substrate for PFK, can also bind to second allosteric site on  

enzyme where ATP can modulate activity of enzyme

2. Think back to discussion of energy charge ATP kept high at 0.9  

concentration, in vivo this becomes as issue, well above 0.9 causes  

problems, ATP bins to allosteric site and slows down activity of PFK

3. Citrate and PEP also inhibit. Citrate – don’t have very much info

4. ADP and AMP primarily are “deinhibitors” – functioning to activate PFK  

by not allowing ATP to inhibit it

5. FBP is an intermediate in the pathway (product of PFK)

6. F-2,6-P – closely related ^ and is the potent activator of PFK, molecule  

made by a separate system – allosteric activator – binds to alternate site  

and modulates activity of enzyme (activates) so that glycolytic pathway  

runs at a higher rate

7. F6P, we won’t talk about

8. NH4+ and Pi, we won’t talk about

iv. PFK

1. Committed step  

2. Control at PFK occurs through cell’s energy

3. Control at PFK occurs through response to hormonal signals

4. Allosteric modulation of an enzyme changes the Km, not the dG’ of a  

reaction, therefore, the reaction is no less favorable, but a higher  

substrate concentration is needed to reach maximal activity

v. Cell Energy Modulation of PFK – Fig. 17.33

1. Role of energy charge is where  

there is always some ATP and some  

AMP, amount of ATP is dependent  

on energy charge

2. We don’t see ADP at all, as ADP  

conc rises, make 1 ADP and 1 ATP

B. Flux Through a Pathway

a. Flux is determined by several factors

b. Allosteric Control (or Covalent)– Fig. 17.32b

i. With an activator like ADP we get the  

structure where F6P binds really well

ii. Sidechain flips depending on active vs.  

inactive state

iii. Arginine has a guanidinium group that is positively charged so it maked a  favorable binding site for F6P (negative b/c of phosphate group)

iv. Inactive form – arginine is flipped so have glutamic acid that has a negative  charge

v. Example of how to use allosteric modulator to alter substrate affinity and  therefore binding affinity – how to regulate activity

c. Hormone Control and F-2,6-P – Fig. 18.25

i. Needed to get maximal activation of PFK in any tissue

ii. Therefore, PFK-2 production is controlled differently in different tissues iii. Enzyme that makes this is the same enzyme that degrades it

iv. Enzyme activity in liver is therefore controlled by phosphorylation

v. Covalent modification is altering activity of an enzyme

vi. Funky enzyme – has 2 activities

1. That take slow conc of F6P and adds phosphate at position 2 (molecule  is the major activator of PFK in pathway we were just talking about)

2. Removes phosphate from activator

vii. In case of liver enzyme – can be particular location related to hormonal control viii. Whole system is complicated because of similarity in names

ix. Remember intermediate has two phosphates at the ends – therefore the one  with the  

phosphate at  

the 2 position  

is the  

intermediate

x. Remember  

enzyme  

pathway  

being spoken about is PFK

d. Substrate Cycles

e. Genetic Control – Fig. 18.24

i. Hexokinase is very good because it’s very  

efficient (ass phosphates to trap in cell)

ii. Glucokinase is slower

iii. Genetic control bc tissues can control  

which enzyme they synthesize which  

changes productions

C. Hormonal Control of PFK

a. When an organism needs to activate glycolysis in a tissue is can turn on the synthesis of  an allosteric control molecule (F-2,6-P0 that will bind to PFK to activate it

b. Tissues respond to hormones differently

c. Heart muscle must start glycolysis during activity

d. Liver doesn’t need to start glycolysis, and in fact must turn it off if glucose is needed in  the blood

e. Hormone Control and F-2,6-P – Fig. 18.25 (again)

i. F-2,6-P is  

needed to get  

maximal  

activation og  

PFK in any  

tissue

ii. Therefore,  

PFK-2  

production is controlled differently in different tissues

iii. Turns out there are a number of different variations of the enzyme b.c there’s  the liver one, the muscles one, another one in other tissues that’s not controlled  by direct hormonal modulation

iv. This kind of modulation allows us to have different kinds of activities

v. Strong area of research

f. Example of a Current Article

Anaerobic Uses of Pyruvate – Fig. 17.24

∙ There are two types of systems that deal with  

O2 deficiency. Both allow organisms to  

continue glycolysis for ATP production without  

O2

∙ They take pyruvate and let pyruvate be an  

electron acceptor

A. Homolactic fermentation, the skeletal  

muscle solution:

a. Pyruvate is reduced to having a hydroxyl group

b. So now have a way to get more NAD+ so we can  

regenerate/continue to use the glycolytic pathway

c. Ex: Hunters know that if you run an animal to exhaustion,  

you get sour meat because of the acid and lactate that build  

up and cause the muscle to be sore when the anima  

undergoes anaerobic activities

d. NADH concentration builds up, don’t have NAD+, can’t keep  

going with glycolysis

e. Get lactate, builds up in muscle to some extent, depending on flow of blood that will  move lactate

f. When lactate gets in the bloodstream, moves back to liver and in liver, where there’s  usually some oxygen, gets reconverted and carries out reaction backwards, produces  more glucoses and muscles can keep functioning

g. Have different forms of lactate dehydrogenase – different tissues have different version,  just know lactate dehydrogenase reaction and that it’s reversible based on levels of  NADH and NAD+

B. Alcoholic Fermentation, yeast’s choice – Fig. 17.25 and 17.26

a. Carry out 2 reactions

b. The first is catalyzed by pyruvate  

decarboxylase, the second is  

alcohol dehydrogenase

c. Yeast can survive in quite a bit of  

ethanol, but its’ competitors can’t

d. We can’t undego alcoholic  

fermentation, but the  

microorganisms in our bodies can

e. Acetaldehyde ultimately gets  

converting to fat

i. Ex: The freshman 15 – production of fat from alcohol that gets consumed – done by reversing this reaction

f. This reaction depends on a cofactor/coenzyme. Not something we have to memorize – don’t learn structure

g. Fig. 17.27 also preview aerobic pyruvate use

i. Cofactor is what we think of as the  

vitamin “thiamin” and if you don’t get  

enough thiamin you end up with huge  

problems because you can’t go  

through a lot of metabolic pathways –

yeast don’t worry about this

C. Alcohol DH Functions Just Like Lactate DH – Fig. 17.30

a. Facilitates the same  

type of hydrolysis

b. “Dehydrogenase” –

typically named for the  

structure that is more  

reduced – carrying electrons

c. Named for reaction from lactate to pyruvate even though we’ve  

been talking about them in the opposite direction

d. Both are set up so you can keep doing glycolysis even when  

there isn’t enough oxygen

D. Summary of anaerobic glycolysis:

a. Muscles, when they’re stressed get glycose to 2 lactate, get net gain of 2 ATP b. In yeast, get glucose to 2 ethanol, this is slightly more favorable

c. Remember that we gain 2 ATP in either system

d. Therefore, fermentation is only 26-31%  

efficient (in vivo is better)

e. Aerobic systems are somewhat better  

because they’re not working with standard  

conditions in terms of ATP  

concentrations/levels

Other Sugars 

A. The basic glycolytic pathway is to break up most sugars, not just glucose

a. Won’t be asked on a test to walk through the reactions

b. Might be asked if a particular sugar can be used in glycolytic pathway

B. Mannose – Fig. 17.37

a. Pretty easy to be put  

through glycolytic pathway  

because it can be easily  

turned into FGP

b. “Kinda boring” bc there is  

nothing different in the pathway

C. Fructose – Fig. 17.35

a. Important bc we get a lot since we usually eat simple sugars

b. Ex: fruit is not enough to be problematic, lots of sweet beverages gives you almost too  much fructose

c. A lot of fructose gets taken  

up in the liver which doesn’t  

have hexokinase it has  

glucokinase which is specific  

to glucose

d. Fructokinase

e. Aldolase gives you DHAP  

and GAP without one part

f. Can use ATP at this step –

not so critical to understand  

details, more important part  

is

i. We have a  

regulatory system  

deciding how much  

sugar is going to get processed immediately or going to be stored

ii. There is no regulatory enzyme here

iii. All fructose in liver gets processes

iv. If we jack up levels of intermediates by not controlling access to pathway, end  up trapping phosphate in cells and don’t have inorganic phosphate, can’t make  ATP

v. Serious problem

g. There are metabolic diseases where the problem is worse and situations where people  with too much fructose in their diets get problems with their livers

h. Just know that liver has to use alternate pathway for fructose

D. Role of fructose and fructokinase – article

a. Example of continuous series of studies on this metabolic scheme and how it can be  problematic

E. Galactose – Fig. 17.63

a. Gets in our diet through milk

b. Part of lactose which is a disaccharide

c. Biggest issue people know of is lactose intolerance where they can’t  

properly digest lactose sugar and digest it into the monosaccharides

d. Are also problems with galactose itself

e. There is a specific kinase that phosphorylates the 1 potision

f. Enzyme crucial for processing is an enzyme that creates an activated  

version of galactose – transfers UDP which is used to put sugars in an  

activated form

g. Activated galactose can be interconverted with glucose

h. Normally if we need to  

make glycoproteins of in a  

lactating female who is  

making a lot of lactose –

we don’t have an issue  

with making lactose but  

enzyme 2 is sometimes  

defective so there is  

evidence of individuals  

who are defective in that  

enzyme in their diet so  

they don’t get any  

galactose in their diets  

(genetic defect)

i. **Galactose can get converted to glucose, can go through glycolytic pathway**

Tricarboxylic Acid Cycle (TCA Cycle)

∙ Not the next section of the book, but rn we have pyruvate hanging around and we need to do  something with it

Section Goals:

∙ Explore the most significant pathway in terms of oxidations of all metabolites ∙ Also, to develop an understanding of how this oxidation is regulated, to help us examine the ability  of enzymes to turn biochemical into chiral molecules and to look at the use of radioisotopes in  studying metabolism – something that is first introduced in initial chapter

Introduction – Aerobic Catabolism – Fig. 21.1

A. Pyruvate Dehydrogenase Cycle

a. One point is that the way it is drawn suggests that its  

part of the citric acid cycle, but its’ not, it’s completely  

separate

b. A lore of Acetyl-CoA comes from fatty acid degradation  

and degrading amino acids

c. Figure starts with pyruvate – all of the carbons in this  

cycle get eliminated as carbon dioxide

d. The second point is that they have chosen to put 2  

enzyme linked intermediates on this diagram

e. If she asks to draw out cycle, don’t have to write out  

these intermediates

Pyruvate Dehydrogenase Complex – Acetyl CoA production 

A. Fig. 21.6

a. The enzymes in this set of  

reactions are part of a huge  

multienzyme complex

b. Figure is an overview of  

what is happening in  

pyruvate dehydrogenase  

activity

c. TPP – cofactor we already  

talked about

d. Release a carbon dioxide, very similar to yeast enzyme

e. 2 carbon fragment is transferred to E2 and is oxidized with electrons transferred into E2  then remaining 2 carbons get released as Acetyl-CoA

B. PDH is a Multienzyme Complex – Fig. 21.4

a. This is a huge comples set up so you can process a lot of pyruvate at the same time b. Has E2 core and E1 & E3 shell

c. E1 = Pyruvate dehydrogenase (PDH)

d. E2 = Dihyprolipoyl transacetylase  

(orange)

e. E3 = Dihydrolipoyl dehydrogenase (pink)

C. Five Coenzymes are Involved

a. TPP – bound to E1

b. Lipoic acid – covalently linked to E2 - huge arm to move things from core to shell c. Coenzyme A (CoA) – substrate for E2 - activates pyruvate and allows it to bind d. FAD – bound to E3 coenzyme - is an electron carrier

e. NAD – substrate for E3 - soluble cofactor that is the ultimate place to which hthe  electrons are transferred

f. ***One more thing is thing is that all of the processing of pyruvate we’re talking about  happens in mitochondria***

i. One of the things that has to happen is that the pyruvate has to get in there

D. Thiamine pyrophosphate (TPP) Bound to E1

a. TP operates almost exactly as in pyruvate decarboxylase  discussed in yeast fermentation except that the two carbon  fragment here is transferred to the lipoamide cofactor

b. In both cases the function is to decarboxylase pyruvate E. Lipoic Acid covalently linked to lysine in E2 – Fig. 21.7 & Fig. 21.8  a. In transfer process, electrons still in 2 carbon  

fragment get moved so they end up in enzyme  

(lipoic acid), carbon gets oxidized

b. Rn it’s a thioester, but before it was a ketone

c. Don’t need to know structures again

d. Electrons gives you reduced form

e. Depending on version of enzyme – have 1 or  

several lipoamide side chains (cofactors attached)  

looks like a long arm – allows you to move 2 carbon  

fragment around in the compound

F. Kidney PDC – Fig. 21.11c

a. Information is based off of bovine  

kidney

b. Taken a combination of structural  

studies and have put together a  

model

c. In our enzyme complexes we have  

an additional subunit that plays a  

role in linking this together

d. Initially didn’t have E3 but now  

we do

e. Large complex and having arm is  

useful if you have components outside and buried

f. Ex: Another interesting thing is that lipoic acid is really  crucial in terms of arsenic poisoning, inhibits lipoic acid.  Darwin – took medicine accidentally – and napoleon –

probably poisoned by his wallpaper – arsenic stops  

processing of glucose and the TCA cycle proper – don’t  have the ability to use it

G. Coenzyme A (CoA) Substrate for E2 – Fig. 21.2

a. CoA is a way for enzymes to recognize that we have the  

activated acetate

b. It’s an activation group, a way to store energy

c. Other molecules can be linked to CoA

d. Accepts the acetyl group from the lipoamide leaving  

reduced lipoamide

H. Flavine-Adenine dinucleotide (FAD) Bound to E3

a. It is reduced by lipoamide so that the lipoamide  

becomes reoxidizes to the disulfide structure that can  

participate in another reaction

b. This is a standard FAD coenzyme electron carrier

I. Catalytic Reaction – Fig. 21.14

a. E3 and FAD are transient and (don’t  

memorize electrons) extended arm can  

move between active sites of enzymes of  

complex and becomes associated  

transiently with active site of E3, loss of  

proton, rearrangement, so by this stage we  

can have cofactor reoxidized and electrons  

sitting in FAD

b. Get an NAD+ that accepts electrons for this  

scheme (soluble e- carrier)

c. Unusual reaction because normally we  

don’t move from FAD to NAD+, but in the  

case of an enzyme bound electron carrier,  

the enzyme active site has a huge effect on its properties – redox potential d. For a family of enzymes that carry FAD and are moving electron at redox state from  keto-aldo carbon for oxidation of carboxylic acid, allows for favorable energy J. Nicotinamide adenine dinucleotide (NAD) Substrate for E3

a. NAD is, again, a classic electron carrier also used throughout metabolism b. It is mobile and removes electrons from enzyme-bound FAD, regenerating the enzyme c. This is one example where FAD donates electrons to NAD

d. Usually it is the other way around

K. Regulation of PDH – Fig. 21.17a

a. The local cell situation can  

regulate PDh using both NADH  

and Acetyl-CoA

b. Without oxygen, NAD hbuilds up  

with no place to deposit  

electrons, lipoamide cofactor  

stays in a reduced state, enzyme  

process slows down/stops

completely depending on how high concentration gets

c. If there’s oxygen and Acetyl-CoA isn’t getting processoed, slows down again b/c CoA  levels will be low bc A-CoA levels high, hard to transfer out A groups

d. 2 products of the comples that negatively regulate activity of the complex, simply by  slowing down the rate

L. Eukaryotic PDH Regulation p Fig. 21.17b

a. Mammals

b. Regulation here is still  

based on cellular needs,  

not on hormonal control

c. Dependent on various

activators/inhibitors

d. E1 is active in  

unmodified form, if it  

gets phosphorylated,  

then it is inactive

e. Each E1 is separately  

regulated by this system

f. Mammals have 2 extra enzymes, pyruvate dehydrogenase phosphatase and pyruvate  dehydrogenase kinase

g. Kinase activated by the two products we just talked about, inhibited by low energy  situations such as high concentration of ADP and moderately high levels of calcium h. Phosphatase modulated a lil bit so that its often fairly active and you have to keep  having the kinase functioning if you’re going to keep E1 inactive otherwise phosphates  will keep being removed by it

i. In our cells there are two mechanisms, this one and the covalent modification of E1 The Citric Acid Cycle – Fig. 21.1

A. Ex: Extra sugar from coffee, doughnut,  

etc. This patheway is functioning

a. If it’s not glucose and its been  

more than 12 hours since  

you’ve eaten it, most likely A

CoA is coming from  

breakdown of AA  

b. Intermediates are regenerated  

in the process – don’t have to  

remember two enzyme linked  

intermediates when  

memorizing stuffs

B. Citrate Synthase – Fig. 21.19

a. Takes Oxaloacetate and A-CoA

b. First step is going from top to  

bottom and second and third step  

are on right hand side of slide

c. The way in which this works is  

always from the same face of the  

oxaloacetate

i. In organic chemistry, using  

metal or other catalysts,  

don’t normally have that  

opportunity

ii. In enzymes, it’s very directional – citrate synthase is making a pro-chiral  molecule

iii. Think of it as sitting down on a flat surface, never have it flipped over

iv. Bc of this, the two apparently identical conformations of central carbon can  actually be differentiated both in terms of making and using in the next step d. Release of product from enzyme relies on hydrolysis of the CoA

e. Reaction is very favorable both in vivo and test tube

f. If oxaloacetate concentration is super low, need to figure out how to make reaction  favorable and this is one way, by having very favorable free energy, end up combining  two reactions

C. Aconitase – Fig. 21.20

a. Looking at C3 of citrate and pro S arm  

with carboxylic acid and R arm

b. If the enzyme needs to associate the  

carboxylic acid on C3, and the hydroxyl  

groyp with the iron sulfur cluster, you  

can’t actually rotate this around, it  

doesn’t work – kinda complicated

i. Think of it as your hands, palm is  

C3 etc. can’t flip around hands

ii. Taking molecule that isn’t chiral  

and bc using enzyme, are able to  

specifically recognize 2 “chemically identical” molecules

c. Moves hydroxyl group

d. Removes water and we get double bond – bond between C2 and C3

e. What happens within enzyme is once you do that it is very favorable to flip this  molecule within the enzyme (look at plains)

f. Can then add water to double bond and now have carboxyl group on C2 instead of C3 g. Totally reversible, goes both ways

h. Not a large driving force either way

i. Operates close to equilibrium

j. Contonuously making citrate and using isocitrate

D. Isocitrate Dehydrogenase – Fig. 21.21

a. Moving hydroxyl group,  

always moving it in this  

direction

b. Always know which carbons  

came from oxaloacetate  

(1,2,3,4) and which came  

from A-CoA

c. Uses NAD+ as electron  

acceptor

d. Favorable reaction, helps  

drive that other reaction

e. Gives you intermediate

f. Releasing CO2

g. If we get complete oxidation, we give 2 CO2, first from this reaction

h. Can’t oxidize carbon without rearranging carboxyl group

i. First real harvest of energy

E. Alpha-Ketoglutarate Dehydrogenae – Fig. 21.1

a. Second energy harvest, favorable reaction, the enzyme is just like the PDH complex  except without the regulatory subunits

b. Bottoms 3 carbons essentially pyruvate

c. Works same way as if it were pyruvate

d. E2 is very similar to E2 from pyruvate  

dehydrogenase complex (PDHC)

e. Except accepts 4 carbon fragment, releasing CO2

f. Transfers remaining carbons onto CoA

g. E3 from AKGDH is the same because you just want  

to get electrons from enzyme to NADH

h. Complex is basically identical to pyruvate  

dehydrogenase complex, E1 and E2 are different because substrate and there is no  covalent regulation

i. Favorable reaction, so again are in situation where we’re helping to drive forward this  reaction

F. Citric Acid Cycle Slide (again)

a. Both carboxylic acid carbons are from oxaloacetate

b. The fact that we have enzymes that create and recognize prochiral molecules allows us  to know where the carbons came from

c. Will walk through how we figured that out, later

G. Succinyl-CoA Synthetase – Fig. 21.22

a. Named for reverse reaction, not making succinyl CoA, we’re using it

b. Enzyme puts us in a position where we can use the nergy from the thioester, transiently  binding, displaces CoA with Pi

c. Pi soluble in mitochondria

d. Phosphoester hydrolysis is favorable

e. Replace one good  

leaving group with  

another good  

leaving group

f. Second step allows  

succinyl phosphate  

to interact with  

active site on  

enzyme and active  

site displaces  

succinyl group  

giving us a  

phosphate linked  

enzyme

g. Now on histidine side  

chain

h. Succinate released  

free

i. Ultimately enzyme  

with phosphate on it  

can bind a nucleotide  

and GTP is equivalent  

to making as ATo

j. Used tioester to make “ATP” is just counting ATPS

k. Have acquired succinate

H. Succinate Dehydrogenase – Page 811

a. This step is slightly unfavorable but not so bad that it won’t happen b. FAD operating at nomrla redox potential

c. Hangs out in mitochondria

d. Can use for energy later

e. Because of the double  

bond, molecule is more  

or less flat (across  

carbons), understand  

that we no longer can  

tell the difference  

between the lil dudes,  

which one was from  

Acetyl CoA and which  

one used to be  

oxaloacetate

f. Have acquired energy

g. Have mixed up the  

carbons

h. Succinate to fumarate

i. Fumarate is always a trans double bond

j. The enzyme is able to specifically abstract the electrons and protons from opposite  faces in a way that you always get the trans product

k. The trans aspect is one of the things you want to keep in mind for structures**** I. Fumerase – Fig. 21.24

a. Can add water across double  

bond

b. Diagram is in textbook – had  

good experimental evidence  

that there were two separate  

pathways. Nice example of  

trying to figure out  

mechanism

c. Outcome is malate

d. Operates ~at equilibirum with  

fumerase

e. Fumerate is converted to  

malate with hydroxyl group  

and hydrogen

f. Very close to oxaloacetate

J. Malate Dehydrogenase – Page 813

a. (S)-Malate to Oxaloacetate

b. Looking at the oxidation of the carbon

c. Produce NADH

d. Actually what’s happening is that the NADH  

is carrying 2 electrons and a proton

e. Oxaloacetate is not typically present in high  

concentrations

f. The way in which the cycle is able to  

continue is that citrate synthase is a very  

favorable reactions because you are able to  

hydrolyze off the CoA, and the fact that its  

so favorable that even in the case where the previous reaction is unfavorable is driven  by this reaction being so favorable

g. Also, the coupling, even though out of order, these enzymes are associated, not  covalently bound, so the fact that one is right before the other makes sense. Channeling  small amount of oxaloacetate right into next reaction

K. Citric Acid Cycle Summary

a. All is net energy recovery from the  

oxidation reactions

b. Actually getting a lot of yield from  

energy

c. Book says a few words about what  

that means in terms of ATP, she’s not  

going to do that at this point bc it’s a  

little more complicated than  

absolute #’s so we’ll talk about it  

later

d. For now, just know yields of reduced electron carriers and GTP & ATP for each portion  of the scheme

L. Discussion on Isotopes

a. Q: Remember the prochiral discussion? Where would the triangle and star be on a ketoglutarate?

i. Star is on CH3’s on pyruvate and ACoA and the triangles are on the carbons next  to those

ii. On citrate the star is on the carbon 1 down from the triangle (Fig. 21.1)

iii. A: The triangle is on the top carbon, the star is right underneath.

b. Q: What about on oxaloacetate?

i. A: Reaction run in a pool of molecules. In fumarate, can’t tell difference  

between 2 carboxylic acid carbons. Triangle is split between the two locations  

(top and bottom). The star is on either of the two middle carbons.

c. If we take labeled oxaloacetate and go back into the cycle again, are we going to release  the two triangle carbons are CO2? Yes. Will be released in next cycle. What happens  then to those carbons as well?

d. Isotope chasing:

i. First time around w/ newly added ACoA, released as free carbon dioxide

e. Starred carbon from a-ketoglutarate is released as CO2 etc. The CO2 released in the step  before that is also from oxaloacetate

f. Should help you learn the TCA cycle better too and what is actually happening Regulation – Fig. 21.25

A. Overall perspective: intracellular considerations, no significant  

hormonal control

B. A-ketoglutarate dehydrogenase is essentially the same E3 as  

pyruvate dehydrogenase that a-ketoglutarate dehydrogenase,  

reaction at bottom left is similarly regulated  

C. High concentrations of calcium activate

D. High levels of NADH negatively regulate isocitrate  

dehydrogenase

E. ATP also negatively regulates isocitrate dehydrogenase

F. Look at diagram to find negative regulation

G. NADH build up ultimately turns off not just pyruvate dehydrogenase and turning on lactate  dehydrogenase, but also you will get complete back up of TCA cycle

Amphibolic Nature – Fig. 21.26

A. Make or eliminate Citric acid cycle  

intermediates

B. Just said excess lactate makes way to  

liver and in liver is converted to glucose

C. In order to make that happen, actually  

need TCA intermediate and in a week or  

so will get to glucose synthesis and the  

pyruvates needed to make glucose, its  

ready to go from lactate to pyruvate but  

its not easy to go backwards

a. Process happens b/c of 4  

carbons fragments to go back to  

get glucose and we get to 4  

carbon fragments from  

pyruvate, so temp addition of  

carboxylic acid carbon

b. This particular reaction, if using  

TCA intermediates to make  

glucose, will require us to make  

new TCA intermediates

D. There are other reasons why you would  

need other TCA cycle intermediates

a. The other thing that should be obvious is that there are AA all over the place (in green) – in black means more than one can come this way

b. Basically, when we consume protein we have way more AA than we need

c. Carbons get broken down and used in fat for storage or (sssomething else). The vast  majority of the AA get converted into TCA cycle intermediates on their way to fat  synthesis

d. Worth recognizing the fact that a lot of input in the TCA cycle is through AA E. Can also use the TCA intermediates to reform AA

a. Last use is the use of citrate for fatty acids and cholesterol

b. Basically what is going on is that citrate is used for taking the ACoA that’s produced from  AA breakdown, glucose breakdown etc., attaching it on so its part of citrate, shove into  cytoplasm (in mitochondria) and in cytoplasm the two components of ACoA get  reunited

c. The 4 carbon part of the citrate gets shoved back into cytoplasm and gets reused d. Citrate is essentially a carrier to get ACoA into cytoplasm where __ is produced F. Constantly in flux, a significant thing it’s doing is giving us energy from foods we consume G. Also heavily involved in some other activities

a. What happens when we have low TCA cycle intermediates and have a need to run the  TCA at a more rapid rate?

H. Oxaloacetate From Pyruvate – Fig. cont

a. Anaplerotic

b. If you have a lot of ACoA, you’re either carrying out a  

lot of activity but you don’t have a enough TCA  

intermediates

c. Its possible that what you need to do is store some  

ACoA, but need to attach it to citrate to shove it into  

cytoplasm

d. So high levels of ACoA shows we don’t have enough  

of the intermediates

e. So we has pyruvate carboxylase, activated by high  

levels of ACoA, provides a way to make oxaloacetate  

from pyruvate and ATP and CO2

f. Use of CO2 in this reaction is essentially temporary, transiently using the CO2 so that we  have more intermediates but that specific carbon gets released at CO2 in TCA cycle  activity so it’s considered a temporary use of CO2

g. This particular reaction is crucial to allow the TCA cycle to function at optimal levels  without always having lots of intermediates that aren’t otherwise necessary

h. Turns out also that b/c of production of oxaloacetate, can use that as first step in  production of glucose, can take pyruvate (liver in conditions where it has lactate), can  easily be converted into pyruvate, just saw how pyruvate can be turned into  

oxaloacetate, oxaloacetate can favorably be turned into malate, and then that can be  turned into glucose ~~ woo provides building block that can be used for the synthesis of  glucose

i. This reaction and its control turn out to be a crucial mechanism

j. Make a point here that we’ve just walked through 2 most intense cycles that will be on  first test. These are the 2 BIG pathways. Start thinking about best way to have all of that  in your head. ☹

Glycogen and Signaling

Section Goals:

∙ For students to develop an understanding of the  

importance of carbohydrate metabolism in the  

body and the mechanisms by which carbohydrate  

storage is regulated

∙ Another goal is to use the hormonal control systems  

involved in glycogen metabolism to illustrate  

general aspects of hormone function

Glycogen Metabolism – Fig. 18.1

∙ Grey ends are non-reducing, addition and removal  

of glucose happens at the non-reducing ends

∙ There is one reducing end where the whole things  

starts, turns out this end is tied up in a protein so  

you can’t oxidize this end

∙ Similar to structure of starch except that glycogen has lots of branch points and starch does not ∙ Each purple circle is a glucose with two chains growing off of it

∙ When you’re either adding glucose or taking them off when needed, have lots of places where you  can remove glucose residues

∙ Good for bursts of energy b/c readily available for rapid activity

Why store glycogen?

∙ Speed

∙ Rapid source of energy

∙ Anaerobic metabolism Muscles that  

store glycogen can carry out a  

significant amount of glycolytic activity  

without the availability of oxygen

∙ Blood glucose-brain function  

∙ Brains need glucose to function and if  

we were dependent on just the  

amount of glucose in our bloodstream  

we would have to eat constantly,  

neurons use a huge amount of glucose so need to have glucose available

∙ A lot of the ATP made from glycolytic pathway is used to maintain ion concentrations across the  membranes of the neurons (signal firing) so bodies work really hard to maintain ∙ ~Constant amount of glucose in bloodstream, easier with a store of it in the form of glycogen.  ∙ Maintaining blood glucose levels happens in the liver and also sometimes some other tissues ∙ Osmotic pressure is low  

∙ Each individual molecule causes a small  

amount of osmotic pressure, glycogen  

reduces osmotic pressure b/c its one  

molecule  

Glycogen Structure – Fig. 18.1

∙ All a-1,4 and at branch point a-1,6

∙ The one position is linked with either 1,4 or  

1,6 which is why there are so few reducing  

ends

A. Glycogen Degradation – Fig. 18.2

a. Carried out by glycogen phosphorylase  

i. Does so in a way that preserves some of the energy  

that is there

ii. Has a complex structure

iii. There is a piece of glycogen that binds to maintain  

the association of the glycogen with the enzyme  

(helps with speed)

iv. Contains active site region that has a cofactor

v. Contains allosteric regions important in modifying  

system

b. Debranching Enzyme

i. Plays role in breakdown of branches to help you get to second tier end c. Phosphoglucomutase

i. Required so that phosphate ends up at 6 position of glucose, critical for  beginning of glycolytic pathway

d. Glycogen phosphorylase – Fig. 18.3

i. Has an active site with a  

cofactor based on vitamin B6

ii. Enzyme binds a nonreducing  

end in active site and a Pi as a  

substitute for the a-1,4  

linkage so you have the one  

position linked to a Pi

iii. Glycogen ends up 1 residue  

shorter

iv. Product released is glucose-1-

phosphate – not the  

intermediate for glycolytic  

pathway

e. Phosphoglucomutase – Fig. 18.4

i. Moves phosphate  

from 6 to 1 position

ii. Functions at  

equilibrium

iii. If you’re degrading  

glycogen, there is a  

constant production  

of G1P and if you  

have a lot of G6P it  

will convert it to G1P

iv. Think of as an  

intermediate in the  

pathway of either synthesis or degradation

v. Enzyme works by having phosphate stuck to it all the time – so a lot like mutase  that shifts locations of phosphate

vi. Now have it in G6P form

f. Debranching enzyme – Fig. 18.5

i. Has 2 separate functions

ii. GPhase is a big enzyme so can’t get into tight  

branches

iii. Debranching enzyme takes a chunk of “limit  

branch” and attaches it onto the basic chain  

that’s closest

iv. Energetically favorable

v. Can also hydrolyze off the a-1,6 linkages, does so because a-1,6 linkages are less  favorable when you hydrolyze them  

vi. dG for a-1,4 is more favorable so it’s not favorable to transfer a-1,6 onto end of  chain so is hydrolyzed as free glucose

g. Majority of glucose residues are released as free glucose, then has to be  phosphorylated, can be sent out to bloodstream directly

B. Glycogen Synthesis

a. Glycogenin (start a new glycogen)

i. People used to think glycogen phosphorylase worked backwards

ii. Disease of glycogen metabolism where glycogen phosphorylase doesn’t work  yet these people have normal glycogen – basically have trouble with rapid  

activity b/c can’t breakdown glycogen

iii. Led to another enzyme – glycogenin (not the one they discovered in response to  disease)

b. Phosphoglucomutase Glucose-6- phosphate -> Glucose-1-phosphate

i. Need or can’t get G1P

c. UDP-glucose Pyrophorylase

d. Glycogen Synthase

i. The enzyme that was discovered in response to metabolic disease

ii. Adds glucose residues onto glycogen

e. Branching Enzyme

i. Makes branches

f. Glycogenin – Fig. 18.8

i. Catalyzes same reaction as glycogen synthase

ii. Depends on UDP glucose

g. UDP-Glucose pyrophosphorylase – Fig. 18.6

i. Have UDPGPP

ii. Have UTP as nucleotide providing energy  

source

iii. Just like GTP situation from before (TCA),  

this is equivalent to ATP so UT P provides  

energy

iv. Linking phosphates b/w G1P and UTP and  

releasing UDP-Glucose (only has 2  

phosphate groups)

v. The Pi in vivo is immediately degraded  

which makes the reverse reaction  

irreversible/impossible

vi. Now committed! Favorable reaction  

because use up phosphoanhydride bond

h. Glycogen synthase – Fig. 18.7

i. UDP-Glucose is an activated sugar – provides the energy, glycogenic or glycogen  synthase, either way is transferred through enzyme into a-1,4 linkage and 2  phosphate residues end up with UDP

ii. Always couple the  

addition of  

phosphate onto  

glucose (hexokinase  

or glucokinase) with  

this reaction then  

get ADP and UDP or  

to think about total  

number of free  

phosphate released  

from the scheme,  

can do by going  

back to previous  

reaction where we  

released Pi

iii. Net use of ADP is 2

iv. Ended up recovering ADP or UDP

v. For each glucose added onto glycogen, have used essentially 2 ATP (1 ATP and 1 UTP)

i. Branching enzyme – Fig. 18.9

i. Makes branches

ii. Segment has to be reasonably  

large but not 2 large (6 long is  

averageish)

iii. More branches you have, the  

more glucose can be released  

very rapidly

j. Energy differences

i. Glucose to pyruvate gives 2  

ATP

ii. Glycogen to pyruvate, short  

term net 3 ATP, but more  

energy is used in synthesizing glycogen when energy is available

iii. Short term 3 comes from breaking down giving G1P, already phoshphorylated  aka don’t need hexokinase and don’t need ATP for that first step

iv. Has significant advantages b/c that’s 1.5x as much energy

v. If you’re gonna release glucose from liver it needs to be free glucose k. How can we control so many processes happening in cytoplasm?

i. Ex: If you eat a lot of breakfast before physics, some glucose is being stored as  glycogen, if you haven’t eaten since last night, body is  

still maintaining blood glucose level for brain so it is  

degrading glycogen made last night. Controlled

through specific enzymes Glycogen phosphorylase and  

synthase

C. Allosteric regulation – Fig. 18.10

a. Scheme tells us a little about the regulation, not really about the signaling b. A lot of things are put into this figure

c. Allosteric changes between active form and inactive form

i. “R form” is active form

ii. “T form” is inactive form

iii. Typically have just these 2  

conformations

iv. In the case of GPhase, also  

have the ability to make  

covalent modifications to  

enzyme that can change it  

activity

v. Phosphorylase kinase  

phosphorylates the enzyme

vi. There is serine in the  

enzyme that gets  

phosphorylated by ^ and  

dephosphorylated by phosphoprotein phosphatase

vii. The modified form of an enzyme that is enzyme is usually referred to as a small  “a” as in “phosphorylase a”, unmodified and less activated is “b” as in  

“phosphorylase b”

viii. Glucose can bind to phosphorylase (GPhase) and puses it to inactive form even  though its phosphorylated

ix. Acts as a control pathway

x. On nonmodified version of enzyme, can further push active conformation to  inactive conformation if you have G6P or ATP, if you have high levels of AMP,  can push inactive to active, releasing more glucose from glycogen

xi. So have enzyme whose activity can be modified by a wide variety of inputs xii. Phosphorylase kinase is very regulated

d. T vs. R Conformations – Fig. 18.11

i. N terminus of T state is  

down in the back but in  

R state is up and pulled  

away, making it more  

accessible

ii. Green loop in T state is  

peptide backbone of  

protein and is blocking  

access to active site

iii. R state does not have  

green loop in the way so  

active site is much more  

accessible

iv. AMP is an allosteric activator, bound to R state

v. If ATP is bound, extra phosphates can cause problems and push it back to T  state

e. Signal Cascade – Fig. 18.12

i. Representative of a  

number of control  

systems, she has named  

the enzymes we care  

about in our pathway

ii. Have covalent  

modifications of an  

enzyme that polay a role  

in activity changes

iii. Fairly straightforward  

cyclic scheme

iv. Phosphorylated is more  

active w/ tiny “a”

v. How much active enzyme you have depends on the kinase that phosphorylates  it as well as the phosphatase that inactivates it, each has a control mechanism vi. At the bottom they’ve written out the two reactions you have to consider when  thinking about whether the enzyme you’re looking at is going to be  

predominantly active or inactive (depending on phosphorylation event) vii. Even more complicated b/c getting both those enzymes in their active forms is a  whole other additional issue

viii. Can have multiple cyclic cascades that just adds one more level of regulation f. Bicyclic Cascade – Fig. 18.13

i. A bicyclic cascade forming another level of complication

g. Overall control of glycogen, muscle – Fig. 18.14

i. Glycogen is only half the photo and to the side is active form and there is  another letter designation, “o” = original, “m” = modified

ii. On the same level of glycogen phosphorylase there is glycogen synthase

iii. GSynthase exists in  

both and active  

and inactive form,  

regulated through  

same cascade as  

GPhosphorylase  

****buttttt for  

glycogen synthase,  

it is more active  

when it is not  

phosphorylated**** Original form is active

iv. Think about this from a logical perspective – situation in your muscles v. EX: ran down hill and exerted muscles. Had to turn on glycogen phosphorylase  Want only one to be on while the other is off.  

vi. Phosphorylase kinase is in charge of phosphorylating  

Glycogen phosphorylase

vii. In its original form it is less active, phosphorylation is active

viii. Multiple steps go into this enzyme being  

activated/deactivated

ix. The alpha and the beta subunits can both be phosphorylated,  

essentially helps remove an inhibitory segment that blocks  

the active site

x. The gamma subunit of glycogen phosphorylase can be  

completely removed and it will still function – delta subunit is  

also an inhibitory subunit, works through sensing calcium  

1. Ex: Running downhill, know muscles increased level  

of calcium. Need to remove inhibitory subunits and have the other part  controlled by calcium and others by phosphorylation. Maximal activity  comes from when all is activated

xi. Can have partial activation by hormonal control etc.

xii. Glycogen synthase has a control mechanism that is more complicated than that  of the phosphorylase mostly because if you follow yellow/orange activation  lines, you’ll see only phosphorylase kinase activates glycogen phosphorylase,  but a bunch of other things can activate glycogen synthase (like protein kinase  A)

xiii. The bottom half of the slide has purple lines that represent dephosphorylation xiv. Protein kinase A at the top is the enzyme that is the easiest to think about in  terms of being a direct link to hormones. Other names: PKA

xv. Enzyme controlled by levels of cyclic AMP in the cell

xvi. Exists in an inactive state and an active state. Phosphorylated in both  conformations so is controlled by the binding of cyclic AMP which removes  inhibitory subunit from catalytic subunit so you get the active enzyme and the  regulatory subunits bound up with cyclic AMP

h. Overall control (cont).

i. Phosphoprotein phosphatase controls the dephosphorylation of the enzymes  from before

ii. In its phosphorylated form, phosphoprotein phosphatase inhibitor controlled  phosphoprotein  

phosphatase

iii. There is modulation  

of the phosphatase  

inhibitor by  

phosphorylation

iv. So, in a hormonal  

situation where you  

want to have  

glycogen  

degradation, one of  

the things that’s important is the inhibitor active, making phosphatase inactive,  with a net result that glycogen phosphorylase stays in its active state longer,  resulting in more degradation of glycogen

v. Control scheme is not the whole story

vi. Another way to control phosphoprotein phosphatase is its association with  glycogen – not included in slide

i. PKA structure – Fig. 18.16

i. Important that we see that  

there is the inhibitory part by  

itself that has 2 cyclic AMPs  

that changes the  

conformation

ii. There is a phosphorylation site  

on the catalytic site of the  

enzyme

iii. Cyclic AMP activates the  

enzyme

j. Back to scheme from before

i. Advantage to the bicyclic thing  

is that you can get amplification of a signal

ii. Each active phosphorylase kinase can active a couple molecules of glycogen  phosphorylase which can then use glycogen

iii. EX: Ten molecules active one enzyme, who  

activate 10 molecules themselves, now  

have 100 glycogen phosphorylase  

activated. A lot at once is better than a  

little trickle

k. Calmodulin – Fig. 18.17 and 18.18

i. Protein that has several helices and a helix  

in between the two

l. Ca2+-CaM activation of Kinases – Fig. 18.20

i. Calmodulin site drawn at the bottom

ii. Need to open up catalytic site

iii. Have a covalent regulatory domain that needs to  

be moved

iv. Little green phone thing is the calmodulin, when  

calcium binds to it, it associates with regulatory  

domain and causes a conformational change that  

removes it from the active site, allowing for  

substrate to come in

v. Phone looks funky now kinda like a music note

vi. There are many kinases regulated through the  

availability of calcium, both involved in regulation  

of myecin activity in the case of glycogen  

phosphorylase in terms of metabolism in muscles

vii. This scheme lets us control the activity of  

phosphorylase kinase in part through calcium calmodulin

viii. Need both this calcium system and the phosphorylation of both alpha and beta  subunits to activate the phosphorylase kinase, optimal activation of glycogen  phosphorylase and inactivation of glycogen synthase to make energy

m. Hormones on muscle – Fig. 18.22

i. Diagram helps you  

understand the second  

aspect of its regulation

ii. Oval represents  

glycogen

iii. Subunit GM  

(m=muscle) and a  

subunits of PP1c  

associated with  

glycogen

iv. Role of subunit is to put  

the phosphatase where it needs to be to function, associating with glycogen so  it can have activity

v. When its associated with the singly phosphorylated it is most active, with two  phosphate groups it is inactive, block its association, phosphatase is released  and it can now bind to soluble inhibitor which now further controls the enzyme

vi. To make the system work, none of the binding interactions and the subunit are  permanent, they are on/off

vii. Mostly have association in terms of double phosphate

viii. It’s pretty easy to understand why we would want the event to happen (double  phosphorylation) because we might not want it running all the time

ix. Two phosphates means enzyme is not localized in an active way and can be  further stopped by its association with an inhibitor, don’t want to keep  removing phosphates if you really need glycogen degradation

x. Muscles are controlling system partially by phosphorylation of enzymes from  before and partially by blocking activity of this phosphatase so that the  phosphate groups stay on

xi. EX: suppose you were in UC eating a parfait, level of  

blood glucose high. Leads to insulin being released.  

Insulin causes a single phosphorylation event on GM1  

which means any available PP1 gloms onto GM and is  

associated with glycogen  

and can remove a phosphate and start process of turning  

system around.

xii. ^Part of the reason why this works well is that in a resting  

state, the most active version of the phosphatase is the  

one associated with a singly phosphorylated GM subunit. Also works b/c of  diagram with green and red boxes (regulation slides) where she told us the  majority of the activation was from phosphorylated but small molecules help as  well.

xiii. Free glucose can force T state of molecule before. By pushing glycogen  phosphorylase into T form, phosphate group is much more available and can be  taken off more easily by PP1

xiv. The role of the small molecules in controlling enzyme activity is so that you can  regulate activity, molecule is more likely to be inactive b/c phosphate group can  be taken more easily

xv. Goal is to manage sugar so we can function

n. Hormones and the liver

i. Low blood glucose effects on metabolism

ii. Low blood glucose cause pancreatic alpha cells to secrete glucagon iii. Glucagon receptors are analogous to epinephrine receptors and activate a G  protein that in turn activates adenylate cyclase

iv. [cAMP] rises activating protein kinase A – endpoint of several steps here v. PKA activates phosphorylase kinase which, in turn, activates glycogen  phosphorylase

vi. G1P is converted to G6P and hydrolyzed to free glucose

vii. Glucose moves through a permease in the membrane to raise blood glucose viii. Glucose is synthesized – next section.

o. Exercise and stress – Fig. 18.27b

i. Turns out liver cells have a second receptor for  

epinephrine, so in the case of fight or flight there  

is also a way to increase calcium levels

ii. Liver has system that moves G6P into the ER and  

makes free glucose

iii. Liver is responsible for keeping free glucose in  

blood  

p. High glucose vs. glucose use – Fig. 18.23

i. Glucose is stored as glycogen

ii. Glucose is degraded and fatty acids are made(F-2,6-

P is made to activate glycolysis)

q. Hormone Receptor interactions – Fig. 19.1a and 19.3a

i. Pancreas makes insulin and glucagon, different  

cells in charge

ii. Secreted, end up in bloodstream and different  

tissues

iii. Lots of tissues respond to these hormones,  

depending, way they respond or don’t is  

whether they have a receptor for the particular  

ligand or hormone in question

iv. At the bottom is a graph from text to remind us  

that hormone binding to a receptor can be thought of the same way as a  substrate bound to an enzyme

v. Response curve creates different levels of response within cell depending on  affinity of cell for hormone

r. G protein coupled receptors – Fig. 19.14

i. Cartoon belies  

complexity – part of a  

whole large family

ii. Turns out glucagon  

receptor that signals low  

blood glucose and the  

epinephrine fight or  

flight hormone work  

through an identical  

mechanism that results  

in the synthesis of cAMP

iii. They have 7 transmembrane segments

iv. When the right hormone is there in good quantity and binds to receptor,  receptor associates with g proteins

v. Association with G protein with the receptor causes a conformational change  with G protein as well, results in GDP being lost

vi. GDP inactivates the G-protein, becomes GTP

vii. ATP comes in with the orange protein and they come together to make cAMP +  PPi

viii. Phosphorylation of glycogen synthase and the other one

ix. To turn off, 1. Hormone disassociates 2. Adenylate cyclases are allosteric  modulators of the G protein itself. G proteins have ability to bind GTP and also  to hydrolyze GTP. Association with adenylate cyclase, there are a bunch, kinda  like a timer and then the GTP with become hydrolyzed and this reaction stops bc  the GDP form does not associate with adenylate cyclase.  

x. Need to keep sending hormone and having receptors on surface

xi. Gives an idea about what’s happening on surface

s. Signal System – Fig. 19.17

i. Have G protein that  

actually looks more  

accurate, a b gamma  

subunits, b and gamma  

subunits are not very  

relevant but play lots of  

roles in other situations

ii. GDP is lost and GTP binds  

rather than a phosphate  

being added

iii. Include changes in gene  

expression which are  

primarily controlled by  

phosphorylation through  

protein kinase

t. Other G protein systems – Fig. 19.54

i. In liver, receptor is also  

available to respond to  

epinephrine so you can get  

optimal regulation of  

glycogen phosphorylase

ii. Mechanism is fairly similar. In  

response to receptor, get  

hydrolysis of a head group of  

a phospholipid that contains  

something bisphosphate,  

produces phosphate free  

thing. Trigger to allow  

calcium transport into cytoplasm. Don’t need to know these details but should  be aware that calcium increases can be induced by several mechanisms u. High blood glucose and the liver

i. Pancreatic beta cells secrete insulin

ii. Insulin binds to its receptors on liver cells  

and other cells are sensitive (variety of cells,  

need to have receptor)

iii. Liver is responsive to insulin just not with  

glucose transport chains

v. Receptor Tyrosine kinases – Fig. 19.25

i. Cytoplasmic domain has enzymatic  

activity

ii. Insulin receptor is the one that looks like  

a spaceship

iii. Insulin binding activates kinase on the  

cytoplasmic side

w. Two RTK Models – Fig. 19.27

i. Have to form a dimer to send a signal

ii. 2 kinase domains come together upon ligand binding,  

phosphorylate each other

iii. All of the enzymes function to add phospjate grops to  

specific tyrosine groups

x. Insulin receptor – Fig. 19.67

i. pY – phosphorylated tyrosine residues

ii. Phosphotyrosines are binding spots,  

depends not on it being phosphorylated but  

also the specific AA around the tyrosine that  

are critical for docking

iii. For insulin receptor, most critical piece is  

IRS proteins

iv. IRS – insulin receptor substrate - proteins serve as a scaffold/an adaptor. When  they associate with phos. receptor, they get phos themselves, pY’s  

v. Don’t need to know all the things that go on in figure

vi. In muscle and in adipose/fat tissue, one outcome of insulin binding is glucose  transport. One critical player is protein kinase C. Also have to have some  

modification of cytoskeleton

vii. Glycogen synthesis - muscles and liver – when there slots of blood glucose, one  response is synthesis of glycogen to lower blood glucose levels (like after a  

donut). Glycogen synthase can be phos and turned off by lots of enzymes, of or  them is GSK3beta which gets turned off by insulin binding to receptor at top

y. MAPK – Fig. 19.40

i. Don’t learn diagram

ii. Primary mechanism by which insulin can  

cause changes in protein synthesis

iii. Multiple steps

iv. Also involved in cell growth

v. Occurs in cytosol  

vi. Pathway that can get turned on in abnormal  

responses like to a cancer or something

vii. One of several related paths

z. PKC and other enzymes- Fig. 19.60

i. Several steps involved in activation

ii. Have a calcium interaction site that’s critical

iii. Have a site that associates with lipids

Glucose Uptake – Fig. 20.11

A. Skeletal muscle and the liver

a. In response to the active insulin receptor, more  

glucose permease is moved to the membrane

b. When glucose concentrations are high (jelly donut),  

you get a rise in insulin, and insulin stimulates  

movement of vesicles from inside of cell to  

membrane of cell

c. This protein is referred to as GLUT4 and vesicle  

transport to cell membrane results in GLUT4 being  

on the membrane and stays there as a response of  

signaling

d. GLUT4 is a permease, moves glucose in am energetically neutral way from higher  concentration to lower concentration

e. Glucose gets phosphorylated and it now trapped and can’t get out

f. Way from body to uptake glucose when there is a lot of glucose in bloodstream g. In liver, also have glucose transporters, do not have GLUT4 (only insulin stimulated  GLUT), have similar transporter (GLUT2, and also have GLUT1 in other cells), but it never  “hides’ as vesicles, is always on cell surface

h. Liver will also take up glucose when high concentration in bloodstream, glucose goes  into both tissue types, don’t want transporter moved in liver is because when blood  glucose is low, liver is shoveling glucose back into bloodstream so if you moved  permease from the membrane you’d have no way to shove it back out

B. How does this work? – Fig. 20.9 and 20.10

a. Glucose binding,  

conformational change,  

phosphorylation

b. Multipass transmembrane  

protein

c. Several parts are charged so  

charged residues are then  

most likely involved in  

providing a hydrophilic  

opening so that the  

glucose/or water  

d. can actually associate (rest is hydrophobic)

C. Glucose transport systems vary – Fig. 20.17 and 20.7b

a. Transporter depends on energy input

b. Because do want to take in  

glucose in the wrong direction  

of a concentration gradient if  

its from intestines – want it  

taken up for body reasons

c. Energy is in the form of a  

sodium gradient

d. Symport system with glucose  

coming in because sodium is  

coming in along its  

concentration gradient, energy  

comes in from making sodium gradient

e. Other ones are ways of bringing in are uniport, symport, and  

antiport

f. Necessity for insulin in terms of moving GLUT4 to membrane

D. Insulin Receptor – Fig. 19.67

a. Modification of proteins involved in cytoskeletal movement –

don’t need to know details

b. Bottom line is insulin is important for GLUT4

c. Liver glucokinase concentration rises in response to PKB – glucokinase helps store  excess glucose in liver glycogen

E. High glucose (cont.) – Fig. 18.22

a. Phosphatase activity is turned on and phosphates are removed from the enzymes. Note  that the phosphatase in liver is actually associated with phosphorylase rather than  glycogen itself

b. In liver, the targeting  

subunits for PP1 are  

associated with  

phosphorylase itself and  

most effectively with  

the T form

c. There are more  

molecules of  

phosphorylase than  

there are of phosphatase

d. To get phosphatase activity, need to have several things happening in the liver. Need to  have phosphorylase in T form – small molecules (glucose) to push phosphorylase back  into inactive conformation

e. When that happens, can have serine residues and PP1 hydrolyzing phosphate off of that  serine which make sit modified unstable T form

f. Lots of phosphorylates around in comparison, have most of phosphorylase in T form,  pushed by glucose concentrations – when that happens the enzymes is now able to take  phosphates off of glycogen synthase

g. In the liver, crucial to have high glucose availability and this response system that allows  you to turn on glycogen synthesis after you’ve turned off glycogen degradation F. Insulin Summary

a. Receptor tyrosine kinase format  

b. Binding of IRS1 and other signaling molecules to phosphotyrosines  

c. MAPK pathway activation  

d. PI3K/PKB/PDK1 and PKC activation  

e. GLUT4 regulation in muscle and adipose  

f. Glucokinase expression in liver  

g. Phosphatase activity increases

h. (If she asks you to write out a control pathway – maybe not insulin – but more the idea  of what you might you if you were to talk about regulation so maybe glycogen  metabolism under certain conditions, list, story, etc. A list like this would be  appropriate)

i. (should you read the book? Book is useful for a lot of things but if you haven’t been  reading it the parts you should read are the ones that can explain complicated ideas like  in regulation. She will not ask about material in book that we haven’t gone over) G. Discussion

a. Consider a fit individual who has recently eaten

b. Would you expect glycogen to be synthesized in either muscles or liver under these  conditions?

i. Guess: I think maybe the muscles because they need to build up a store  

whereas the liver doesn’t have the same urgency bc its always around

ii. Answer: Both tissues would be making glycogen, how much depends on how  much sugar

c. Which enzymes would be active? Liver glycogen phosphorylase?

i. Guess: Yes in muscle cells bc need to keep glucose in the cell

ii. Answer: NO. Glycogen phosphorylase degrades glycogen so it wouldn’t be  active

d. Muscle hexokinase?

i. Guess: would be active to add a phosphate group to keep the glucose in the cell ii. Answer: YES. Phosphorylates hexoses or glucose in particular in this case. Need  G6P so need to go back and trace to hexokinase, so it would be active. Enzyme  is not committed to glycolytic pathway

e. Liver pyruvate carboxylase?

i. Answer: NO. Converts pyruvate to oxaloacetate. If you need more TCA cycle  intermediates. Not active bc have lots of energy and don’t need to make glucose  rn

f. Liver PP1?

i. Answer: YES. Takes phosphate off of enzyme phosphorylase. Takes phosphate  off of inhibitor. Glycogen synthase. PP1 takes phosphates off of different  

proteins. Will be functioning, don’t want glycogen phosphorylase active, so we  take the phosphate off. Taking phosphate group off of glycogen synthase which  activates it

H. Diabetes is a defect in insulin function

a. Type I, Insulin-dependent diabetes is typically juvenile

onset and due to damage to the pancreatic beta cells,  

these individuals respond to insulin

i. Damage to cells that synthesize insulin. People  

that need to take insulin and manage their levels.  

Those individuals are responsive to insulin, so if  

you’re at the proper levels, it can be fairly easy to  

manage. Autoimmune disease, so it’s the immune  

system that attacks the beta cells

b. Type II, Noninsulin-dependent Diabetes is typically late  

onset and more prevalent in obese individuals and those  

with a genetic predisposition. Elevated levels of free fatty  

acids can contribute to insulin resistance which may  

develop up to 20 years before NIDDM.

i. If you catch it early, you can treat it if you can do  

dietary changes. Farther into the disease is harder  

to do.

I. Mechanism of free fatty acid effect – Fig. 27.10

a. Phosphorylation of serine and threonine residues.

b. Tyrosine getting activated has downstream effects.

c. Can get phosphates on serine and  

threonine that make IRS 1 less  

susceptible to activation, less  

downstream, less GLUT4 to  

membrane, less glucose uptake - net  

result is difficulty in regulating blood  

glucose

d. Muscle uptake of glucose from  

bloodstream is super crucial, liver  

can’t do it all, important to reduce  

blood glucose levels

e. When were less responsive to insulin  

we have more glucose in blood  

stream which is bad. Attaches to  

proteins, causes vascular diseases, neuropathy (loss of feeling), damage to blood vessels  in eyes and kidneys.

f. There are other drugs that treat noninsulin diabetes

J. Leptin (information about fat)

a. Tells body how much fat is stored (in adipocytes)

b. Has effects on brain, in most individuals if fat levels  

are too high it stops having an effect, happens in brain  

and in peripheral tissues where it can be burned

c. In obese individuals, does not work as well in the brain  

but still work in peripheral tissues

d. Information is best identified in chapter 27 which his  

books attempt to pull tissues stuff into one chapter,  

good place for review at end of semester.  

e. Signaling mechanism discussed in chapter 19, don’t learn details

K. Leptin uses Jak/State Pathway – Fig. 19.48

a. Receptor system that has  

kinases associated with it  

but the receptor itself ifs not  

a kinase

b. Depends on  

phosphotyrosine  

recruitment which is kinda  

similar to insulin

c. Proteins recruited are  

transcription factors that  

move to nucleus if activated  

and alter transcription

L. Thyroid hormone – Fig. 19.4

a. Look at slide

b. Control metabolic activity

c. Does so through receptor – intriguing bc made by  

starting w very large protein – thyroglobulin –

iodinated on tyrosine residues

d. What you get is the side chain of one tyrosine linked  

to another one w three or four iodines

e. After proteolysis, end up with aromatics with iodine  

on them

f. People used to use iodized salt

g. T3 and T4, depending on how many iodines are on it

h. Binds to a transcriptor associated with DNA, inside nucleus

i. Receptor is a transcription factor, group that directly modulates transcription Carbohydrate Synthesis

Section Goals

∙ Develop and understanding of the reactions involved in glucose synthesis in response to low blood  glucose and the coordinate regulation of gluconeogenesis with glycolysis and glycogen metabolism ∙ Gain an appreciation etc

Gluconeogenesis – Fig. 23.1

∙ Liver using lactate as a starting point in anaerobic  

conditions

∙ Subset of muscle fibers have very little in way of  

mitochondria which are muscles fibers used for  

short term bursts of activity. Chickens that fly  

minimally, muscles are used for short bursts,  

muscles function primarily through anaerobic  

metabolism, few mitochondria.  

∙ The more muscles fibers you have that are  

specialized for fast burst activity, the more lactate  

you produce

∙ Lactate from the muscles at the top of diagram

∙ Converted to pyruvate in liver

∙ In green, a large number of AA (lysine and leucine  

not included) can give you carbon skeletons they  

can use to make glucose. Protein turnover. Longer it  

has been or few carbohydrates, a lot of what  

happens is the degradation of AA

∙ When glycogen levels get low. Enough glycogen  

sores for 12 hours. By time of 12 hours, definitely degrading protein. If you had some peanut butter  (high protein low sugar) AA could be used for this process. Could be dietary protein or muscle stored  protein

∙ Mostly the opposite of glycolytic pathway

Overview of Gluconeogenesis – Fig. 23.8

∙ Deal with three enzymes where dG is too high

∙ Called bypass reactions

A. Pyruvate Kinase Bypass – Fig. 23.2

a. Pyruvate carboxylase turns pyruvate  

into oxaloacetate, adds carbon  

dioxide

b. Take off CO2 when making glucose,  

PEPCK, requires GTP as an energy  

source, get PEP

c. Reverse pyruvate kinase step

B. Pyruvate Carboxylase – Fig. 23.4

a. Has a cofactor – biotin,  

micronutrient found in diet,  

but your gut also makes it a  

lot

b. Biotin is the transient site  

to white CO2 is added and  

is where you need the ATP

c. Phosphate donated to CO2  

then CO2 transferred onto  

biotin

d. Biotin on long arm, lysine  

residue, very flexible, can  

move from one location to  

another

e. Moved again to make  

oxaloacetate

C. Model for pyruvate carboxylase – Fig. 23.5a and 23.5d

a. Only way enzyme  

functions is if it  

exists in multimeric  

form, so reaction  

sites are close to  

each other

b. First step on one monomer and the second on a  

different polypeptide that’s in the same complex so  

two sites can be close enough together

D. PEP transport – Fig. 23.7

a. In mitochondria, where oxaloacetate is

b. Diagram is designed to tell you we have to transport  

components, ether the PEP or the oxaloacetate which  

does not have a mitochondrial carrier (don’t learn  

routes now, but will come back to it later, transport is  

crucial for the movement of electrons)

E. PFK and hexokinase bypass – Fig. 23.8

a. Cant reverse phoshofruktokinase and make ATP

b. Simply hydrolyze off the phosphate group, reasonably  

favorably, clearly if we were just doing the two, we  

would just keep burning up ATP

c. Only happening in tissue that makes glucose, enzyme doesn’t exist in muscle

F. Glycolysis vs. Gluconeogenesis

G. How Does Blood Glucose Regulate Gluconeogenesis?

a. Control of PFK/FBPase

b. Control of Pyruvate Kinase/Pyruvate Carboxylase

H. Regulation – Table 23.1

a. These are the controls that occur in the liver

I. PEPCK

a. When the liver is responding to glucagon, transcription factors (CREB in particular) are  phosphorylated.

b. This binds to CRE (cAMP response element) in the control region  

of the PEPCK gene resulting in more enzyme production.

c. This helps to increase glucose synthesis.

Biosynthesis of Other Carbohydrates (Start of Exam 2 Material)

A. Glyoxylate – Fig. 23.11

a. Plants can start with fat  

and make carbohydrate,  

we cannot do that What  

they need is only 2  

enzymes that we don’t  

have – isocitrate lyase and  

malate synthase

b. In glyoxisome, can take  

two 2 carbon fragments  

and TCA intermediate and  

if you have TCA  

intermediates, you can  

make glucose

c. Can also degrade to succinate and glyoxylate

d. Isocitrate lyase carries out this reaction

e. Glyoxylate can be combined by malate synthase with ACoA to get malate which is a TCA  cycle intermediate

f. *Not how seeds get enough sugar to grow

g. Mechanisms by which plants can create sugars from fat which we cannot do B. Oligosaccharides  

a. Structures that are attached to proteins

b. N-linked oligosaccharides in ER and Golgi – by far the  

most commons of the bunch

c. O-linked in Golgi

d. GPI-linked in ER – not talk about these at all

e. Fig. 23.16

1. Summarizes oligosaccharide synthesis in ER

2. Lots of details

3. Oligo’s are not initially added directly to the  

protein

4. Started by linking sugars one by one to a  

lipid that hangs out in ER membrane and has  

an interesting property that it can be flipped  

back and forth across ER membrane

5. Sugar transport mechanism dolichol  

phosphate has to be activated (by a phosphate)

6. There are a whole host of nucleotides used in activation

7. Turns out that when the nucleotide itself is linked onto something to serve as an  activator --not ATP

8. Each sugar gets activated like activation of glucose from glycogen synthesis 9. Every sugar gets activated in a similar mechanism – using a nucleotide- sugar w  phosphate group on it

10. Nucleoside triphosphate group, linking, releasing inorganic phosphate

11. Look at diagram and can figure it out

12. Gets linked to very specific sequences in a protein

i. Usually polypeptides that are in synthesis

ii. Have dolichol pyrophosphate

iii. Another flippase that moves it back out

iv. Can start process over again

13. Basic mechanism used for all N-linked oligosaccharides

14. Happening in ER

f. Fig. 23.18

1. Figure summarizes relationship  

with n-linked to proper protein  

folding

2. Main things to point out are  

that the extra glucose residues  

are a mechanism that can help  

to ensure that the proper  

folding happens

3. Down to a single sugar,  

proteins can be processed by  

one of CRT or CNX –

chaperone’s help to get folding correct and can pull off  

extra glucose --if folding isn’t correct, glucose can be  

re-added at that point and can start folding again

4. Should have general sense that proper folding  

happens in the ER

5. Once proper folding happens there is a removal of one  

of the mannose residues

6. The glycoprotein now holding n-linked is transferred  

by vesicles to golgi

7. In golgi get additional processing

g. N-Linked ER in Golgi – Fig. 23.20

1. Activated sugars UDP sugars primarily but not only

2. At bottom there is a CMP sugar, etc.

3. Only know that final processing  

happens with N-linked sugars in Golgi

Pentose Phosphate Pathway 

A. One additional way to oxidize glucose

B. 2 crucial functions

C. Three roles for Pathway

a. Synthesis of NADPH

i. Had talked about AND as accepting  

electrons in glycolytic pathway or TCA

ii. NADPH is the version of the electron carrier crucial for biosynthesis, can’t make  lipids, cholesterol, or steroid hormones without it

iii. Concentrations of NADPH are like that of ATP and are kept high

b. Synthesis of Ribose-5-phosphate

i. Need because of nucleotides

ii. Want to make DNA, replicate DNA etc. need ribose

c. Interconversion of odd sugars

i. We’ll see how that interconversion works

D. NADPH synthesis – Fig. 23.26

a. Oxidizes G6P, G6P dehydrogenase,  

gets 6-phosphoglucono thingy  

lactone

b. Controlled by concentrations of  

NADP+ and NADPH

c. Oxidized glucose can be opened  

more easily than just glucose

d. 6-phospho-glucono-lactonase  

intentionally opens glucose

e. Second oxidation enzyme – 6-

phosphogluconate dehydrogenase  

makes Ru5P

f. Each glucose that goes into  

pathway in this format, get 1 CO2, 2 NADPH, and one  

5 carbon sugar

g. Regulation dependent on NADPH and NADP+

h. Starting with G6P

i. Have two oxidation steps that together result in a 5  

carbon sugar and release of CO2

j. Oxidative stage is stage 1

k. Look at stage 2 the interconversion of 5 carbon  

sugars so if primary goal is to make R5P we can do  

that through this pathway

E. Ribose-5-Phosphate and Xylulose-5-phosphate synthesis –

Fig. 23.29

a. Lots of ribose needed for nucleotide biosynthesis

b. If ratios of NADPH for Ru5P needed are balanced,  

could be only thing going on in this pathway (isn’t,  

but could be)

c. If the balance of NADPH and ribose 5 phosphate is satisfied by the reactions we’ve looked  at, that’s the primary way in which this pathway works

d. Often the case that you need more NADPH of R5P and don’t need the balance this  pathway gives you

e. To deal with issue and for other convenience purposes…

F. Fig. 23.26

a. Have whole other set of  

additional reactions that allow  

us to interconvert a variety of  

different sugars

b. Use two different enzymes so  

just to lay out where we are,  

on the right hand side have  

R5P and X5P below it, created  

on previous slide

c. Making 5 carbon sugars allows  

us to recover all of our carbons  

and glycolytic intermediates

d. First reaction is catalyzed by  

transketolase – takes 2  

carbons from a kept sugar (X5P in this case) these are shaded in green and the carbons  are green

e. 2 carbon fragment dumped onto the top of R5P giving S7P and remaining 3 carbons  from X are G3P

f. Transketolase does this 2 carbon thing

g. There is an enzyme called transaldolase which is similar to aldolase in glycolytic pathway h. Moves 3 carbon fragments so top 3 carbons of S7P (shaded in blue) dumped onto 3  carbon fragment G3P now have F6P which we now what we can do with – glycolytic or  back to G6P easily

i. Get 4 carbon fragment

j. Then come back and get 1 more molecule of X5P and we use transketolase – takes 2  carbons same as first but now dumped onto 4 carbon sugar not 5

k. Get a second F6P and a G3P

l. End of process get 2 F6P and a G3P having starting with 3 5 carbons sugars --one R two X m. Useful for if you don’t need much R5P but need NADPH

n. Could be true if making lots of fat for storage (talk about in 2 weeks) could be true for  other tissues that have a huge need for reducing power for biosynthesis but aren’t  proliferating much themselves

Electron Transport Chain and Oxidative Phosphorylation

∙ Made NADH in glycolytic pathway

∙ Made NADH and FADH2 in CAC

∙ Sources of energy

A. Where did the electrons  

come from? – Fig. 22.1

B.

a. When we break  

down fat and  

amino acids we  

get a lot of NADH

b. This is the  

mechanism or  

scheme  

regardless of  

what kind of  

energy source we  

have

C. Harvesting the Energy

a. We’re not going to walk through all the  

math in class

b. We have a situation where if you  

completely oxidize glucose and do  

calculations for the energetics of the  

complete oxidation of glucose and  

reduction of oxygen we end up with a  

whopping amount of energy  

c. Have seen so far that we’ve been able  

to carry out this oxidation

d. Converting chemical energy to electrical energy

e. All energy in form of NADH and a little FADH2 as far as we’ve talked about f. Conversion of electrical energy into chemiosmotic energy

i. ETC basically uses a flow of electrons to set up a proton gradient across inner  mitochondrial membrane

g. Then we’ll look at the conversion of chemiosmotic energy back into chemical energy in  the form of energy

i. Reduced electron carriers making ATP

D. Electron Carrier Types – Fig. 22.22

a. Have briefly seen structures before

b. Here are several different hemes that  

have this basic protoporforin structure  

and various side chains

c. Iron is there, which can switch between 2+  

and 3+ which is where the electron gets  

carried

d. Each heme group ends up being buried  

within a protein

e. DON’T LEARN STRUCTURES

f. Can exist at a variety of redox potentials

E. Monitoring Carriers – Fig. 22.20

a. Monitoring absorption

b. Take a population of  

cytochromes and do spectral  

analysis which allows you to  

see several bands

c. Levels change depending on  

talking about oxidized or  

reduced form

d. Using spectroscopy to get this  

information

e. When looking at entire mitochondria membranes the diagrams get more complicated F. FMN – Fog. 22.17

a. Flavin Mono Nucleotide

b. Part of the FAD+ molecule

i. TCA cycle

ii. Pyruvate dehydrogenase

c. One of the interesting things about  

FAD+ and FMN, Flavin molecule can  

carry one or two electrons but NAD+  

and NADH needs two electrons

d. Here can get a semiquinone  

intermediate which is a free radical  

but is possible to get

i. Turns out for electron  

transport sometimes only need to move one electron

ii. Flavin is therefore useful then

iii. DON’T LEARN STRUCTURES

G. Fe-S clusters in Proteins – Fig. 22.15

a. Iron-Sulfur clusters exist within proteins

b. Majority of structure types we’ll find

c. Another version where one iron is missing  

d. Get coordination sometimes to cysteine

e. Sometimes to sulfur

f. Sometimes to histidine

g. Not showing specific charge bc clusters are managed by protein in which they exist so  binding site within protein for specific iron sulfur cluster  

i. Single goes between +1 and +2 state

ii. But others can be at many levels

iii. Ex: 2Fe-2S can start out as +3+3 and take one electron, can go to +3+2 or could  be in most oxidized state +3+2 and then be +2+2

h. Combination between protein in which they exist and the overall redox situation and  the specific range of oxidation states for any particular iron atom is

i. The iron sulfur clusters share the charge over all of them – not localized, spread H. Coenzyme Q – Fig. 22.17

a. Have seen carriers free in  

solution, some bound to  

proteins

b. Coenzyme Q (also called  

ubiquinone or abbreviated  

CoQ) is a small molecule, fairly,  

that is very hydrophobic so it is  

soluble in membranes

c. If you look at it, the electron  

carrier part of it is actually a  

small aromatic ring with a  

number of sidechains and what  

makes it soluble is a set of  

isoprene groups

i. The image shows you one with an N at the end of it

ii. Kind of chain that exists in dolichol sugar carriers

iii. This lonng hydrophobic tail is the reason why this is a very lipid soluble molecule d. Carrier very useful because it can ferry electrons between large protein complexes that aren’t very mobile

e. Can carry either one or two electrons at a time

i. While cytochromes only carry one electron

Location is Everything 

A. Location – Fig. 22.2

a. Sitting on membranes

b. Most of activity is within  

inner membrane of the  

mitochondria

c. Inner membrane quite  

impermeable while outer is  

very permeable  

(impermeable to large  

proteins but small ones can even get through sometimes)

d. Large shape range and small size range

B. Mitochondrial structure – Fig. 22.4

a. Rat liver mitochondrion

b. Inner membrane is in a yellow

color

c. Outer membrane in a red color

d. Cristae (folds) are in green

i. While they are continuous with inner  

mitochondrial membrane, have isolated  

regions

ii. Would have to have a fair amount of  

diffusion in an awkward setting to get  

significant material between the inner  

and outer mitochondrial membranes  

and all the way to this contiguous but somewhat isolated space within the  cristae

e. ETC sets up proton gradient – might be somewhat different in regular mitochondria vs.  rat liver mitochondria tho

C. Getting NADH There the Easy Way – Fig. 22.8

a. Critical to get the NADH that’s produced in the cytoplasm into the mitochondria b. NADH produced in aerobic glycolysis has to get in** important

c. In TCA cycle it’s already in

d. 2 different main shuttles

e. Easy way (tissue specific) - know DHAP  

from glycolysis and 3PG we know as  

the backbone for lipids and triacyl

glycerols (TAGs)

i. One way we can process NADH  

is starting with 3PG DH

ii. Electrons get used to reduce  

DHAP to 3PG

iii. 3PG shows up at inner  

membrane surface

iv. (Outer membrane has pores in  

it – basically anything that isn’t  

a protein can come into the  

mitochondria, ballpark 10K)

v. 3PG shows up at inner membrane surface

vi. Enzyme has FAD+ bound, collects electrons from 3PG, now have FADH2 vii. Carbons get re-oxidized to DHAP so it can be a shuttle

viii. Chain functions in inner mitochondrial membrane

ix. Exists in bees, also works in our brains and some other  

tissues as well

x. Simple, effective, takes advantage of glycolytic  

intermediates (DHAP)

xi. Electrons in FADH2 are at a lower redox potential than in NADH so there’s not  as much energy there, however, used very effectively in specific locations

D. NADH Conservation and Movement – Fig. 22.27

a. This is the second, more complicated way

b. Happens in skeletal muscle and in a number of other tissues

c. Mechanism depends on having amino acid skeletons (carbon skeletons) around d. Takes advantage of several transporters in inner mitochondrial membrane e. Oxaloacetate cannot get through inner mitochondrial membrane, need malate f. Situation allows us to transfer electrons not in NADH but in malate

g. Malate DH in TCA cycle, malate is more favorable than oxidized oxaloacetate – more  energetically favorable

h. Malate made in cytoplasm

i. Have a transporter which is an antiporter, transports malate for alpha ketoglutarate  (AKG – at the bottom of the circle) (TCA intermediate)

j. Once in mitochondria, able to recover malate to oxaloacetate and CAC can drive this – happens by citrate synthase being so favorable

k. NADH in mitochondrial matrix, now have oxaloacetate which can’t get out – so the rest  of the scheme is try to get it back out now

l. Since it would be worthless to go back to malate, now convert the carbons to aspartic  acid – amino transferases interconvert alpha keto acids and alpha amino acids so we get  aspartic acids because we use an amino group from glutamate and the glutamate  becomes AKG

m. Aspartic acid goes through second transporter (AA transporter) which exchanges  glutamic and aspartic acid across inner mitochondrial membrane – neutral exchange n. End up with aspartic acid in cytoplasm and glutamic acid we needed in matrix o. Equivalent aminotransferase in the cytoplasm so can regain oxaloacetate and our  glutamate

p. The 5 carbon molecule (AKG) is just there for the ride or provide the amino group or the  anti-port molecule

q. 4 carbon fragments (oxaloacetate, aspartic acid, malate) are what end up carrying the  electrons

r. Have both molecules cycling together with a net movement of NADH across the  membrane

s. Also move a proton because you start with both of them on one side and end with both  on the other side

t. So we are using proton gradient as part of the energy to drive this

u. Gets us NADH and not FADH2 across the membranes

v. Tissues that use it can save energy bc they have NADH, stronger reducing potential  molecule

w. LEARN WHICHEVER ONE MAKES MORE SENSE TO YOU -- DON’T LEARN BOTH The Electron Transport Chain – The Order of the Carriers 

A. Order – Fig. 22.9

a. Can start talking about ETC  

itself and this diagram does  

a really nice job of lining up  

various players that are  

involved in the ETC and  

gives a sense of the redox  

differences

b. When trying to figure out if  

electron movement is  

favorable, take value for  

electron acceptor and  

subtract from that the  

value of the donor and if its  

positive its favorable (-dG)

c. Ex: NADH is the most  

negative one we’re looking  

at -0.315

i. Oxygen, ultimate  

acceptor, +.815

ii. Subtract -.3 from .8 get a much more positive values

d. Instead of having a direct transfer of electrons from NADH to oxygen, have  intermediates that are set up in a way so that you can use that significant amount of  energy to make more than one ATP

e. There are three steps shown there redox potential is large enough where you could  synthesize an ATP – not actually true but is suggested on figure

f. Each complex gives a specific redox value for the reaction that is the electron transport  across the complex – redox values for specific players in complexes

g. Giant multiprotein structures

h. Succinate DH with FADH2 or transport system we just talked about with FADH2 show up  at same level of CoQ so show up at lower redox potential than NADH which is why you  get lower energy yield

i. Cytochrome c is a small protein intermediate that contains one heme c group i. Intermediate between complex III and IV

j. Biggest jump is at bottom and where we get biggest transfer of electrons onto oxygen to  give us water

k. Have tools that are able to block electron transport at specific complexes and we will  use those as how they help us understand how the ETC works

i. Each functions by blocking a particular complex function

ii. Blocks ATP synthesis from that transfer but bocks electron transfer at that  

function

2/24/2017

The Electron Transport Chain – the Order of Carriers (cont.) 

A. The Order (cont.)

a. Makes the assumption that the electrons are floating from carrier to carrier to carrier b. This kind of alignment/flow is what is actually happening

c. We’ll then talk about the complexes and how the e flow is moving protons across the  inner mitochondrial membrane

B. Carrier Order by O2 Use – Fig. 22.11

a. Similar alignment to the  

previous figure

b. Representation of the  

kind of experiment one  

can do to study electron  

transfer in the  

mitochondria (in a test  

tube)

i. Specialized test  

tube that has an  

oxygen sensing  

electrode in it

ii. Using a  

particular DH as  

a source of NADH as long as there is beta-hydroxybutyrate

c. Have mitochondria, supplying NADH, getting a decrease in the concentration of oxygen d. If they add some specific inhibitors, find that they can block that electron transfer e. Inhibitors function in a way that they can bypass the inhibitors if they add succinate - which we’ve talked about in TCA cycle

f. The succinate DH enzyme reaction results in the transfer of electrons to FADH2 i. Not subjected to same inhibition

ii. Can deplete amount of oxygen present

g. If we add an antibiotic that inhibits the function of the succinate DH and FAD+ system,  we stop the use of oxygen and electron transfer

h. Electron source is ascorbate/vitamin C and a small hydrophobic carrier that will interact  and transfer electrons to the electron transport scheme – will get additional use of  oxygen

i. If we add cyanide, all electron transfer terminates

j. So oxygen is the sink and we look at different ways to mess with it

C. What is a P/O Ratio? – Fig. 22.13

a. What this means in terms of coupling this to ATP synthesis

b. Graph in textbook has been updated but we use this one for the historical sense of how  this work was done initially where the expectation was that you could just relate oxygen  use to ATP production

c. P/O is essentially how much ATP is made from ADP per amount of oxygen used d. The assumptions were that you would have to have whole numbers – couldn’t have  partial relationship between oxygen and ATP use

e. Same sense you’d get from first figure we started out with when they had ATP directly  linked to the complexes

f. Trying to measure the oxygen use which we just talked about and the amount of ATP  production – running to the end of ADP in each case so you’re not making any more ATP g. In this experiment once you run out of ADP you stop the oxygen use

h. This tells you there is a significant coupling

i. If you look at the current, updated graph

i. When these  

experiments are done  

more carefully and get  

a better understanding,  

P/O ratios are not  

whole integers, closer  

to 2.5 and 1.5

ii. Tells you the coupling  

between these electron  

transfer schemes is  

different than direct  

interaction than a  

system that makes ATP  

and the electron transfer couple

iii. If you’re starting with NADH at 2.5 and succinate at 1.5 (succinate is not the only  place you could get electrons at the level of FADH2 – on Weds we talked about  the G3PDH system that was one mechanism by which you could transfer  

electrons into the mitochondria and there are other redox enzymes)

j. Looking at last step, there seems to be an integer value of approximately 1

D. Table 22.1 Redox Potentials

a. Not just the levels at which you’d donate  

electrons

b. Textbook has many of the individual electron  

transfer players within each complex and she  

chose not to put them on there, more of a  

sense of what we were just talking about

c. Between succinate and CoQ (intermediate  

electron carrier) there is very little difference  

which is why you don’t get enough energy out  

of this electron transfer to contribute to ATP  

synthesis

d. Includes actual table too – Table 22.1

i. Each iron sulfur complexes listed is a part of  

complex I

ii. Each heme b is part of complex III

iii. Basically separates them

iv. Have hemes and coppers which play a role in  

electron transfer

v. As you go down list, numbers go from  

negative to positive

vi. NADH is better electron donor

vii. O2 is best electron acceptor

viii. If you look at second level “complex II” there  

isn’t a whole lot of difference so can’t really  

contribute energy for ATP synthesis through  

complex II which she also explained in the  

simplified table above

ix. Also means you can’t transfer protons here bc  

the redox potential isn’t large enough

E. Electron Flow – Fig. 22.14

a. This is useful in terms of a big picture of what’s going on, not in terms of what’s actually  happening in the complexes

b. Each complex that has a large difference in redox potential has the ability to move  protons across the mitochondrial membrane and the proton differential is what is used  to drive ATP synthesis

c. Complex I spans the electron potential difference between NADH and CoQ d. Complex III spans the electron potential difference between CoQ and Cyto-C/oxygen e. Figure lays out some of the players, doesn’t tell you how electron or proton flow is being  accomplished

F. Complex I – Fig. 22.14, 22.18

a. This is misleading slide

b. Structural analysis of prokaryotic one

c. Has a huge transmembrane arm and  

a peripheral arm and they lay out the  

locations of these

d. Because this comes from a bacterial  

system it’s actually sitting in the  

cytoplasm and that’s what they’re  

showing you

e. If we were looking at the  

mitochondrial complex I we’d be  

looking at the matrix – facing matrix  

where NADH is

f. Name of the complex is actually NADH CoQ reductase

i. Using NADH to transfer electrons and reduce  

CoQ (give it electrons and protons)

g. Has, in general, different functionalities between arm in matrix and arm in inner  membrane

h. Transmembrane arm has the easiest color scheme to figure out

i. Transmembrane helices that are purple, blue cluster, green cluster, etc. each  cluster represents a protein

ii. Combined together to make the arm

iii. Have a structural relationship to proteins used to serve as antiporters –

exchange protons with sodium ions (not these, the ones that they’re similar too) iv. The idea that became popular when this first structure of a bacterial complex I  was solved was that the side chain parallel to the membrane was maybe a

mechanical arm that somehow altered the conformation of the transmembrane  segments that helped proton flow – not actually the case

i. What actually happens is that the electron transfer occurs in bunchy region and force is  communicated to transmembrane arm

i. Protons and electrons flow from the binding  

site of the NADH (at top in peripheral arm –

bunchy region) to an FMN

ii. Electrons can then flow to a series of sulfur  

complexes within the various protein

iii. The protons cannot bind to Fe-S complexes

iv. Protons that come in are separated from the  

electrons and essentially get dumped out

j. CoQ has a binding site at the bottom rightish and  

recovers protons from a different source – not  

necessarily the dumped ones - and gets electrons  

from Fe-S clusters

G. A Giant molecular Proton Pump – Structure and Mechanism of Respiratory Complex I a. Pic not included in slides

b. Shows you peripheral arm on the side  

c. Still based on bacterial system

d. Bacterial systems that have been characterized – are around 14-15, in humans there are  45

e. All extra subunits are involved in aspects of function including assembly and protection  from environment

f. Critical because there are a couple of diseases associated with complex I– some fatal g. Cartoon suggests a slightly different functionality of membrane arm but piece most  helpful to think about is that the transmembrane parts (most) have discontinuities in  the middle of the membrane

h. Now seems more likely that there is a link somewhat electronic but also have  mechanical changes between electron flow and the transmembrane segments of these  proteins so have more work to do to figure it out

i. Diagram suggest proton flow is happening through these transmembrane proteins so  the proton flow is being regulated by the electron transport but not by pulling this  parallel arm back and forth

j. Net result of this is transferring 4 protons across the scheme – one for each of the  locations – for each pair of electrons transferred from NADH

k. Basically for the whole movement of the electrons through this scheme, get flow of 4  protons across inner mitochondrial membrane, such that the protons accumulate on the  outside of the inner mitochondrial membrane

l. Can remember direction of proton flow is that the yellow jellybean diagram does it right H. Complex I Electron Movement – Fig. 22.19a/b

a. Expansive look at the  

peripheral arm

b. Shows you the series of  

iron sulfur clusters  

c. Can kinda see the FMN  

location at the top -

where the NADH binds  

d. The electrons transfer  

(and FMN can transfer  

one or two electrons at a  

time so get 2 electrons)

e. Then one transfers down,  

then the second transfers

f. Proton gained in Q is probably also occurring one at a time since you add electrons g. ^All happening in membrane

I. Complex II – Fig. 22.22

a. Several things you can see –

bound FAD+ in complex itself

b. Is succinate DH, the other  

enzymes that carry out similar  

FAD+ are similarly placed in inner  

mitochondrial membrane

c. In each case, while they have  

some transmembrane subunits,  

there is no mechanism to move  

protons so all that’s happening in  

complex 2 is that you are  

funneling electrons into the  

electron transfer scheme to get  

to CoQ – not collection of energy

d. dE value is very small – not enough to collect energy or  

make a proton gradient

e. So, enzyme essentially just gets the FADH2 electrons and  

moves them through a series of Fe-S clusters

f. There’s a coordinating heme group then they get moved  

to CoQ  

g. Now have this whole pile of electrons (most from NADH,  

some from succinate, some from the G3P DH, oxidation  

of fatty acids)

h. At this point we just end up with a collection of CoQ  

without adding to the proton gradient

J. Redox Loops – Fig. 22.30

a. This is how complex III functions, it is not a pump like complex I

b. Have electrons in Q – critical in  

getting electrons transferred to  

cytochrome C and also getting  

proton flow

c. When we talked about complex I we  

talked about a pump mechanism

d. Complex III works in a redox loop/Q  

cycle which depends on having 2  

types of electron carriers – one  

carries electrons and protons and  

one carries electrons – kinda saw it  

in complex I but it didn’t function  

this way  

e. Complex III – have a carrier that  

brings in both electrons an protons  

f. Delivers the electrons to an electron carrier (or maybe more than on) and dumps  protons on opposite side so other than a pumping mechanism, it’s a question of where  the protons come from and get dropped off

g. Picks them up on one side and dumps them on the other side

h. Electrons go to oxygen but it’s not the immediate receptor

i. Initially how people thought  

complex I worked too but it’s  

actually a pump

K. Complex III – Fig. 22.23

a. Dimer structure

b. Color scheme shows lots of proteins  

that come together

c. Have buried all the players in this  

structure

d. Can see two heme groups – one closer to inner membrane space side and one to matrix  side

e. Can see an iron sulfur cluster

f. Can see heme C1 – almost in inner membrane space

g. Can see docking site for cytochrome C – only one of them docked at a time h. Some protein that are on inner membrane space, some mostly transmembrane i. A lot of the proteins extend at least one (lilac) transmembrane segment through to  other side in matrix

j. The other thing they have in this diagram is an inhibitor – stigmatellin that helps us  figure out how scheme works

k. On the right are the redox potentials isolated

i. Cyto bH carries a single heme b

ii. bL carries another

iii. Rieske carries an Iron sulfur

iv. If you pull the players out they don’t give you  

100% in order redox values

v. CoQ for whole thing is pretty close to the Cyto  

b’s, cyto c is slightly more positive that the C1

vi. Rieske is more positive than you’d expect

vii. In vivo, redox potential is not exactly the .28  

shown here for isolated form

L. How Complex III works – Fig. 22.31

a. This is the first half – the top half – of the  

redox loop

b. Shows you a Coenzyme 2 from complex I  

or II

c. Reduced electron carriers soluble in  

membrane move to binding site close to  

inner membrane space, binds to where  

stigmatellin inhibitor was before

d. CoQ encounters situation where there  

are options for it to transfer various  

components

e. No place to put 2 at a time, only one electron carrier here

f. The closest electron acceptor is the Fe-S protein and the transfer  

of one electron there is quite favorable

g. Q has an interesting property – if you transfer one at a time the  

redox potentials are different

h. From QH2 to QH dot (radical) is much more positive so really only  

effective way to get rid of an electron is easily transferring electron  

to Rieske Fe-S protein

i. Protons get dumped into inner membrane space (out of  

mitochondria)

j. Rieske has conformational shift that moves the Fe-S protein away from the binding site k. Left with the radical Q with a negative charge and it’s in the vicinity so it can transfer an  electron to cytochrome and the second electron transfer potential is .03 which is similar  to cyto b’s

l. All close enough that you can transfer the electron

i. Shown going down as if there’s a big change but its pretty much similar

m. There’s a second Q binding site for first round

i. Second Q binding site contains a completely oxidized Q

n. Now were in the matrix side (orange) – picks up an electron so Q’s are equivalent just in  different locations, associated with the protein, not just free in solution

o. Electron in Rieske moves to cytochrome C 1 and differential is not favorable i. Now have electron in C1 that’s at the inner membrane space side of the inner  mitochondrial membrane and at the docking location that cytochrome c comes  to bind (mobile, water soluble)

p. Cytochrome C docks very quickly and leaves very quickly – relative affinity, gives  orientation and stuff  

q. Electron transfers favorably from C1 into cytochrome c and Cyto C diffuses away r. Left at end of first cycle with radical CoQ, way this get resolved is cycle 2 M. Complex III (Cont.)

a. Another reduced CoQ comes in exactly  

how the first did, gets to same reaction  

site, transfer electron to Fe-S protein,  

once it gets rid of electron moves back to  

original location

b. Whole process is identical to the first one^

c. This transfer is the same as well except at  

second Q binding site, see waiting for it is  

the free radical Q which is a better  

electron acceptor than oxidized Q

d. Picks up second electron (more favorable)  

and picks up protons from the matrix

e. What we’ve done is created a scenario  

where two Q’s have moved their protons  

across the inner membrane space – one  

has ended up in its oxidized form (namely from cycle 2) the one  

from cycle 1 is completely reduced again, picking up 2 protons  

from matrix and one electron from first cycle and one from  

second cycle

f. QH2 is free to come in and start all over again

g. From 2 cycles have net transfer of 2 e and 4 protons into the  

inner membrane space

h. There’s a little bit of motion (like in Rieske) but mostly  

functioning by redox loop in complex III rather than pump system  

found in complex I

N. Cytochrome b orientation – Fig. 22.23 a. What’s happening at inner  

membrane space is where we  

have cyto c picking up electrons  

to move one at a time

b. Can also see the binding site for  

Q that first comes in and the  

second Q binding site which is  

at the bottom of the protein  

(matrix side)

O. Cytochrome C – Fig. 8.42

a. Mobile electron carrier

b. Met it in Biochem I as a heme

carrying protein that has been  

used as an evolutionary clock

c. Have looked at its sequence bc its  

easily available and can study to  

figure out evolutionary  

relationships based on sequencing  

(more than 30 years!)

d. Freely soluble in inner membrane  

space

e. Able to move from Complex III and  

complex IV

P. Complex IV – Fig. 22.24

a. Can see in diagram a nice layout of it in  relation to membrane

b. Also a dimer

c. Has a docking site for cytochrome c in  inner membrane space

d. Doesn’t have as much matrix side  

structure as some of the other complexes e. Buried in each side are components  involved in electron transfer – can see a  copper center by the aqua balls on the  

top left

Q. Complex IV Electron Carriers – Fig. 22.25 a. Can see the copper and the specific  

protein side chains to which its  

associated

b. Have a redox potential for copper a

c. Can see heme groups at the bottom and  another copper part of the complex

The ETC 

A. Cytochrome C Oxidase – Fig. 22.24

B. Complex IV Electron Carriers – Fig. 22.25

a. Each carrier within complex 4  

can carry only electrons, no  

protons

b. Proton movement has to be  

through a pump mechanism

c. The copper complex is closest to  

where cytochrome c docks

d. Get an electron, shared between  

two coppers

e. Get copper 1 copper 1 but without the electron you just have a share 1 and 2 f. Electrons move one at a time from  

center

g. Move through a heme A and get  

shuttled into complex here where they  

get transferred to oxygen

h. Can’t transfer electrons one at a time  

trying to get them into oxygen it just  

doesn’t work

i. It has been studied and oxygen gets  

associated and gets reduced one  

electron at a time so there is a hypothetical pathway trying to tell you what’s going on j. Start with an oxidized center

C. Electron Transfer Steps – fig. 22.27

a. Rapidly followed by second electron and proton

b. Release molecule of water

c. Oxygen was  

associated/coordinated with  

the iron of heme A3 and it  

came from the previous round  

of the cycle

d. Have water this water release,  

iron 1 copper 2

e. Now can have new oxygen  

bound in coordinated with the  

heme and evidence suggests a  

fairly rapid reorganization of  

the structure of this and allows  

you to get an electron from the  

tyrosine, see where the proton moved from the oxygen that entered

f. Last electron needed to get structure coordinated correctly comes from the tyrosine

g. Add electron and release water

h. Have original atom of oxygen coordinated with water

i. Donated electrons so very oxidized

j. Copper now in original format

k. Can add electron and proton and get back to starting intermediate

l. Very quick cycle – pretty hard to catch all of this in process so some of it is best guess  and some of it is supported by spectroscopic measurements

m. Some protons are added to the system to become part of water

D. Complex IV Also Moves Protons – Fig. 22.23

a. Doesn’t tell us about how this causes  

a proton gradient

b. Before evidence, people guessed it  

moved protons with a pump

c. Have better evidence now for  

complex iv but there is more  

clarification

E. Bacteriorhodopsin – model – Fig. 22.34

a. Don’t learn details

b. Essentially depends on a gating  

system so protons can’t blow backwards across membrane

c. Partially due to the fact that the center of the  

pathway is gated in a resting state without light

d. Light activates retinal which is converted into a cis  

form – basically excited

e. Now have a rearrangement of the structure such  

that the proton that was up there can actually  

move to become associated with the aspartic acid

f. Structural change and now a chance for second  

proton to move to a pair of electrons on the  

nitrogen

g. Finally able to release the proton that had moved  

to the carboxylic acid carbon and have another  

proton come in and take the place of the one  

from the carboxylic acid and start over

h. Basic chemistry tells us we don’t just have free  

protons – are actually thinking of H3O+ (not H+ even though we write it this way – bc  we lazy)

F. Complex IV Channels – Fig. 22.35

a. Looks as if there are two separate channels in complex iv

b. One channel (K) we don’t have a lot of players but we probably have a water – have a  situation where we can imagine with the right water possibilities a way in which we can

get protons up to the copper b or the site where the  

oxygen is going to be present – no way we can align  

the channel with an exit on the outside so hypothesis  

is that this channel provides the protons for the  

water production

c. D channel – see 3 separate potential parts – trying to  

show you pathways that make sense based on what  

we know about structure – have potential  

connection all the way across the membrane so the  

evidence at this point is that through conformational  

changes that as the oxygen is being reduced ( e  

coming in) have changes that allows protons/push  

protons from binding site to binding site – end up  

with a situation where you can imagine a series of  

hops the protein can take from appropriate electron  

source to e source up to exit channel and where hypothetical exit channel is we have an  aspartic acid there for possibilities of a last step before it leaves

d. Another unclear thing is that in addition to asp 91 there are other potential side chains  to recruit protons

e. Best evidence is that as we move electrons from center – reducing oxygen – we make  some conformational changes we basically push protons through channel

f. 4 protons across membrane and were using protons to become part of the water – 4 for  the oxygen

g. Decrease of protons in the matrix as were moving the electrons from cytochrome c to  water as a final source

G. Super-complex I1III2IV1 – Fig. 22.28

a. Shows an additional picture in class??

b. Q binding site for complex 3 and complex 1, you can imagine  

a very simple transfer instead of a huge jumping

H. The Chemiosmotic Hypothesis – Fig. 22.29

a. We’ve talked about how we move protons

b. Can calculate what that means in terms of energy byu looking at pH difference between  the two locations can get a measurement of the membrane potential  

c. Talking about a single charge

d. Can calculate that for each proton, costs us ~21kJ/mol to move protons out e. We’re expending a lot of energy from ETC to make the process work and to push the  protons out

f. High proton concentration is a source of energy that we can use for other purposes Oxidative Phosphorylation – Fig. 22.36

∙ Can see matrix in here and what look like lollipops and they are a portion of the enzyme that  synthesizes ATP – the F1 particles

A. F1 Particles – Fig. 22.36

a. Are available through sonication – get vesicles where lollipops are sticking out b. Can dissociate with urea which doesn’t  

cleave bonds but changes interaction so  

F1 particles get dissociated and can be  

reconstituted so there’s no permanent  

change

B. ATP Synthase F1 structure – Fig. 22.38

a. Has a couple of proteins part of a stalk  

structure running through middle of  

globular structure

b. If you look top-down at the lollipop you  

can see alternating beta residues and  

alpha residues in the lilac color – 3 of  

those – and the way this is shown, it’s  

shown also with nucleotides and each

biding site – can see 5 – the catalytic part of this enzyme is in the beta subunits c. All three betas are the same and all three alphas are the same but sometimes the  structures change depending on where we are in the cycle

d. If you isolate the particle san give ATP, this hydrolyzes it

e. Tells you that the only way this is synthesizing ATP is if its associated with a part in the  membrane

f. Works in an energetically favorable way  

– hydrolyzing ATP

g. Isolating hydrolytic part hydrolyzes ATP

h. Catalytic part is within F1 structure

C. Top Down View – Fig. 22.39

a. Tunnel/channel part

b. Uses sodium ions so purple circles are  

sodium ions

c. Shows a series that span the membrane

d. Each has a spot to bind sodium  

ions/partially charged small molecules

D. ATP synthase – Fig. 22.41

a. Have cytoplasm and paraplasm – bacteria don’t have  

mitochondria

b. Has a peripheral arm (orange) – made up of both a  

protein that interacts with the c subunits and a long  

arm that spans from the membrane up over the top  

of the catalytic portion of the structure

c. Book goes back and forth between terminology for  

mitochondria system and bacterial system so only  

remember the parts that are the same – STALK and  

PERIPHERAL ARM and that’s good

E. Diagram of Action – Fig. 22.45, 22.46

a. Aspartic acid which is a potential binding site for a  

proton

b. What is believed to happen is the ring  

of c subunits 10-15 and the side  

protein associated in the membrane  

has a way in which the proton can get  

through from the matrix to the  

aspartic acid in one particular c  

subunit – which causes a ink, kink  

pushes rotator and moves a proton  

that’s gone all the way around to an  

exit site in same protein from which it  

can exit out of the complex to the  

inner membrane space

c. So every time a proton come sin it pushes the rotator – with a lot of protons ca move  rotator pretty quickly

d. Allowing protons from high concentration to associate with c proteins, kink, turn, turns  stalk, stalk doesn’t turn F1 portion, rubs against it

e. A and peripheral arm don’t move, stalk has to turn against this full structure held in  place by peripheral arm

f. Have looked at AA inside alpha and beta and it’s clear that there’s nothing holding them  together – lubricated

g. Causes conformational changes in alpha and beta subunits which causes ATP synthesis h. Gradient of protons that is creating this rotation, causes conformational changes  causing synthesis

F. Isolated ATP synthase can be turned on by a proton gradient – Self fig

a. Can mimic and create vesicles  

with ATP synthase and if you set  

up a proton gradient so that it is  

able to let protons enter c  

sections and come through  

synthase, actually see ATP  

synthesis

b. Can mimic without complex 1 3 4

c. Shows it’s the movement of  

protons driving ATP synthesis

d. Without gradient, there is no synthesis (right side of image)

G. dpH is Critical! – Fig. 22.46

a. For liver mitochondria with a pH  

difference of 0.75, DG = -21.5 kJ/mol  

to move the H+ in. About 3 H+ /ATP  

appear to be needed.

b. Trend negative bc favorable for  

protons to come in

c. 3 protons to synthesize an ATP

d. Can take a solution of mitochondria  

and add in a small hydrophobic  

proton carrier and will dissipate  

proton gradient

e. Get no ATP synthesis

f. People have shown it is not the membrane potential difference, it’s the pH difference H. ATP Synthase sites – Fig. 22.42

a. Alpha and beta subunits are the critical ones

b. Pointer is the stalk

c. Have three sites – open site, no binding

d. Loose site – can have initial binding of substrate

e. T site – only ATP can be here,  

conversion of loose site to T  

site is the catalytic force that  

drives the synthesis

f. Have energy in the form of  

rotation – pointer changes  

conformation, convert all  

sites tight becomes open,  

loose become tight, open  

becomes loose and allows  

binding  

g. Open site releases ATP that was synthesized (and water) now have additional energy we  can move back

h. Happens rapidly

i. Can cause conformational changes within the beta subunits that creates this continuous  cycle of synthesizing ATP as long as there is proton flow through motor

I. Three Sites are Coupled – Fig. 22.42

a. Conformation of the active sites  

in the ATP synthase is modulated  

by proton flow through the F0  

portion of the ATP synthase and  

how that drives the  

conformational changes and the  

changes drive the synthesis of  

ATP and if we don’t have it we  

just have it free in solution

b. Enzymes stabilize the highest  

energy state to which the  

intermediates have to get in the  

reaction and so clearly its forced in a direction here that we get ATP synthesis under  conditions where the proton gradient exists

J. An uncoupling proton channels – Fig. 22.47

a. In mitochondria present  

in a specific type of fat or  

adipose cells namely  

brown fat they have the  

ability to uncouple the  

proton channel  

essentially transfer  

protons across inner  

mitochondrial membrane  

similar to 24DNP  

(dangerous chemical that  

can kill you if you have

too much) but in a controlled way a number of organisms can do the same thing  effectively

b. Proton channel also referred to as an uncoupling protein and there are a couple of types  – present in brown fat, hibernating animals and human infants primarily in the upper  back region

c. As scientists have done more work there is a certain amount of brown fat in adults it  just not clumped

d. Deals with heat production by taking proton gradient and getting rid of it simply making  heat

e. Alternate way to allow protons across the inner mitochondrial membrane that is gated  by free fatty acids

f. Free fatty acid activate the thermogenin channel

g. High level of nucleotides can block the channel’s function

h. Common situation is if you have low energy and therefore high levels of ADP or GDP but  usually in fat you aren’t going to be looking at high levels of fat in cytoplasm in any case i. Free fatty acid release happen sonly under conditions where triacylglycerols are  hydrolyzed by enzyme triacylglycerol lipase

j. Only works in brown fat because of thermogenin(?)

K. Uncoupling in vivo – Fig. 22.47

a. Top half of figure

b. We’ve seen G  

protein coupled  

receptors

c. Didn’t bother to put  

G protein in pic but  

it’s there

d. G protein picks up  

GTP after activation  

interacts with  

adenylate cyclase

e. Catalytic subunits of  

protein kinase A is released wen 4 molecules of cyclic Amp interact with regulatory  subunit dimer

f. Etc etc Activated triacylglycerol lipase

g. Heat important in hibernating animals  

and infants who don’t have a lot of  

insulation

h. Control of enzyme is a general control

L. Integration of Control – Fig. 22.48

a. Regardless of when you ate you have to  

be making ATP

b. As long as you have oxygen you have a  

source for electrons to be deposited

and a generation of proton gradient and ATP synthesis

c. If ATP concentration gets super super high or no ADP or no oxygen and NADH builds up d. High concentrations of NADH inhibit citric acid cycle in very part where NADH is a  product (DH) and also at citrate synthase that controls the entry of acetyl-CoA into the  cycle

M. Control of ET and OX/Phos

a. NADH/NAD+

b. ATP/ADP

c. Cytochrome c reduced/oxidized – in ETC  

can control level of cytochrome C bc  

ultimate place of electrons is oxygen so if  

complex IV can’t put them anywhere the  

cytochrome C levels build up in reduced  

form  

d. Control of complex IV – related to above --integrates with previous slide N. Control (cont.):

a. Control of pyruvate dehydrogenase complex  

by TCA cycle activity and also control by  

NADH/NAD+ ratio

b. Some positive control by calcium and energy  

charge if you will so a higher ADP can push  

this forward

c. Very high ATP can also inhibit

d. All control is based on what is going on in cell  

in question and oxygen levels which isn’t just a  

local cell issue

O. Control (cont.)

a. If citrate gets high messes with PFK

b. When talking about liver and skeletal we  

didn’t talk about F26BP but not about  

control with oxygen

c. In brain and heart etc where you can’t have  

anaerobic run into situation where you can  

have high levels of AMP which activates a  

kinase (AMPK) which can phosphorylate a  

specific version of the enzyme that makes  

F26BP, a way to activate the glycolytic  

pathway even when there isn’t enough  

oxygen for the short term just to keep  

going

d. AMPK is taken advantage of in cancer cells

P. Energy Yield Discussion

a. How much ATP/pair of electrons?

b. NADH ATP/2.5