BIO 201 With Todd Hennessey Week Seven Notes
BIO 201 With Todd Hennessey Week Seven Notes BIO 201
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This 33 page Class Notes was uploaded by ChiWai Fan on Sunday March 13, 2016. The Class Notes belongs to BIO 201 at University at Buffalo taught by TODD HENNESSEY in Spring2015. Since its upload, it has received 101 views. For similar materials see CELL BIOLOGY in Biology at University at Buffalo.
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Date Created: 03/13/16
Cell Bio on Mar 7, 9, 11 (All images taken from Professor Todd Hennessey’s slides) There are metabolic and structural. We will talk about metabolic today. Metabolism Anabolism: Synthesis Catabolism: Breakdown Energy derived from catabolism (like the breakdown of sugars) can be used to drive anabolism (like the synthesis of sugars) The Circle of Life From another book: “Solar energy is the ultimate source of all biological energy.” We need the SUN in order to get the stuff we need for survival. Chloroplasts use energy from light to make carbohydrates and generate O 2 Mitochondria produce water and CO from c2rbohydrates and O 2 Glycolysis—metabolism. Let’s focus on animal cells Glycolysis Generates ATP, NADH and Pyruvate You take a 6-carbon sugar and modify it giving you phosphate-charged glucose then split it in half giving you two 3-carbon compounds which gives you two pyruvates. Thus glycolysis giving you 2 ATP and 2 NADH Can’t run glycolysis without ATP The enzymes of glycolysis—each enzyme is a different gene product Example of one enzyme Hexokinase uses ATP to phosphorylate Glucose; Hexokinase puts a phosphate on glucose; you are storing energy You get 2 moles of ATP, 2 moles of NADH and 2 moles of pyruvates The first steps of glycolysis These first three steps use ATP to charge up the sugar Each step is a different, specific enzyme This converts Glucose to Fructose 1,6-bisphosphate ATP is needed for glycolysis; imagine an investment of ATP, using ATP to make ATP Several more enzymes are involved in cleaving this sugar and modifying one of the products How to make it into pyruvate? This series of steps generates three types of high potential energy compounds: ATP, NADH, pyruvate. o 2 NADH o 4 ATP (we want lots of ATP so we use NADH to do this) o 2 pyruvate Each round of glycolysis yields TWO ATP because you need 2 ATP to start it. What can you do with pyruvate? Convert it to Acetyl CoA inside the mitochondria. Why? Because Acetyl CoA is what you want. To go into the Citric Acid cycle (aka Kreb’s cycle or TCA cycle) Summary so far: o Glycolysis yields 2 ATP, 2 NADH and 2 pyruvate o The 2 pyruvate go into the mitochondria o Inside the mitochondria, they are converted to 2 Acetyl CoA o What do we do with the Acetyl CoA? o The Acetyl CoA go into the citric Acid Cycle in the matrix (insoluble area) of the mitochondria Three big steps from Glucose to lots of ATP 1. Glycolysis in the cytoplasm (you get a little bit of ATP) 2. Citric acid cycle in the matrix (inside) of the mitochondria (you get none ATP here but it gives stuff to make ATP) 3. Oxidative phosphorylation across the mitochondria inner membrane A redox cycle inside the mitochondria Different names for the Citric acid cycle o Kreb’s cycle o Tricarboxylic acid cycle o TCA cycle What do you get out of this cycle? (FYI) Purpose of Kreb’s: Get NADH Acetyl CoA (2 carbon)42+4=6Lose a CarbonLose a Carbonback at four An example of one of the reactions Malate is getting oxidized. This happens in the matrix of mitochondria Reduced compound Oxidized form Other nutrients can feed into this process to provide energy Other products can be used for anabolism (synthesis) Glycolysis isn’t only doing one thing; it has a lot of reactants and products. There are several points for either activation or feedback inhibition We’re making more than enough ADP stimulates and ATP inhibits here ATP or NADH inhibit here If a lot of ADP but not a lot of ATP, you might want to stimulate it If a lot of ATP, we might want to tell the glycolysis to slow down (inhibit it) Regulation of cellular biochemical pathways What do you need NADH for? NADH is an electron donor + NADH + H+ + 1/2O 2 NAD + H O 2 o The oxidation of NADH by O y2elds -52.4 kcal/mol (to make ATP) o The hydrolysis of ATP yields -7.3 kcal/mol o Synthesis of ATP requires +7.3 kcal/mol ( Oxidized form= NAD+ Reduced form= NADH (Then make ATP from this) You’re getting electron and hydrogen ions from NADH Using the power of redox What do you get out of this? Energy to set up a proton gradient (like energy in a bucket) The Secret of Life: anything alive can do this Electrons (e-) and protons (H+) 1. Membrane e- transport is used to set up a H+ gradient as electrons are passed from NADH to O . 2 2. The energy of the H+ gradient is used to make ATP NADHenergy stored in proton gradientmakes ATP Redox—converting the forms of energy Mar 9, 2016 A reduced compound gives electron and an oxidized compound want electrons If you want more oxidized compound A, add more reduced compound A and oxidized compound B What would you need to do to get more Oxidized compound A? Add more Reduced compound A AND Add more Oxidized compound B Oxidation, Reduction and Electrons LEO the lion goes GER: In mitochondria, NADH is the first electron donor and oxygen (O ) is the final 2 electron acceptor. NADH is a reducing agent. It gets oxidized as it loses e-. Glycolysis in the cytoplasm and Electron Transport in the mitochondria The three parts: (Compartmentalized) Glycolysis in the cytoplasm Citric acid cycle in the matrix of mitochondria Electron transport across the inner membrane of mitochondria (You get most ATP out of this step) What do you get out of this? 32 ATP/molecule of glucose, H 2 and CO 2 Glycolysis does not give you any ATP; it is when glycolysis offers pyruvate to make acetyl CoA in the Kreb Cycle to generate NADH. Then we use NADH to make a lot more ATP. And Glycolysis operates by using ATP! Mitochondria Outer membrane: More permeable. Protein/lipid about 1:1 Inner membrane: Less permeable. Protein/lipid about 3:1. This is where electron transport happens Matrix: Soluble (aqueous) area inside the inner membrane (The pink stuff on the image below) Intermembrane Space: Aqueous area between the inner and outer membranes Mitochondrial electron transport and Oxidative phosphorylation What is the pmf? Proton Motive Force The pmf provides the electro-chemical energy to make ATP. It is dependent upon: A. Membrane potential. This is the electrical driving force (Vm) B. H+ gradient or proton gradient. This is the chemical driving force Our Rationalization (to simplify this): If the membrane potential is held constant, a change in the pmf is mainly controlled by changes in the H+ gradient. (Thus more influx) Watch for 3 steps: Step 1: Electron transport from NADH to O prod2ces energy (-ΔG) higher proton concentration in Intermembrane space than in matrix Step 2: That energy is used to set up the H+ gradient by pumping H+ against their electro-chemical gradient (+ΔG) from matrix to outer space Step 3: The energy from the H+ gradient (-ΔG from the pmf) is used to make ATP (+ΔG) Water Analogy to help understand Mitochondrial e- transport We’re not destroying energy, we are just changing its form 1. Pump water up into bucket. This takes energy (like e- transport) 2. The water in the bucket is potential energy (like the H+ gradient) (high proton concentration outside than inside so they want to come in) 3. Pour the bucket out into the pipe and the energy of falling water is like the pmf. It can be used to turn the turbine. 4. This generates electricity (like making ATP) The Oxidation of NADH and FADH in2the Respiratory Chain: Lets oxidize these! What do you start with? NADH—first electron donor (Complex 1) FADH 2 What do you get? Energy Water What can you do with this? Set up a proton gradient. Ubiquinone is a lipid. Cytochrome C is a peripheral protein sitting inside Intermembrane space, its low salt! Not all transport chains are proteins!! Are these endergonic or exergonic reactions? How can you tell? EXERGONIC because we are going from high energy to low free energy state which is negative delta G thus it is exergonic Mitochondrial electron transport Transport of electrons. Complex II is left out for simplicity UQ is ubiquinone which is a lipid. Final electron acceptor: OXYGEN. If no oxygen, no electron transports. If pH doesn’t change, this thing shuts down! The purpose is to get high concentration of protons outside and low concentration of protons inside. High protons mean low pH. So we want low pH outside and high pH inside. Oxidative Phosphorylation: The coupling of Electron Transport from NADH to O 2 and ATP synthesis 1. In cytoplasm: Glycolysis breaks down sugars into smaller pieces in the cytoplasm 2. In matrix: Those smaller pieces go into the TCA cycle inside the mitochondria, generating NADH 3. Within the inner membrane: Electrons are passed from NADH, through the electron transport chain, to oxygen (that’s why it is aerobic) 4. Across the inner membrane: H+ are pumped across to set up a H+ gradient 5. In matrix: The energy of the H+ gradient is used to make ATP 6. ATP is exported from the mitochondria to the cytoplasm. So is CO and 2 water as by-products What does mitochondrial electron transport produce? 1. ATP? No 2. A proton gradient across the inner membrane? Yes 3. Higher H+ inside or outside of the inner membrane? Outside 4. Why? To generate an inward proton motive force (pmf) to provide the energy for ATP synthesis in the matrix Three energy steps to link electron transport and H+ gradient to ATP synthesis Step 1: Electron transport from NADH to O p2oduces energy Step 2: That energy is used to set up the H+ gradient by pumping H+ against their electro-chemical gradient Step 3: The energy from the H+ gradient is used to make ATP How ATP Is Made The Focomplex is the H+ channel The F1complex makes the ATP, but it can’t do it by itself Why? The –ΔG from the pmf provides the energy to overcome the +ΔG for ATP synthesis 1. H+ diffuses through this channel 2. This subunit rotates 3. Rotation causes this subunit to change its 3D shape to expose the active site for ATP synthesis Some Mitochondrial poisons 1. Cyanide, CO (carbon monoxide): Inhibit e- transport to O . 2ith no final e- acceptor, e- transport backs up and stops. Blocks Step 2 2. Detergents: Break down H+ gradient. Blocks Step 2. 3. DNP (dinitrophenol): Uncouples the energy of e- transport from ATP synthesis so the energy is lost as heat (non-shivering thermogenesis). Blocks Step 3. DNP is a H+ shuttle DNP breaks down the H+ gradient This dissipates the pmf so there is not enough energy to make ATP Where does the energy go? HEAT Non-shivering thermogenesis This is used in babies and hibernating animals to generate heat March 13, 2016 For animal cells, oxygen is the final electron acceptor Bacteria don’t have mitochondria but they still make ATP Other electron acceptors instead of oxygen Bacteria don’t have mitochondria. How do they make ATP? It can be better for bacteria to live in acid because it can use the proton to make ATP! (proton gradient) Microbial fuel cell Waste from sewage treatment plants inElectricity out You take anode and collect with cathode, when waste from sewage plant comes in, electricity is created Four stages of energy generation by aerobic metabolism Four stages of energy generation by aerobic metabolism Stage 1: Glycolysis in the cytoplasm If 2 is present (aerobic), then: Stage 2: Pyruvate oxidation to Acetyl CoA in the mitochondria Stage 3: Citric acid cycle inside the mitochondria (gives NADH) Stage 4: Oxidative phosphorylation across the inner membrane of the mitochondria (gives ATP) What if there is no oxygen (anaerobic? Anaerobic energy production (no O present2 Aerobic and anaerobic | Oxygen and no oxygen There’s much more ATP made under aerobic conditions, which means when there’s oxygen!! Two main types of anaerobic fermentation processes When there’s no oxygen, then it is fermentation then there’s lactate or ethanol 1. Fermentation of pyruvate to lactate (as in muscle) 2. Fermentation of pyruvate to ethanol (as in yeast) This regenerates NAD+ for glycolysis and produces 2 ATP for each glucose molecule NAD+ is needed for glycolysis to make NADH. Pyruvate’s choices Glucose goes into pyruvate o If oxygen present, then you go to Acetyl CoA then Citric acid Cycle o If no oxygen, then you go to fermentation then either ethanol in yeast or lactate in muscle Anaerobic fermentation to lactate o This is often seen in muscle when the oxygen gets used up; we want to get more ATP but no oxygen present o Lactate dehydrogenase is reversible, so it can be used to regenerate pyruvate when oxygen returns o When there’s no oxygen, pyruvate makes lactate causing you 2 ATP giving you NAD+, if oxygen is present, it can reverse and make a bit more pyruvate o The purpose of fermentation is to make NAD+ to start glycolysis o Where does the NAD+ usually come from under aerobic conditions? o NAD+ helps start glycolysis. It will run out when they’re no oxygen o If electron transport is transferring electron to NADH, if there’s no oxygen there to accept it, everything backs up and STOPS. Leading to lots of NADH but no NAD+ o What does an anaerobic condition do to a cell? It stops the oxidation NADH to NAD+ o Under anaerobic conditions, we don’t get enough NAD+, we need NAD+ to make things like glycolysis This is muscle because lactate Key concept: Anaerobic is all about the NAD+ Glycolysis can’t run without NAD+ Where does the NAD+ usually come from? Mitochondria e- transport (if O i2 present) If anaerobic, everything backs up giving you more NADH than NAD+ because no oxygen. So you can get NAD+ from anaerobic fermentation Anaerobic fermentation to ethanol o This is done by small microorganisms (like yeast) because they don’t need as much ATP as we do o Yeast has mitochondria but they don’t use it because they are anaerobic o Yeast can run on just glycolysis alone as long as they can get some NADH, and they’re so small that they don’t need much ATP o Why does yeast make ethanol? To get the NAD+ needed for glycolysis Chapter 10 Chloroplasts and Photosynthesis Comparison of mitochondria and chloroplast Mitochondria o F1 faces the matrix o High H+ outside of the inner membrane o ATP is made inside the MATRIX due to influx Chloroplasts are like inside-out mitochondria in what they do Chloroplast o CF1 faces the stroma o High H+ inside the thylakoid membranes o The inner membrane is now inside out for the chloroplast o Space in chloroplast is called Stroma instead of inner membrane for mitochondria and the matrix in chloroplast is called the Lumen o Now ATP gradient is facing out instead of facing in, the proton gradient has flipped o ATP is made outside due to efflux The CF1/CF0 can make ATP in vitro All of this is done in the dark What provides energy to make ATP? Is it Electron transport or Proton Gradient? o No light no electron transport! o What is membrane potential here? I will guess the pmf is inward because High proton concentration is so high out than in. just let it sit over night until in and out has same concentration. o We’re changing pmf going in to now pmf going out by incubating it. We add ADP+Pi, so pmf is outward then now we made ATP by setting up artificial proton gradient Electron transport is necessary for ATP synthesis in VIVO not VITRO ATP synthesis in vitro Thylakoids are like inside-out inner mitochondria membranes: 1. They need a higher H+ inside than outside 2. The pmf should be outward if you want to make ATP 3. The pmf provides the energy for ATP synthesis outside After a long time (incubation) in pH = 3.8, the inside becomes pH = 3.8 What does this do? It loads up the inside with high [H+] This sets up an artificial H+ gradient Does this provide all of the energy that is needed to make ATP? this shows pmf outwards Could this be done with inside-out mitochondria inner membranes? YES. Because F1 will be on outside, load inside with proton concentration. Would it require NADH? NO, because light provides energy for plants to set up proton gradient. Now we have ARTIFICIAL proton gradient, so we don’t need NADH. Overall scheme of photosynthesis: o CO 2s reduced to make carbohydrates by reductive biosynthesis in the Dark Reactions. o Reductive biosynthesis makes sugars by reduction. We take those reduced sugars and oxidize them. It is a CYCLE o This requires ATP and NADPH from the light reaction Two parts of photosynthesis: light and dark. The light reaction doesn’t make sugars, the dark reactions do. But dark reactions require ATP and NADPH which comes from light reactions. (NADH for mitochondria, NADPH for chloroplast) Chloroplast, a plant organelle Thylakoid membranes are high in protein and have no cholesterol two membranes: outer and inner 1. The envelope membrane is a double membrane. These membranes are not the photosynthetic membranes 2. The thylakoid membranes are the photosynthetic membranes inside the chloroplasts. The light harvesting proteins (pigments), electron transport proteins and the ATP synthetase are all on or in the thylakoid membranes 3. The lumen is the aqueous area inside the thylakoids 4. The stroma is the aqueous area outside the thylakoids (Inside mitochondria matrix, as you go out you see inner membrane then inner membrane space then cytoplasm. Inside chloroplast, I’m in lumen as I move out I see thylakoids, lumen is inside thylakoids you come out, you see stroma) 5. No TCA cycle in chloroplasts. They are different than mitochondria. They do have their own DNA but 90% of their proteins come from nuclear-encoded genes Pathway for electron transport in chloroplast thylakoids o PQ is plastoquinone. It is a lipid o PC is Plastocyanin. It is a peripheral protein o Light is not the first electron donor. It is from WATER. Electrons are then passed on from water to PQ to PC, the final electron acceptor is NADP+ to make NADPH (here we set up a proton gradient to make NADPH) light provided the energy to split water into Hydrogen and electrons Mitochondria and chloroplast electron transport Mitochondria What do you get from this first electron transport? A proton gradient Chloroplasts What do you get? A proton gradient and NADPH (NADPH is important to chloroplast) In both mitochondria and chloroplasts, H+ flux is coupled to ATP synthesis In both, ATP is made when H+ flow from the low pH side to high pH side, the pmf is enough to make H+ on the other side Is this a chloroplast or a mitochondria membrane? How can you tell? You can’t Chloroplasts H O oxidized to O 2 2 Energy required (light) Makes sugars from CO 2 H+ high inside thylakoids CF 1aces out H+ efflux during ATP synthesis NADPH (Nicotinamide adenine dinucleotide phosphate) Mitochondria O 2educed to H O 2 Energy produced (ATP) Makes CO fr2m sugars H+ high outside inner membrane F 1aces in H+ influx during ATP synthesis NADH (Nicotinamide adenine dinucleotide)
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