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by: Shelly Notetaker

Lec18_1spp.pdf 405

Shelly Notetaker
GPA 3.0
Jim Hurley

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About this Document

Jim Hurley
One Day of Notes
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This 33 page One Day of Notes was uploaded by Shelly Notetaker on Thursday November 6, 2014. The One Day of Notes belongs to 405 at Louisiana State University taught by Jim Hurley in Fall. Since its upload, it has received 48 views. For similar materials see Biochem in Biochemistry at Louisiana State University.


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Date Created: 11/06/14
Lasft tlimel S1 Citric acid Cycle Ksygh M Today Electron Transport very brief Oxidative Phosphorylation Review if time 39 QJW gt9 a quotw Overview FA 5 POLYSAl 1ARDL39gt Fatty acids and Glucose and glycerol other sugars Acetyl 1 oA Electron transport makes a proton gradient across the inner membrane 2 PRO LIN Amino acids 2 co CoA T Stage Stage I I Stage Mitochondria convert reduction potential into a proton gradient and then into ATP h 0qj P k sez H no K 39 o C u r O o wk 2 Outer membrane 39 s Inner membrane Rough endoplasmic reticulum Figure 172b Fundamentals of Biochemistry 2e 2006 John Wiley amp Sons lntermembrane s ace p Cyt cat ooooooch voooooo Inner mitochondrial V p 6 Cyta membrane K 1 t C 7 Q8 00000 C uoooo Matrix A NMJDH Complexl Complexlll 02 2395 Complex IV CAC 39 H20 39 re 178 Fundamentals of Biochemistry 2e 06JohnWiIeyampSons D Q D H The Proton Gradient and ATP synthesis are coupled Electrons cannot flow it the pH Gradient is too steep Intermembrane H nigh Ir 4 space f 39 e 0 h o complexlv o o o o o o o o o o 0 0 Inner 39 mitochondnal brn 0 390 390 390 396 0 39 t 4 I V t Matrix Complex II Low IH l wntw Protonmotive force rotates F0 relative to F1 944 6maln The next few slides show some experimental approaches to studying oxidative phosphorylation Chemical UDCOU IGFS UISSI ate the roton radlent Matrix Cylosol high pH low pH 0 OH OH 0 j j VT T N02 O2 diffusion O2 NO2 N0 N02 N02 N02 24Dinitrophenol DNP Other uncouplers are FCCP ptrifluoromethoxy carbonyl cyanide phenyl hydrazone CCCP carbonyl cyanide mchloro phenyl hydrazone EI ionisable proton x x N CN NC DNP dinitrophenol Powerful slimming drug gt100000 people treated in 1930 s Converts fat directly into heat Verynarrow therapeutic window Slight overdose causes catstrophic loss of energy in the body Excessive sweating extreme thirst cararacts irreversible damage to heart liver lungs Ultimately paralysis death First drug to be banned by FDA 1938 Oligomycin and venturicidin inhibit the ATP synthase H Oligomycin venturicidin O2 e 2 H Matrix ATP Pi Membrane Stryer 5e Demonstration of chemical uncoupling using isolated mitochondria 02 cons tuned Add venturicidin or oligomycin 3 W U ncou pled ATP S I1th SiZ d Example of a mitochondrial O2 Consumption Rate OCR analysis Oxygen Consumption Rate 1 c e a b g Time 0 a Buffer alone b Mitochondria c Pyruvate fuel d ADP substrate for ATP synthase e Oligomycin Inhibits ATP synthase f FCCP uncoupler g Antimycin blocks respiratory chain How much ATP is made Every NADH that Every FADH2 that 9 t9FS C0mP399X I enters complex II yIeds about 25 ATP yields about 15 ATP These numbers can change degending on how UNCQQELED the mitochondria are 12 Mitochondrial uncoupling Nx V MT Brown Fat generates heat Hibernating animals Newborn infants Lots of mitochondria in fat tissue makes it brown Action of Uncoupling Protein H H H H up1 Electron V transport 39o39o39939939939o39o39939 i 339o g 9 O 0 CdS 39 39V39V39quot 0 0 0 0 0 0 0 0 o 0 gt 0 0 0 0 o g 9 9 9g ac Iva e ooooooo channel a 02 0 AKA nmldL M P1 14 Oxygen Consumption Rate Liver Mitochondria Brown Fat Mitochondria tightly coupled mostly uncoupled f f 11 d d C C e a 4 I M E cuM C E 3 air b g O0 a bl Mnu L Time 0 Time ruff 6 a Buffer alone b Mitochondria c Pyruvate fuel d ADP substrate for ATP synthase e Oligomycin Inhibits ATP synthase f FCCP uncoupler g Antimycin blocks respiratory chain How is the energy in the proton gradient captured to make ATP ATP Synthase F1 particles Mltochondnon Outer membrane Matrix Inner membrane Structure of ATP Synthase Can be separated into 2 sub structures F1 fragment The knob Subunits 3 0c 3 3 1 y 1 6 1 8 3 catalyticsubunit Structure of ATP Synthase 2nd substructure F0 fragment Integral membrane protein 0 pC arquot subunits 1 a 2 b 1012 c Cytoplasm Cell membrane Periplasm I9 Structure of ATP Synthase Ii L H l39 0 cf W B Z ATP N 13 C Plasma A THIS IS THE MATRIX b H Z SIDE C JR S 56 membrane y cytoplasm Q ya l9wm r OR MITOCHONDRIA H 739 I5 39 3 39 39 L 6 2 Q Periplasmic space i we 1826 13 John Wiley amp Sons Inc All rights reserved 20 How does the proton gradient drive ATP synthesis The proton binding and dissociation cause rotation of F0 compared to F1 The rotation induces conformational changes in the oc3 comlexes of F1 A given 3 subunit cycles between three conformations 39 3 ADP ADP bound Loose 3ATP ATP bound tightly Tight 3 Empty Open Mechanism of ATP synthesis Each unit is an H5 Complex Proton driven rotation of F0 L T ADP P ADP P 139 Energy 39 1 ATP H 2O X 3 1 J T4 0 l L y subunit green tu 3ADP L conformation binds substrate and product loosely 39 3ATP T conformation binds ATP tightly 3 empty O conformation is not binding ATP or ADP 3 lt2 Structure of ATP Synthase E 4 The next slides show experimental evidence that A ATP supports this rotary model B Hjdroyd 4 HP 391 1k 39 Plasma As can b membrane e H y cytoplasm H H Pj C c Periplasmic space Figure 1826 2013 John Wiley 8 Sons Inc All rights reserved 23 Actin lament Streptavidin cCys2 biotin His tag Figure 1828a 2013 John Wiley 8 Sons Inc All rights reserved 24 Key concepts for Oxidative Phosphorylation Respiratory chain pumps protons across the inner mitochondrial membrane Uncouplers and inhibitors of ATP synthase can influence oxygen consumption ATP synthase functions as a rotary motor with three active sites and a ring that moves as it binds and releases protons 26 REVIEW E there is remaining time at the end of this lecture I may review material from the past 7 lectures We may discuss one or two examples of metabolic diseases from this list Do NOT memorize this ist Lactose Intolerance Fructose Intolerance McCarde Syndrome Warburg Effect Cancer Pyruvate Kinase Deficiency Hemolytic anemia Hexokinase Deficiency Hemolytic anemia Triose phosphate lsomerase Deficiency DH Succinate Dehydrogenase Fumarase mutations Cancer Oncometabolite alpha hydroxyglutarate PFK1 deficiency Glycogen storage disease Arsenic poisoning LDH deficiency Leigh Syndrome PDH deficiency 27 Some key things to understand about carbohydrates Aldoses Ketoses Hemiacetal D vs L configuration Enantiomers Epimers Anomeric carbon oc vs 3 Glycosidic bonds Reducing vs nonreducing sugars Types of disaccharides polysaccharides Amylase Glucose isomerase HFCS 28 Some Key Concepts for Glycolysis Sources of energy ATP Phosphocreatine glycolysis respiration Types of Glucose Transporters and hexokinases Importance of coupled reactions Overall scheme for glycolysis 10 steps of glycolysis and their significance Investment vs Payoff phases lsomerase vs Mutase Significance of 23 BPG Which steps use and which steps make ATP How NADH is regenerated in animals vs in yeast How alcohol is made and how it is metabolized 29 Some key concepts about regulation of Glycolysis Regulation occurs at steps with the highest free energy difference Hexokinase PFK 1 Pyruvate Kinase Be familiar with positive and negative regulators Be familiar with types of regulation Regulation depends on the purpose of the tissue example muscle vs liver PFK1 stimulation by F26BP and relationship to ATP and AMP regulation Fructose metabolism differs from glucose metabolism 30 Key Concepts for Pyruvate Dehydrogenase Huge multienzyme complex types of subunits types of prosthetic groups types of reactions they catalyze how they interact with each other Regulation Inhibition by products NADH and Acetyl CoA Phosphorylation of Pyruvate Dehydrogenase Regulation of PDH Kinase Regulation of expression Mechanism by which HIF senses Oxygen 31 Key concepts of the citric acid cycle 1 Hub of cellular metabolism 2 The net reaction of the citric acid cycle is Acetyl CoA 3NAD FAD GDP Pi 2H2O 2CO2 3NADH FADH2 GTP 2H CoA Regulation occurs at steps with largest free energy drop The two carbons from Acetyl CoA are fully oxidized to CO2 TCA intermediates are precursors for other compounds 4 pairs of electrons are transferred to oxidize Acetyl CoA to CO2 Shuttles transfer reducing power from to the cytosol to the mitochondrial matrix FDUquot39gt 32 Key concepts for Oxidative Phosphorylation Respiratory chain pumps protons across the inner mitochondrial membrane Uncouplers and inhibitors of ATP synthase can influence oxygen consumption ATP synthase functions as a rotary motor with three active sites and about a ring that moves as it binds and releases protons 33


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