lecture17_sb_08.pdf

BC405 Fall 208 Friday October 31st Kane 130, 10:30 - 11:20am Lecturer: Dr. Brockerhof 2 Metabolic Overview C CH 2 CH 2 COOHO COO COO Citrate CH 2 C CH 2 COO COO O ?-ketoglutarate C CH CH 2 COO COO COO H HO Isocitrate CH 2 C CH 2 S COO O CoA Succinyl-CoA CH 2 COO CH 2 COO Succinate HC COO CH COO Fumarate CH 2 COO CH COO HO Malate CH 2 COO C COO O Oxaloacetate H 3 C C S CoA O Acetyl CoA CO 2 CO 2 1/2 1/2 C CH 2 CH 2 COOHO COO COO Citrate CH 2 C CH 2 COO COO O ?-ketoglutarate C CH CH 2 COO COO COO H HO Isocitrate CH 2 C CH 2 S COO O CoA Succinyl-CoA CH 2 COO CH 2 COO Succinate HC COO CH COO Fumarate CH 2 COO CH COO HO Malate CH 2 COO C COO O Oxaloacetate H 3 C C S CoA O Acetyl CoA CO 2 CO 2 5 6 Balanced Equations ?Glucose + 2P i + 2ADP + 2NAD + -> 2pyruvate + 2ATP + 2NADH + 2H + 2H 2 O ?Pyruvate + NAD + CoA -> acetylCoA + CO 2 + NADH + H + ?Acetyl CoA + 3NAD + FAD + GDP + P i + 2H 2 O -> 2CO 2 + 3NADH + FADH 2 + GTP + 3H + CoA 7 Balance Shet ATPNADHFADH 2 Pyruvate to Acetyl CoA (per mole pyruvate) citric acid cycle (per mole pyruvate) Total/mole pyruvate Total/mole glucose Glycolysis (per mole glucose) Overall total/mole glucose 0 1 0 1 3 1 1 4 1 2 8 2 2 2 0 4 10 2 8 Fundamentals of Biochemistry Review Oxidation Reduction Principles Chapter 13 (Second Edition): Pgs(414-419) Chapter 14 (3rd Edition): Pgs (469-474) Copyright © 2006 by John Wiley & Sons, Inc. Donald Voet ? Judith G. Voet ? Charlotte W. Pratt 9 10 Overview ?During oxidative phosphorylation, ?The potential energy of the electrons in NADH and FADH 2 is converted to a proton gradient. ?The energy in the proton gradient is converted to ATP. ?Phase 1: Three electron driven proton pumps create a proton gradient ?Phase 2: ATP synthase converts the proton gradient into ATP. 1 Reduction Potential ?A measure of the potential energy of electrons. ?Electron transfer potential. X + 2H + + 2e - <-> XH 2 oxidized formreduced form ?XH 2 and X are a redox pair. 12 Measurement of Reduction Potential B BH 2 X XH 2 voltmeter salt bridge electrode ?X + 2e - + 2H + <-> XH 2 ?B + 2e - + 2H + <-> BH 2 ?X + BH 2 <-> XH 2 + B 13 Reduction potential of XH 2 compared to a reference std. ?Standard conditions: 1 M everything. ?Compared to a reference 2H + 2e - <-> H 2 (arbitrarily set at zero). ?E 0 (V): If negative, e - ?s flow towards the ref. cel. If positive, e - ?s flow towards the X & XH 2 half cel. 1M H + 1 atm H 2 1 M X 1 M XH 2 1 M H + salt bridge E 0 14 The biochemical standard state: pH 7.0 ?E? 0 : 1 M everything except 1 x 10 -7 M H + . ?NOTE: The reference is the same. 1M H + 1 atm H 2 salt bridge 1 M X 1 M XH 2 pH 7.0 E? 0 15 Some Standard Reduction Potentials ? Aceptors Donors E? 0 ? 1/2 O 2 H 2 O +0.82 ? FAD FADH 2 +0.03 ? fumarate sucinate +0.03 ? oxaloacetate malate -0.17 ? NAD + NADH -0.32 ? ?-ketoglutarate isocitrate -0.38 16 E = E ' 0 + 2.303 RT n F lo g [ a c c e p to r ] [d o n o r ] The Nernst Equation ?n is the number of electrons ?F is Faraday?s constant: 96.5 kJ volt -1 mol -1 ?R is the gas constant: 8.315 J K-1 mol -1 ?T is in degres Kelvin ?If [aceptor] = [donor] then E = E? 0 17 ?E? 0 ?Acetaldehyde + NADH + H + <--> ethanol + NAD + Two half reactions: ?acetaldehyde + 2H + + 2e - <-->ethanol -0.197 NAD + + 2e - + H + <--> NADH -0.32 ??E? 0 = E? 0 aceptor - E? 0 donor ?Which is the electron acceptor? What is ?E? 0 for this reaction? 18 ?E? 0 ?Acetaldehyde + NADH + H + <-> ethanol + NAD + Two half reactions: ?acetaldehyde + 2H + 2e - <->ethanol -0.197 NAD + 2e - + H + <-> NADH -0.32 ??E? 0 = E? 0 aceptor - E? 0 donor ??E? 0 = - .197 - (-0.32) = 0.123 19 ?Consider two half reactions: ?X + 2e - + 2H + <-> XH 2 ?B + 2e - + 2H + <-> BH 2 ?X + BH 2 <-> XH 2 + B ?At equilibrium E X = E B ?Acording to the Nernst equation, then ?E? 0 and the equilibrium constant 20 ?E? 0 and the equilibrium constant E ' 0 X + 2.303 RT n F lo g [ X ] [ XH 2 ] = E ' 0 B + 2.303 RT n F lo g [ B ] [ BH 2 ] E ' 0 X ! E ' 0 B = 2.303 RT n F lo g [B ] [ BH 2 ] ! lo g [ X ] [ XH 2 ] " # $ $ % & ? ? ! E ' 0 = 2.303 RT n F lo g [ B ][ XH 2 ] [BH 2 ][ X ] " # $ $ % & ? ? 21 ?E? 0 and the equilibrium constant ! E ' 0 = 2.303 RT n F lo g K e q ! G ' 0 = " 2.303 RT lo g K e q ! G ' 0 = " n F ! E ' 0 X + BH 2 <--> XH 2 + B ! E ' 0 = 2.303 RT n F lo g [ B ][ XH 2 ] [BH 2 ][ X ] " # $ $ % & ? ? 2 ?E? 0 and the equilibrium constant ?If ?G? 0 is negative, ?E? 0 is positive and the reaction is exergonic. ?If ?G? 0 is positive, ?E? 0 is negative and the reaction is endergonic. 23 24 25 Malate-aspartate shuttle. This shuttle is particularly important in heart and liver. It is a shuttle for NADH that does NOT cost ATP. The malate dehydrogenase (MDH) occurs as separate enzymes in mitochondria and cytosol (cMDH & mMDH). Transamination reactions will be covered in more detail next quarter. Transaminase CMDH Transaminase MDH Mod. From Stryer 5/e 26 Fundamentals of Biochemistry Electron Transport Chapter 17 (Second Edition): Problems 1,2,3,4,5,6,1 Chapter 18 (3rd Edition): Problems 1,2,3,4,5,6,1 Copyright © 2006 by John Wiley & Sons, Inc. Donald Voet ? Judith G. Voet ? Charlotte W. Pratt 27 28 Electron Transport Chain Intermembrane space Matrix 4H + 4H + 2H + succinatefumarate NADH + H + NAD + 1/2 O 2 + 2H + H 2 O Low [H + ] High [H + ] 29 Cytochromes: Electron carriers ?All have heme prosthetic groups ?Pass one electron Fe 3+ --> Fe 2+ ?Reduction potentials vary from 0.077 to 0.55 volts ?Different micro-environments in the protein. 30 Electron cariers: Iron-Sulfur Proteins ?Reduction potentials vary from -0.65 V to +0.45 V depending on microenvironment in the protein. 31 Electron carriers: FMN 32 Electron carriers: FMN 3 Electron cariers: Ubiquinone (Q) ?Can acept two electrons ?Pas them on one at a tie. ?Forms a radical or seiquinone. 34 Electron Transport Chain NADH + H + NAD + E? 0 =-0.32V 2e - ?s FMN FeS 1/2 O 2 +2H + -->H 2 OE? 0 =0.82V Cyt a, a 3 , Cu FAD FeS sucinate fumarate not a protein soluble protein Cyt b,c 1 , FeS Integral membrane proteins in the inner membrane Susan Brockerhoff lecture#17_sb_07.ppt