4 Glyoxylate cycleThe glyoxylate cycle results in the net conversion of two acetyl-CoA to succinate instead of 4 CO2 in citric acid cycle.Succinate is transferred to mitochondrion where it can be converted to OAA (TCA)Can go to cytosol where it is converted to oxaloacetate for gluconeogenesis.Net reaction2Ac-CoA + 2NAD+ + FAD OAA + 2CoA + 2NADH +FADH2 + 2H+Plants are able to convert fatty acids to glucose through this pathway
6 Electron transport and oxidative phosphorylation Complete oxidation of glucose by molecular oxygen can be described as:C6H12O6 + 6O2 6CO2 + 6H2O Go’=-2823 kJ/molCan be broken down into two half-reactions with the transfer of electronsC6H12O6 + 6H2O 6CO2 + 24H+ +24e-6O2 + 24H+ + 24e- 12H2O12e- from the oxidation of glucose are not transferred directly to O2, go to NAD+ and FAD to form 10NADH and 2FADH2These are reoxidized, passing their electrons to the electron- transport chain to reduce O2 to H2O causing the mitochondrion to create a proton gradient.This pH gradient is used to drive the synthesis of ATP via oxidative phosphorylation.
7 Chemiosmotic TheoryElectron Transport: Electrons carried by reduced coenzymes are passed through a chain of proteins and coenzymes to drive the generation of a proton gradient across the inner mitochondrial membraneOxidative Phosphorylation: The proton gradient runs downhill to drive the synthesis of ATPElectron transport is coupled with oxidative phosphorylationIt all happens in or at the inner mitochondrial membrane
8 Outer Membrane – Freely permeable to small molecules and ions Outer Membrane – Freely permeable to small molecules and ions. Contains porins with 10 kDa limitInner membrane – Protein rich (4:1 protein:lipid). Impermeable. Contains ETR, ATP synthase, transporters.Cristae – Highly folded inner membrane structure. Increase surface area.Matrix- “cytosol” of the mitochondria. Protein rich (500 mg/ml) Contains TCA cycle enzymes, pyruvate dehydrogenase, fatty and amino acid oxidation pathway, DNA, RNA, ribosomesIntermembrane Space – composition similar to cytosol
9 Mitochondrial transport The inner membrane is impermeable to hydrophilic substances. Has special transport systems for the following:Glycolytically produced cytosolic NADH.Mitochondrially produced metabolites (OAA, acetyl-CoA) for cytosolic glucose formation and fatty acid biosynthesis.Mitochondrially produced ATP must go to cytosol where ATP-utilizing reactions take place.Example: cytoplasmic shuttle systems transport NADH across inner membrane.
11 Malate-aspartate shuttle 2 phasesPhase ACytosolic NADH reduces OAA to malate (malate DH).Malate--ketoglutarate carrier transports malate from cytosol to mitochondrial matrix, exchanged for -KGIn the matrix, NAD+ reoxidizes malate to make OAA and yield NADH.Phase B4. Transaminase converts OAA to Asp and Glu to -KG.The glutamate-apartate carrier transports Asp from the matrix in exchange for Glu.Transaminase in cytosol converts Glu to Asp.3 ATPs for every NADH but loses 0.3 ATP because each NADH enters matrix with proton (yield 2.7 ATP).
13 Glycerophosphate shuttle Simpler but less energy efficient than the malate-aspartate shuttle.Supplies electrons in a manner similar to succinate dehydrogenaseApprox. 2 ATP per NADH, about 0.7 ATP less than the malate-aspartate shuttle.Advantage-irreversible so it operates efficiently even when cytoplasmic NADH is low relative to NAD+.Malate-aspartate shuttle is reversible - driven by concentration gradients.
16 Figure 22-9 The mitochondrial electron-transport chain. Page 803
17 Electron TransportFour protein complexes in the inner mitochondrial membraneA lipid soluble coenzyme (UQ, CoQ) and a water soluble protein (cyt c) shuttle between protein complexesElectrons generally fall in energy through the chain - from complexes I and II to complex IV
18 Standard reduction potentials of the major respiratory electron carriers.
19 Cofactors of the electron transport chain Fe-S clustersCoenzyme Q (ubiquinone)Flavin mononucleotideFADCytochrome aCytochrome bCytochrome cCuACuB
20 Iron-sulfur clusters 4 main types of iron sulfur clusters [2Fe-2S] and [4Fe-4S] cluster coordinated by 4 Cys SH[3Fe-4S] is a [4Fe-4S] lacking one Fe atom.[Fe-S] is only found in bacteria, liganded to 4 CysRieske iron-sulfur proteins [2Fe-2S] cluster but 1 Fe is coordinated by 2 His.Oxidized and reduced states of all Fe-S clusters differ by one formal charge.
21 Figure 22-15a. Structures of the common iron–sulfur clusters Figure 22-15a Structures of the common iron–sulfur clusters. (a) [Fe–S] cluster.Page 808
22 Figure 22-15b. Structures of the common iron–sulfur clusters Figure 22-15b Structures of the common iron–sulfur clusters. (b) [2Fe–2S] cluster.Page 808
23 Figure 22-15c. Structures of the common iron–sulfur clusters Figure 22-15c Structures of the common iron–sulfur clusters. (c) [4Fe–4S] cluster.Page 808
24 Figure 22-16 X-Ray structure of ferredoxin from Peptococcus aerogenes. Page 809
25 Figure 22-17a Oxidation states of the coenzymes of complex I. (a) FMN. Can accept or donate 1 or 2 e-Page 810
26 Figure 22-17b Oxidation states of the coenzymes of complex I. (b) CoQ. Coenzyme Q’s hydrophobic tail allows it to be soluble in the inner membrane lipid bilayer.Page 810
27 Figure 22-21a. Visible absorption spectra of cytochromes Figure 22-21a Visible absorption spectra of cytochromes. (a) Absorption spectrum of reduced cytochrome c showing its characteristic a, b, and g (Soret) absorption bands.Page 813
28 Figure 22-21b The three separate bands in the spectrum of beef heart mitochondrial membranes indicate the presence of cytochromes a, b, and c.Page 813
29 Figure 22-22a Porphyrin rings in cytochromes. (a) Chemical structures. Page 813
30 Figure 22-22b. Porphyrin rings in cytochromes Figure 22-22b Porphyrin rings in cytochromes. (b) Axial liganding of the heme groups contained in cytochromes a, b, and c are shown.Page 813
32 Complex I NADH-CoQ Oxidoreductase (NADH dehydrogenase) Electron transfer from NADH to CoQMore than 30 protein subunits - mass of 850 kD1st step is 2 e- transfer from NADH to FMNFMNH2 converts 2 e- to 1 e- transfer6-7 FeS clusters.Four H+ transported out per 2 e-NADH + H+FMNFe2+SCoQNAD+FMNH2Fe3+SCoQH2
33 Complex II Succinate-CoQ Reductase Contains the succinate dehydrogenase (from TCA cycle!)four subunitsTwo largest subunits contain 2 Fe-S proteinsOther subunits involved in binding succinate dehydrogenase to membrane and passing e- to UbiquinoneFAD accepts 2 e- and then passes 1 e- at a time to Fe-S proteinNo protons pumped from this stepSuccinateFADFe2+SCoQFumarateFADH2Fe3+SCoQH2
34 Q-CycleTransfer from the 2 e- carrier ubiquinone (QH2) to Complex III must occur 1 e- at a time.Works by two single electron transfer steps taking advantage of the stable semiquinone intermediateAlso allows for the pumping of 4 protons out of mitochondria at Complex IIIMyxothiazol (antifungal agent) inhibits electron transfer from UQH2 and Complex III.UQUQ.-UQH2