Transcript Fundamentals of Biochemistry 2/e

Chapter 14
Chem 341
Suroviec Fall 2013
I. Citric Cycle Overview
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8 reactions
Oxidizes acetyl group of Acetyl CoA to 2 CO2
1.
2.
I. General Features
Circular Pathway oxidizes acetyl
groups from many sources
Net reaction
3.
In eukaryotes all enzymes of CAC are located in
mitochondria
4.
5.
1 CO2 produced in 1 round of the cycle
Oxidation of acetyl groups to 2 CO2 requires transfer of 4 pair of
electrons
II. Synthesis of Acetyl CoA
A.
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Pyruvate Dehydrogenase: Multienzyme Complex
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Group of non covalently associated enzymes that catalyze 2+ sequential steps in
metabolic pathway
Formed from pyruvate through oxidative
decarboxylation
– Pyruvate dehydrogenase (E1)
– Dihydrolipoyl transacetylase (E2)
– Dihydrolipoyl dehydrogenase (E3)
a)
24 E2 proteins associated as trimers at the corners of cube
b)
24 E1 proteins form dimers that associate with E2 core along the 12 edges.
The 12 E3 proteins form dimers that attach to the 6 faces of E2 cube
c) Combining a) and b) forms a 60 subunit complex
B. Pyruvate Dehydrogenase
Overall Reaction
1.
Pyruvate dehydrogenase (E1)
A TPP requiring enzyme
TPP acts as electron sink in the reaction
2. Lipoamide
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Hydroxyethyl group transferred to E2
Lipoamide uses lysine
Cyclic disulfide reversibly reduced
3. E2 transesterification
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Yields acetyl CoA and
dihydrolipoamide-E2
4. Regenerate E2
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E3 becomes reduced
Regenerates E2
Disulfide interchange reaction
5. Reoxidize E3
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Reoxidize E3
III. Enzymes of CAC
1.
Citrate synthase
1.
Aconitase
3.
Isocitrate dehydrogenase
3.
a-ketoglutarate dehydrogenase:
5.
Succinyl CoA synthetase
5.
Succinate dehydrogenase
7.
Fumerase
7.
Malate dehydrogenase
III. Enzymes of the CAC
A.
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Citrate Synthase
Catalyzes the condensation of acetyl-CoA and
oxaloacetate
Free enzyme is a dimer
Active site closes when oxaloacetate binds
Conformational changes seals oxaloacetate in binding
site and shuts out the solvent
B. Aconitase
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Catalyzes reversible isomerization of citrate to
isocitrate
C. NAD+ Dependant Isocitrate Dehydrogenase
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Catalyzes oxidative decarboxylation of isocitrate to a-ketogluterate
D. a-ketoglutarate dehydrogenase
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Catalyzes oxidative decarboxylation
of a-ketogluterate
Is a multienzyme complex
E1: a-ketoglutarate dehydrogenase
E2: dihydrolipoyl transsuccinylase
E3: dihydrolipoyl dehydrogenase
E. Succinyl-CoA Synthetase
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Cleaves “high-energy” succinyl-CoA to synthesis of GTP
Reaction almost energy neutral.
F. Succinate Dehydrogenase
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Catalyzes stereospecific dehydrogenation of succinate to fumerate
Inhibited by malonate
G. Fumerase
Catalyzes the hydration of double bond of fumarate to form
malate
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H. Malate dehydrogenase
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Regeneration of oxaloacetate
IV. Regulation of CAC
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Availibity of substrates
Need for CAC intermediates
Demand for ATP
A. Regulation of pyruvate decarboxylation
1.
2.
Product inhibition by NADH and acetyl-CoA
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NADH, acetyl-CoA compete with NAD+ and CoA for binding sites
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Drive with E2 and E3
Covalent modification by phosphorylation/dephosphorylation of E1
B. Rate-Controlling Enzymes
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Same method as in glycolysis
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Flux of metabolites through the CAC is proportional to the rate of cellular
oxygen consumption
3 main mechanisms
– Substrate availability
– Product inhibition
– Competitive feedback inhibition
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B. Rate-Controlling Enzymes
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Regulators are acetyl-CoA, oxaloacetate, NADH
Flux varies with substrate concentration
V. Pathways that use CAC intermediates
1.
Glucose biosynthesis
2.
Fatty acid synthesis
3.
Amino Acid synthesis