Download Lecture 3section7

Lecture 3:
November 29th
Dr. Moxley
(Tricarboxylic Acid Cycle)
Functions of TCA cycle
Oxidizes Acetyl CoA derived from pyruvate or fatty acid oxidation
Production of NADH and FADH for the synthesis of ATP
Side reactions produce substrates for biosynthesis
Pyruvate dehydrogenase
Acetyl CoA can be derived from amino acids or free fatty acids
Major source is pyruvate
bridge between glycolysis and the TCA cycle is the oxidative decarboxylation of
pyruvate by the pyruvate dehydrogenase complex
Mitochondrial enzyme complex
Consists of multiple copies of 3 enzymes
pyruvate decarboxlyase
dihydrolipoyl transacetylase
dihydrolipoyl dehydrogenase
Regulation of PDH
Primary inhibitor - ATP
Also inhibited by acetyl CoA and NADH
influences of above is enhanced by presence of long chain fatty acids
Allosterically activated when AMP, free CoA and NAD accumulate
In vertebrates the complex contains two additional enzymes
kinase and phosphatase
kinase tightly associated with complex
phosphatase more loosely associated
act on 3 serine residues in pyruvate carboxylase
phosphorylated form of enzyme complex is inactive
Kinase allosterically activated by
ATP, Acetyl CoA and NADH
inhibited by ADP and pyruvate.
dephosphorylated PDH is active
phosphatase affected by Ca++
increased Ca++ increases association with complex increases
activity
Essentially energy charge controls activity as reflected by the following ratios
CoA/Acetyl CoA, NAD+/NADH and ADP/ATP
Regulation of TCA cycle proper
Review of pathway and regulatory sites
Regulatory enzymes
citrate synthase
isocitrate dehydrogenase
∀ ketoglutarate dehydrogenase
Note “logic” of molecule which affect enzyme activity: Relate to what this indicates
about needs of cell
Citrate synthase
Positive effectors
Availability of substrates Acetyl CoA and OAA exerts positive affect on
activity
ADP
Negative effectors
feedback inhibition by citrate or succinyl CoA
NADH
ATP
Isocitrate dehydrogenase
Positive effectors
ADP
NAD+
Negative effectors
ATP
NADH
∀ Ketoglutarate dehydrogenase
inhibited by Succinyl CoA and NADH
Again energy charge of the cell influences activity of pathway
Remember pathways are integrated
Rates of glycolysis and TCA cycle are matched so that only as much glucose is
metabolized to pyruvate as is need to provide Acetyl CoA for the cycles
Rate of glycolysis is matched to the TCA cycle by ATP and NADH levels. Also
remember the citrate is a negative allosteric effector of PFK-1
Glyoxylate Cycle
Alternative metabolic fate for acetyl CoA
present in plants and microorganisms
allows anabolism to 4 carbon level rather than catabolism to 1 carbon
Five steps two unique to glyoxylate pathway catalyzed by
isocitrate lyase
malate synthetase
remaining steps are TCA cycle enzymes
Indispensable pathway in species which can grow on 2 carbons substrates
In plants cycle is compartmentalized to glyoxisome
TCA cycle enzymes are organelle specific isozymes
Function of Pathway
Bypass 2 steps in TCA cycle which release CO2
2 acetyl CoA used converted to a four carbon unit – succinate
succinate can then be converted to malate which can be used in anabolic
pathways.
Why can’t this happen in the TCA cycle?
Regulation of Glyoxylate cycle
Eukaryotes
lyase subject to allosteric inhibition by
AMP
PEP
and TCA cycle intermediates
Prokaryotes
Regulation of glyoxylate cycle via phosphorylation of isocitrate
dehydrogenase
Phosphorylation inhibits activity
bifunctional kinase/phosphatase control flux through glyoxylate pathway.
Inhibition of IDH cause increase in level of isocitrate thus increasing
velocity of isocitrate lyase
Regulation of IDH phosphorylation
Inhibit kinase activity and activate phosphatase activity
ADP, AMP
isocitrate, oxaloacetate, ∀ ketoglutarate
PEP, 3-phophoglycerate, pyruvate
Mechanism for monitoring such large number of metabolites?