Humboldt State University ® Department of Chemistry

Richard A. Paselk

Chem 431	Biochemistry	Fall 2001
Lecture Notes:: 14 November	© R. Paselk 2001
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KREB'S TCA CYCLE, cont.

Note that because all catalysts (oxaloacetate, enzymes etc.) must be regenerated in looking at the overall operation of the cycle, only the acetyl group of acetyl-CoA can be oxidized completely. Some intermediates, such as citrate, can be partially oxidized, but Kreb's cycle intermediate catabolism requires leaving the cycle at oxaloacetate and then returning as acetyl-CoA. Note that this requires leaving the mitochondria for some reactions, and since the extremely low concentrations of oxaloacetate don't allow its efficient transport across the mitochondrial membrane (the K_m of the carrier is much higher than [oxaloacetate]), malate is the species which actually leaves the mitochondria. We looked at the details of some of these reactions when we looked at gluconeogenesis.

Regulation of the TCA Cycle

In order to understand the regulation of the TCA cycle we need to look at the DG values for the various reactions and the kinetic properties of the enzymes. Values for the non-equilibrium reactions are tabulated below:

Data from E. A. Newsholme and A. R. Leach Biochemistry for the Medical Sciences. John Wiley & Sons, New York (1983) pp 101 & 110.  Enzyme Substrate Substrate Conc. (mM)  Km (mM) DG (kJ/mol)  Effectors  Citrate synthase acetyl-CoA 100-600 5-10 -53.9 Succinyl CoA (-), ATP (-), NADH (-) oxaloacetate 1-10 5-10  Isocitrate DH (NAD+) isocitrate 150-700 50-200 -17.5  Ca2+ (+), ATP (-), ADP(+), NADH (-)  2-Oxoglutarate DH 2-oxoglutarate 600-5900 60-200 -43.9  Ca2+ (+), Succinyl CoA (-), NADH (-)

Note that normally the concentrations of ATP, ADP, NAD⁺, and NADH are relatively constant in the mitosol and are thus unlikely to be very effective as allosteric regulators under most circumstances. On the other hand the availability of NAD⁺ and FAD as substrate will affect the rate not only of the reactions in the table , but also the near-equilibrium dehydrogenases. Note that NAD⁺ availability in turn is determined by the activity of the electron transport system, whose activity is closely coupled to the availability of ADP. Thus high [ATP] will slow the TCA cycle since high [ATP] means low [ADP], which will slow the ETS resulting in low [NAD⁺]!

In muscle, Ca²⁺ does show significant changes in concentration in the mitosol (recall that an increase in [Ca²⁺] concentration initiates muscle concentraction). Succinyl CoA will also show significant concentration changes under differing conditions and can thus also serve as an effective regulator, indicating carbon status in the second half of the cycle.

Integration of TCA Cycle with Glycolysis: The concentration of citrate also affects PFK activity as a negative effector.

Interconversion of metabolic intermediates: The TCA cycle has a central place in metabolism (even in anaerobic organisms) via its use to interconvert metabolites as summarized in Figure 20.22 in G&G (p 662). We will look further at these interactions as the course proceeds.

Mitochondrial Electron Transport

Mitochondria: First let's review mitochondrial structure (V&V Figure 17.2 & 17.3) [overhead 18.2 P]. Recall that the inner membrane is very tight - that is the passage of polar molecules and ions is prevented without a specific transport vehicle. The inner membrane is also protein rich. If carefully broken down we find it is very rich in five protein complexes: I -IV are large protein complexes involved in electron transport, while V is the ATP sythatase driven by proton gradients.

Pathway Diagrams

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Last modified 3 December 2001