|Lecture Notes: 17 November||
The Tricarboxylic acid cycle is in many ways the central pathway of metabolism, both catabolically and anabolically: it is involved in the breakdown and synthesis of a variety of compounds. Right now we want to focus on its catabolic role in aerobic catabolism: the oxidative breakdown of the acetyl group of acetyl CoA. In this instance we can consider the entire cycle to be a catalyst for this breakdown.
The problem is that the C-C bond of the acetyl group is chemically very resistant. Recall that in organic chemistry generally get C-C bond cleavages at alpha-beta bonds to carbonyl carbons, but with the acetyl group there is no beta carbon. So the TCA Cycle creates an alpha-beta bond by first attaching the acetyl group onto a carrier molecule, oxaloacetate.
Note that for pathways with a carier the carrier must be regenerated to consider the pathway completed!
Let's look at an overview of the Kreb's TCA Cycle. First condense the acetyl group with a four carbon carrier to get a six carbon tri-acid. This is then rearranged and oxidized with loss of carbon dioxide to give a five carbon di-acid ketol very similar to pyruvate in structure. An irreversible DH Complex then creates a four carbon CoA derivative with the release of a second carbon dioxide. At this point it appears that acetyl has been released as carbon dioxide, however, the carrier has been reduced, and modified. A series of reactions now regenerates the original carrier.
The first reaction of the cycle is an aldol condensation catalyzed by
Unfortunately the resulting citrate is a tertiary alcohol which cannot be readily oxidized. Aconitase catalyzes the rearrangement of citrate to give an oxidizable secondary alcohol. This reaction involves an elimination/addition sequence, catalyzed by an iron-sulfur cluster (Fe4S4), with an alkene intermediate, cis- Aconitate:
We have now converted the 3° alcohol, citrate, into an oxidizable 2° alcohol, isocitrate. The next reaction is the first oxidation of the TCA Cycle.
The isocitrate alcohol can now be oxidized with NAD+ by Isocitrate DH to give an enzyme bound intermediate. The intermediate has a carboxyl group beta to a carbonyl carbon, so it has an excellent leaving group, CO2, attached to a stabilized carbanion. Thus it immediately rearranges to lose carbon dioxide:
The resulting 2-oxo-glutarate (-ketoglutarate) looks just like pyruvate with an R-group attached to the -carbon, so it is broken down by a DH Complex, the -Ketoglutarate DH Complex, just as pyruvate was. As a DH Complex this is an irreversible enzyme, making the entire cycle irreversible. This gives succinyl-CoA and releases a second carbon dioxide. Note that at this point two carbons have been released, so formally, we have released the two carbons of Acetyl-CoA (Though neither of them came from the acetyl CoA we added)! We have also produced two NADH's (4 NADH/Glucose) which will result in the production of 5 ATP's (or: 4 x 2.5 ATP/NADH= 10 ATP's/Glucose). However, we have not regenerated the carrier. The remainder of the cycle is involved in this regeneration. (Again in the first four reactions two carbons have been lost as CO2 - as many carbons have been lost as were picked up with acetate. In a sense the rest of the cycle is regenerating our carrier - oxalacetate!)
Succinyl-CoA, like acetyl-CoA, has a high-energy bond. However in this case the energy will be captured, using Succinyl-CoA Synthetase, to give a GTP which is energetically equivalent to an ATP (2 ATP's/Glucose). The mechanism of this reaction first involves the phosphorolysis by inorganic phosphate of the thiol ester bond to give a phosphoric-carboxylic mixed acid anhydride, followed by formation of a phosphorylated enzyme and finally transfer of the phosphate onto GDP.
The reactions beginning with succinate are representative of a common pattern, the "Mainline Sequence," seen repeatedly in biochemical pathways.
Last modified 18 November 2008