biochem exam 4 (TCA)

what are the fates of pyruvate after glycolysis?

-pyruvate can be reduced in anaerobic fermentation or oxidized for greater metabolic yield

-each carbon of pyruvate is converted into CO2 in the citric acid cycle

what is pyruvate converted to? why?

-pyruvate is converted to acetyl CoA because it is higher in energy

-acetyl CoA contains an RNA component, a fatty acid component, and the thiol part does the chemistry

how is pyruvate converted to acetyl coa? what are the byproducts?

-pyruvate is converted to acetyl coa through oxidative decarboxylation

-CO2 and NADH are produced as byproducts

what is the TPP cofactor?

-the cofactor involved in the conversion of pyruvate → acetyl CoA
-it is a ylide, so it has both a positive and negative charge right next to each other

why is the TPP ylide cofactor useful? why is the CoA thioester useful?

-the CoA can be used as a nucleophile

-the ylide is an aromatic ring that can be used as an electron sink because the ring is positively charged and can pull electrons to itself

-this means pyruvate can kick off a CO2 without being unstable

what is the pyruvate hydrogenase complex?

-three enzymes catalyze 5 different reactions with 5 different cofactors

what is TPP necessary for?

-TPP is necessary for decarboxylation

-without this cofactor this reaction doesn’t happen

-pyruvate is an alpha ketoacid so it can’t just spontaneously decarboxylate

what are NAD and FAD used for?

-NAD and FAD are necessary for redox, they hold the electrons produced by decarboxylation

what does lipoate do?

-lipoate acts as a carrier and aids in redox

-it has long arms that swing around

what is acetyl coa? why is it so high energy?

-acetyl coa is the final product, it is an activated acyl group

-a thioester, which is the highest energy carboxylic acid derivative that hydrolyze irreversibly

-super electrophilic and does not want to be there

what are the 3 enzymes of the pyruvate dehydrogenase complex?

-pyruvate dehydrogenase (TPP)

-dihydrolipoyl transacetylase (lipoamide)

-dihydrolipoyl dehydrogenase (FAD)

what is the pdc doing?

-turning pyruvate into the higher energy acetyl coa

-decarboxylation with TPP

-oxidation and transfer with the lipoamide

-regeneration of NADH with FAD because FAD is giving the electrons from pyruvate to NAD+ to make it NADH

is the pyruvate dehydrogenase step (TPP) reversible?

-the PDH step is irreversible, it is the first committed step

what is the first step of the pyruvate dehydrogenase mechanism of converting pyruvate → acetyl coa?

-the TPP ylide negative charge attacks the pyruvate ketone

-the ketone carbonyl grabs a proton from a nearby acid

-this results in the ylide being covalently attached to the pyruvate which now has an OH instead of a =O

what happens now that we have this TPP-pyruvate complex?

-we have an extended electronic delocalization system and there is the double bond nitrogen of the TPP ylide that can act as an electron sink

-this means that we can decarboxylate! the O- attacks the carbonyl which attacks the alpha-beta bond so the electrons are moving into the electron sink

-losing CO2 is irreversible

now pyruvate has lost CO2, what happens from here?

-lipoic acid swings in with an oxidized disulfide (no Hs on the sulfurs)

-the disulfide is electrophilic and the leftover part of pyruvate bound to TPP is nucleophilic

-so the nucleophilic pyruvate-TPP attacks the sulfur of the oxidized disulfide which breams the disulfide bond, the bond grabs a proton from an acid

-this means that lipoic acid is now attached to the pyruvate-TPP complex too

what happens now that the lipoic acid disulfide bond has been broken?

-we want to kick the TPP off, so a base grabs a proton from the HO pyruvate part

-the electrons push around to break the TPP ylide off of the lipoic acid and pyruvate part

what happens now that it is just the pyruvate part and lipoic acid?

-this is a thioester

-oxidized

-we have regenerated the ylide

what needs to happen before we go to regenerate the enzyme?

-we need to swap out what is attached to protein/enzyme to what is attached to CoA

-CoA acts as the nucleophile because the carbonyl is an electrophile, a base deprotonates the CoA and it attacks the nucleophile

-this causes another tetrahedral intermediate because the carbonyl grabs a proton to become OH, then the intermediate collapses when a base deprotonates the OH and the electrons push around so that the S of the lipoic acid grabs a proton and that bond breaks

-this leaves us with acetyl CoA and reduced lipoic acid

how do we regenerate the lipoic acid enzyme?

-using FAD

how does FAD regenerate the lipoamide?

-hydride transfer
-a base grabs a proton from one of the reduced sulfur hydrogens

-the electrons push to the bond between sulfurs and to the other H-S bond which attacks the center ring N of FAD

-the electrons push around the ring so that the double bond by the other ring N grabs a proton

-this results in FADH2 (reduced) and lipoamide regenerated (oxidized)

how do we then return FADH2 to FAD?

-using NAD+

-a base grabs a proton from the FADH2 and the electrons push back down around so that the N-H bond attacks the carbon attached to the hydrogen

-this adds a hydrogen to that carbon to form NADH

-we always will regenerate the whole complex

what happens to the NADH made by turning FADH2 → FAD?

-NADH can go to the ETC to make ATP