Lec 11: Thiamine pyrophosphate, glutathione, pentose phosphate pathway

thiamine pyrophosphate function
coenzyme to pyruvate decarboxylase; is a good nucleophile that can attack pyruvate and is needed by pyruvate decarboxylase to get CO2+ acetaldehyde
thiamine pyrophosphate structure
amino pyrimidine ring attached to thiazolium ring attached by 2 carbons to a pyrophosphate; thiazolium ring is where chemistry and nucleophilic attack occurs; pyrophosphate negative charges orient the coenzyme perfectly in the active site
cyanide example
cyanide has a similar functionality to TPP in that it is a very good nucleophile that can attack a carbonyl carbon on pyruvate and facilitate removale of CO2; CN tends to bind to hemes, making it toxic in biological systems but it can be used in a lab for this purpose
how TPP works
TPP binds to active site of pyruvate decarboxylase, Glu 51 acts as an acid catalyst and protonates N on amino pyrimidine ring of TPP which triggers electron rearrangement that allows sterically hindered H to leave which forms a carbanion, carbanion is stabilized by nearby electron sink N+ on thiazolium ring; stabilized carbanion is a good nucleophile that can attack the carbonyl carbon of pyruvate and trigger the release of CO2
why does proton on amino pyrimidine ring want to leave
it is sterically hindered with H on thiazolium ring; Glu 51 acid catalyst rearranges electrons and makes N less able to hold on to H; stable ylid forms which allows carbanion to exist long enough to nuc attack
where is TPP found
pentose phosphate pathway, PDH complex, Krebs cycle all require a very good nucleophile
what makes TPP such a good coenzyme
pyruvate can’t self-eliminate CO2 because the carbanion that results is unstable; pyruvate must be nucleophilically attacked by something as good as a stabilized cation; TPP is stabilized by resonance and the nearby electron sink on N+; negative charge is distributed all around the thiazolium ring
glutathione function
behaves like mercaptoethanol and reduces SH group; it is recycled back to its reduced form by glutathione reductase and NADPH; example: CoA may attach to each other by sulfur group and can only work with a free SH group so they need to be reduced by free-floating glutathione
glutathione structure
glutamate attached by its gamma carbon to cysteine which is attached to a lgycine; the cysteine houses an SH group which is the business end of glutathione that reduces disulfide bonds
glutathione formation
glutamate is bound with ATP by gamma-glutamyl cysteine synthase to make acyl phosphate; acyl phosphate is a high energy intermediate and the alpha amino of cysteine can bind to the synthase active site and displace phosphate; a synthetase uses nucleoside triphosphate to add glycine to cysteine using ATP
pentose phosphate pathway function
produce NADPH and pentose phosphates
why make NADPH
reduces glutathione back to useful form; used in reductive biosynthesis to create fatty acids, sterols, etc
oxidative phase of pentose phosphate pathway
glucose dehydrogenase uses NADP+ to oxidize glucose-6-phosphate to 6- phosphogluconate which is oxidized by NADP+ to create ribulose-5-phosphate which becomes a series of pentose phosphates that can be used in DNA, RNA, coenzymes, etc; as NADP+ is converted to NADPH it is used to reduce glutathione and fatty acid precursors; overall oxidative phase is driven by release of CO2 which increases entropy
why does releasing CO2 increase entropy
CO2 is small, nonpolar, and can easily pass in and out of the cell
nonoxidative phase of pentose phosphate pathway
carbons aren’t lost, just reshuffled between 5 and 6 carbon things; transketolase and transaldolase reversibly convert ribose-6-phosphate to glucose-6-phosphate
takes 2 carbons and moves them to aldose in nonoxidative phase pf pentose phosphate pathway; requires TPP to attack its active site with a carbanion
takes 3 carbons to move onto aldose; uses protonated Schiff base as a nucleophile instead of TPP; both TPP and protonated schiff base are stabilized by resonance and electron sink

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