62 by: Nicholas Simunac

Student – Nicholas Simunac

Name of Enzyme & E.C. number – Pyruvate Dehydrogenase 1.2.4.1

Where is the enzyme found? The enzyme, pyruvate dehydrogenase can be found as a part of the pyruvate dehydrogenase complex in the inner mitochondrial membrane of aerobically metabolizing eukaryotes. It is the first enzyme of 3 total enzymes in this reaction complex.

What does the enzyme do?  The pyruvate dehydrogenase (PDH) enzyme converts incoming pyruvate to hydroxyethyl-TPP, which is then utilized by the other two enzymes in the pyruvate dehydrogenase complex (PDHC). The whole pyruvate dehydrogenase complex is a key part of cellular respiration in which glucose is processed into ATP through a series of reactions and reaction systems. Before delving into the molecular nature of this enzyme’s mechanism, it is worth noting that the PDHC has 5 coenzymes, 1 of which, thiamine diphosphate (TPP), is bound to PDH (Huang & Kuo, 2014). TPP consists of 3 regions, a pyrimidine, a thiazolium ring, and a diphosphate region (NCBI, n.d.). The active site of PDH has 5 key amino acids that play some role in the reaction. Glutamate 59B, which undergoes acid-base chemistry with the thiamine pyrophosphate to activate the coenzyme (Holiday et al, n.d.). Histidine 128B and 271C act as general acids and bases, and Arginine 267C and Tyrosine 281C stabilize the reactive intermediates (Holiday et al, n.d.). The first step is the activation of the thiamine pyrophosphate coenzyme by deprotonation from glutamate 59B which triggers a series of double bond rearrangements and subsequent deprotonation of the thiazolium ring, leading to a stable carbanion formation (Holiday et al, n.d.). This form of thiamine pyrophosphate is the active form, as is how it is found in nature (Huang & Kuo, 2014). When a pyruvate is presented to the thiamine pyrophosphate, the carbanion will perform a nucleophilic attack on the pyruvate’s carbonyl group, leading to a carbon-carbon bond formation between TPP and pyruvate (Holiday et al, n.d.). The oxyanion formed deprotonates histidine 271C to form OH, and the carboxyl group of pyruvate undergoes decarboxylation, ejecting CO2 from the reaction (Holiday et al, n.d.). This is hydroxyethyl-TPP.

The coenzyme must be regenerated, and this is creatively coupled with the transfer of the hydroxyethyl group to the coenzyme, lipoamide from enzyme 2 of this complex. The carbanion of the hydroxyethyl group undergoes a nucleophilic attack of the least substituted sulphur of the lipoamide, where the other sulphur of the 5-membered ring deprotonates histidine 128B, forming a TPP-hydroxyethyl-S-SH bond (Holiday et al, n.d.). The histidine 271C deprotonates the hydroxyethyl group, which oversaturates the central carbon, leading to a bond breakage between TPP and the hydroxyethyl, thus transferring this group over to the lipoamide, forming acyl-lipoate which enzyme 2 of the PDHC uses to make acetyl-COA (Holiday et al, n.d.). If pyruvate metabolism stops, the newly reformed carbanion of the TPP will deprotonate a nearby nitrogen and trigger a double bond rearrangement within TPP, returning itself to its original state. If pyruvate breakdown continues, the TPP will remain in its carbanionic state and attack more pyruvate. Any of the amino acid residues that were deprotonated and didn’t get reprotonated from the reaction will receive protons from water molecules floating nearby (Holiday et al, n.d.).

Any other interesting facts or important information on your enzyme – Pyruvate dehydrogenase is feedback inhibited by elevated levels of all three of ATP, NADH, and acetyl-COA exclusively of each other (Huang & Kuo, 2014). Actual information regarding how this inhibition works is hard to find. Still, given that PDH is the first enzyme in a greater complex, it is essential that it, instead of the other 2 enzymes is inhibited to shut down the rest of the complex and avoid accumulation of byproducts and intermediates (unless the intermediates are desired).

Deficiency in the whole pyruvate dehydrogenase complex leads to severe neurological deficiency due to poor energy output as the body primarily relies on lactic acid fermentation to survive (Medline Genetics n.d.).

References

Hewlett, M. (2021). Pyruvate dehydrogenase. HHMI BioInteractive. https://www.biointeractive.org/classroom-resources/pyruvate-dehydrogenase

Holliday, G. L., Almonacid, D. E., & Bartlett, G. J. (n.d.). Pyruvate dehydrogenase (acetyl-transferring). Mechanism and Catalytic Site Atlas. European Bioinformatics Institute. https://www.ebi.ac.uk/thornton-srv/m-csa/entry/106/

Huang, Y., & Kuo, Y. (2014). Metabolism of selenium: Mechanisms and mechanisms of toxicity. The Journal of Nutritional Biochemistry, 25(5), 465-475. https://doi.org/10.1074/jbc.R114.563148

National Center for Biotechnology Information. (n.d.). Pyruvate dehydrogenase deficiency. MedlinePlus Genetics. https://medlineplus.gov/genetics/condition/pyruvate-dehydrogenase-deficiency/

National Center for Biotechnology Information. (n.d.). 2-(1-hydroxyethyl)thiamine diphosphate(2-). PubChem. https://pubchem.ncbi.nlm.nih.gov/compound/2-_1-Hydroxyethyl_thiamine-diphosphate_2

Vo, L. (n.d.) Pyruvate Dehydrogenase Complex Regulation. BioRender. https://www.biorender.com/template/pyruvate-dehydrogenase-complex-regulation

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