Mechanistic Studies on the Single-Turnover Yeast Thiamin Pyrimidine Synthase: Characterization of the Inactive Enzyme
2022
Lai, Rung-Yi | Mondal, Anushree | Fedoseyenko, Dmytro | Begley, Tadhg P.
The eukaryotic thiamin pyrimidine synthase, THI5p, has been identified as a suicidal/single-turnover enzyme that catalyzes the conversion of its active site histidine and lysine-bound pyridoxal phosphate (PLP) to the thiamin pyrimidine (HMP-P). Here we identify the histidine and PLP fragments using bottom-up proteomics and LC-MS analysis. We also identify the active form of the iron cofactor and quantitate the oxygen requirement of the THI5p reaction. This information is integrated into a mechanistic proposal for this remarkable reaction.
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