The enzymatic synthesis of thiamine monophosphate.

The catalysis, by the enzyme thiaminase, of reversible baseexchange reactions between thiamine and aromatic tertiary amines was first reported by Fujita et al. (1). Related findings with mammalian diphosphopyridine nucleotidases (2) and with methionine activation (3) drew attention to the energy-rich nature of “onium” (4-6) compounds and suggested that the alkylation of thiazole must involve activation of the pyrimidine, 2-methyl-4-amino-5-hydroxymethylpyrimidine. That the pyrimidine might be activated by phosphorylation was suggested by the observation of Harris and Yavit (7) that in crude extracts of bakers’ yeast, the phosphate ester of the pyrimidine was a better precursor of thiamine than the pyrimidine compound. In independent studies, Leder (8) and Nose, Ueda, and Kawasaki (9), using synthetic substrates, showed that the reactants were the pyrophosphate ester of the pyrimidine and a phosphorylatecl thiazole and that the initial reaction product was not thiamine, but a phosphorylated form of thiamine subsequently identified (10, 11) as thiamine monophosphate. Camiener and Brown also showed (12) that 2-methyl4amino5-hydrosymethyl pyrimidine monophosphate and a second pyrimidine derivative identified, on presumptive evidence, as the corresponding pyrophosphate ester, were formed enzymatically in partially purified yeast preparations from 2-methyl-4amino5-hydroxymethyl pyrimidine and adenosine triphosphate. Evidence for the sequential formation of these esters by separate enzymes has been reported by Lewin and Brown (13). The results of these studies may be summarized by the following equations: