Thiamine biosynthesis in yeast—evaluation of 4-hydroxy-5-hydroxymethyl-2-methylpyrimidine as a precursor

Abstract

The putative role of 4-hydroxy-5-hydroxymethyl-2-methylpyrimidine 2 as a late precursor in the biosynthesis of thiamine 1 by Saccharomyces cerevisiae has been examined using the deuteriated derivative 2a. To enable measurement of deuterium incorporation into the pyrimidine ring fragment of 1, a procedure for the degradation of 1 to the aniline derivative 6 using thiaminase I from Bacillus thiaminolyticus has been developed. While 4-amino-2-methyl-5-hydroxy[²H₁]methylpyrimidine 3a was incorporated into thiamine, deuterium from hydroxy-pyrimidine 2a was not incorporated to any significant extent, indicating that amination of compound 2 is not a late step in the biosynthesis of the pyrimidine ring.