Biosynthesis of thiamin thiazole in eukaryotes: conversion of NAD to an advanced intermediate

J Am Chem Soc. 2007 Mar 14;129(10):2914-22. doi: 10.1021/ja067606t. Epub 2007 Feb 20.


Thiazole synthase catalyzes the formation of the thiazole moiety of thiamin pyrophosphate. The enzyme from Saccharomyces cerevisiae (THI4) copurifies with a set of strongly bound adenylated metabolites. One of them has been characterized as the ADP adduct of 5-(2-hydroxyethyl)-4-methylthiazole-2-carboxylic acid. Attempts toward yielding active wild-type THI4 by releasing protein-bound metabolites have failed so far. Here, we describe the identification and characterization of two partially active mutants (C204A and H200N) of THI4. Both mutants catalyzed the release of the nicotinamide moiety from NAD to produce ADP-ribose, which was further converted to ADP-ribulose. In the presence of glycine, both the mutants catalyzed the formation of an advanced intermediate. The intermediate was trapped with ortho-phenylenediamine, yielding a stable quinoxaline derivative, which was characterized by NMR spectroscopy and ESI-MS. These observations confirm NAD as the substrate for THI4 and elucidate the early steps of this unique biosynthesis of the thiazole moiety of thiamin in eukaryotes.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenosine Diphosphate / metabolism
  • Adenosine Diphosphate Ribose / metabolism
  • Eukaryotic Cells / metabolism*
  • NAD / metabolism*
  • Saccharomyces cerevisiae / enzymology
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Substrate Specificity
  • Thiamine / metabolism*
  • Thiamine Pyrophosphate
  • Thiazoles / metabolism*


  • Saccharomyces cerevisiae Proteins
  • THI4 protein, S cerevisiae
  • Thiazoles
  • NAD
  • Adenosine Diphosphate Ribose
  • Adenosine Diphosphate
  • Thiamine Pyrophosphate
  • Thiamine