Summary: A 1·7 kb DNA fragment isolated from an genomic library was able to complement the thiamin requirement of strains carrying the and mutations. The three genes encode hydroxyethylthiazole kinase, hydroxymethylpyrimidine (HMP) kinase and phosphomethylpyrimidine (HMP-P) kinase, respectively. Sequence analysis revealed that the 1·7 kb fragment contained two ORFs of 708 bp and 801 bp. The former ORF complemented the mutation and the latter ORF both the and mutations. The latter ORF was cloned into the expression vector pET3a, and the encoded protein was purified through three successive column chromatographies. The purified protein was able to convert HMP to its monophosphate and the monophosphate to its pyrophosphate. These results suggest that the two distinct enzyme activities, HMP kinase and HMP-P kinase, are indeed a bifunctional enzyme encoded by a single gene, designated


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