Genetic Analyses of Thiamin Pyrophosphate Biosynthesis in Escherichia coli

NAKAYAMA Hideo

doi:10.20632/vso.64.11_619

A series of reactions catalyzed by six enzymes participating in the terminal stage of TPP formation in Escherichia coli has been examined by mutant analyses as to whether they are involved as true biosynthetic reactions or not. Mutants auxotrophic for intact form of thiamin produced from JC1552, a prototrophic strain for thiamin, include two types : one lacks phosphomethylpyrimidine kinase, and the other lacks thiamin-phosphate pyrophosphorylase. Mutants lacking each of the remaining four enzymes were produced in stepwise fashion from JC1552, via intermediate production of mutants whose metabolic sequence had been blocked prior to the reaction which was expected to block by the second mutation. We found five new genes for which we propose the designations thiD, thiK, thiL, thiM and thiN, each governs the activities of phospomethylpyrimidine kinase, thiamin kinase, thiamin-phosphate kinase, hydroxyethylthiazole kinase, and hydroxymethylpyrimidine kinase, respectively. By conjugation and P1 transduction they were mapped on the chromosome at about 25 min, 9 min, 46 min, 46 min, and in the 46-51 min region, respectively. When the first block had been deleted from the doubly blocked mutants by transduction, mutants carrying one of the thiK, thiM, and thiN mutations became to prototrophic for thiamin, suggesting that the enzymatic defects caused by the second mutations do not appear to be involved in the de novo synthetic pathway for TPP.

Genetic Analyses of Thiamin Pyrophosphate Biosynthesis in Escherichia coli

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