Identification of Bacillus subtilis men mutants which lack O-succinylbenzoyl-coenzyme A synthetase and dihydroxynaphthoate synthase

Abstract

Menaquinone (vitamin K2)-deficient mutants of B. subtilis, whose growth requirement is satisfied by 1,4-dihydroxy-2-naphthoic acid but not by o-succinylbenzoic acid (OSB), were analyzed for enzymatic defects. Complementation analysis of cell-free extracts of the mutants revealed that there are 2 groups, as already indicated by genetic analysis. The missing enzyme in each group was identified by complementation of the cell-free extracts with o-succinylbenzoyl-CoA synthetase and dihydroxynaphthoate synthase extracted from Mycobacterium phlei. Mutants found to lack dihydroxynaphthoate synthase and which therefore complement with dihydroxynaphthoate synthase of M. phlei, were designated as menB; those lacking o-succinylbenzoyl-CoA synthetase and complementing with o-succinylbenzoyl-CoA synthetase, were designated as menE. The menB mutants RB413 (men-325) and RB415 (men-329), when incubated with [2,3-14C2]OSB, produced only the spirodilactone form of OSB in a reaction that was CoA and ATP dependent.