Cloning and Expression of the Glutamate Racemase Gene of Bacillus pumilus

Abstract

A glutamate racemase gene (murI) was found in Bacillus pumilus cells and cloned into Escherichia coli WM335, a D-glutamate auxotroph, by means of a genetic complement method. MurI of B. pumilus encodes a 272-amino acid protein with an unusual initiation codon, TTG. The deduced amino acid sequence shows significant similarity with those of glutamate racemases from E. coli (ratio of identical residues, 28%), Pediococcus pentosaceus (44%), and Staphylococcus haemolyticus (49%). B. pumilus MurI was expressed as a fusion protein connected to the N-terminal 12 residues of β-galactosidase; the fusion protein showed glutamate racemase activity, and resembled the enzyme of P. pentosaceus in physicochemical and enzymological properties.