Biosynthesis of poly(γ-glutamic acid) in Bacillus subtilis NX-2: Regulation of stereochemical composition of poly(γ-glutamic acid)

Abstract

The stereochemical composition of poly(γ-glutamic acid), γ-PGA, produced by Bacillus subtilis NX-2 could be effected by Mn 2+ in the culture medium. When the concentration of Mn 2+ varied from 0 to 0.09 g/l, the proportion of d-glutamate increased from 18 to 77%. The d-amino acid aminotransferase and the glutamate racemase activity were also analyzed with Mn 2+ concentration changing in this range, the former was stabled at 0.12 ± 0.02 U/mg, while the latter increased from 0.20 to 0.44 U/mg. Moreover, the glutamate racemase activities also increased while adding 0.03 g/l Mn 2+ to cell extracts. These results suggested that different Mn 2+ concentration could change γ-PGA stereochemical composition through regulating glutamate racemase activity, which was different from other γ-PGA producers in species of B. subtilis ever reported. On the other hand, when Mn 2+ concentration was 0.03 g/l, the proportion of d-glutamate of γ-PGA remained approximately 75% during γ-PGA production, this also made it different from other γ-PGA producers. To understand why it was different in γ-PGA stereochemical modulation, the glutamate racemase gene in B. subtilis NX-2 was cloned, and the amino acid sequence was compared with those in B. subtilis 168 and γ-PGA producer B. subtilis IFO3336. There are four different amino acids (Leu2, Ala230, Asn231 and Arg271) in B. subtilis NX-2 glutamate racemase.