A poly-gamma-glutamic acid (gamma-PGA) productive strain, halotolerant bacterium WX-02 was isolated from the saline soil of China (Yingcheng). By physiological, biochemical, and 16S rDNA sequence analysis methods, the strain was identified as Bacillus licheniformis. The effect of NaCl concentration on gamma-PGA production by WX-02 was investigated in modified E (ME) medium. It was found that the gamma-PGA production was salt-inducible, and the highest volumetric yield of gamma-PGA (13.86 g/l) was attained with 8% of NaCl. It was also observed that the molecular size of gamma-PGA decreased when the NaCl concentration increased. This was the first report of isolation and identification of a gamma-PGA productive strain, halotolerant B. licheniformis. This study provided a simple strategy for controlling the yield and molecular size of gamma-PGA by WX-02.

BACKGROUND The poly‐γ‐glutamic acid (γ‐PGA) hydrolase of PgdS affects the molecular weight and yield of γ‐PGA in Bacillus subtilis, while the effects of PgdS on the γ‐PGA synthesis in Bacillus licheniformis have not been investigated. RESULTS By heterologous expression in Escherichia coli, the PgdS was characterized. The purified PgdS had an optimal γ‐PGA degradation activity from pH 5.5 to 6.5, with an appropriate reaction temperature from 37 °C to 45 °C. The enzyme activity could be enhanced by Zn2+, Mn2+, Ca2+ and SDS, while inhibited by Hg2+ or Cu2+. By enhanced expression of pgdS gene in vivo, the enzyme activity, protein concentration and gene transcriptional level of PgdS were improved, and γ‐PGA molecular weight decreased from 1000–1200 kDa to 600–800 kDa. Moreover, the γ‐PGA yield reached 20.16 g L‐1, increased by 54% than the native strain (13.11 g L−1). Gene transcript levels of glutamate transporters and poly‐γ‐glutamate synthetase were confirmed to be improved, which might be the reason for the increased γ‐PGA production. CONCLUSION Enhanced expression of pgdS gene in B. licheniformis WX‐02 helped to reduce the molecular weight and improve the yield of γ‐PGA. This study provided a novel strategy to regulate the molecular weight and yield of γ‐PGA. © 2013 Society of Chemical Industry

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The glr gene, which encodes glutamate racemase involved in the conversion of L-glutamic acid to its D-isomer, was cloned and expressed in Bacillus licheniformis WX-02. Overexpression of the glr gene not only increased the production of poly-γ-glutamic acid (γ-PGA) by 22.5% but also increased the proportion of D-glutamate in γ-PGA from 77 to 85%. The activity of glutamate racemase was higher than in the original strain throughout cultivation. This is the first report that overexpression of the glr gene could enhance the L- and D-glutamate conversion in B. licheniformis WX-02 and increase the proportion of D-glutamate in γ-PGA and the yield of γ-PGA.

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