Abstract
Several γ-glutamyl compounds produced from gamma-glutamyltranspeptidase (GGT) have alluring features for food, pharmaceutical, and biotechnology applications. GGT from Bacillus altitudinis IHB B1644 was cloned, followed by an expression in pET-47b(+) Escherichia coli BL21(DE3). Recombinant GGT (BaGGT) gene subsisted of 1755 bp encoding protein with 585 amino acids, predicted molecular weight, and theoretical pI of 63.3 kDa, and 4.92, respectively. Ectopic expression resulted in inclusion bodies formation at 37 °C. The protein was solubilized and different strategies like dialysis, rapid dilution, and on-column refolding were tried to get active recombinant protein (BaGGT1). To get protein in soluble fraction, GGT was over expressed at low temperature (20 °C), and IPTG concentration (0.025 mM) (BaGGT2). The heterodimeric enzyme consisted of molecular weight of 40, and 22 kDa for large and small subunits, respectively. The specific activities for BaGGT1 and BaGGT2 were 263.9, and 497.45 U mg−1, respectively. BaGGT1 and BaGGT2 showed optimum temperature, and pH at 37 °C and 9, respectively. Km values of 0.8 and 0.2 mM, and Vmax of 666.67 and 333.33 U mg−1 of protein, were calculated for BaGGT1and BaGGT2, respectively. The results demonstrate potential of BaGGT in biosynthesis of γ-glutamyl compounds with industrial application.
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