Cloning, expression and characterization of γ-glutamyltranspeptidase from escherichia coli and bacillus subtilis

The enzyme γ-glutamyltranspeptidase is the key enzyme involved in glutathione metabolism, and has wide applications in biocatalysis. In this paper, the GGTs from Escherichia coli and Bacillus. subtilis were cloned, recombined and expressed. The catalytic properties and stabilities of the recombined GGTs from E. coli (rE_GGT) and B. subtilis (rB_GGT) were investigated, respectively. The results show that, although the affinity of rE_GGT toward GpNA (K_m) is higher than that of rB_GGT, the catalytic constant of rB_GGT (k_cat) is as high as 3.48×10⁵ s^-1, which is 20 folds as high as that of rE_GGT. Stability studies reveal that rE_GGT is more stable than rB_GGT under the condition of pH 6~9 and T<45°C. While the temperature is above 45°C, rB_GGT is more stable. The studies for the amino acid composition and subunit interface interactions of r_EGGT and rB_GGT show that the rE_GGT has more hydrophobic bonds to help stabilizing its quaternary structure and to make it more stable under moderate temperature. The percentages of charged and aromatic residue in rB_GGT are higher than that in rE_GGT, which is helpful to improve the stability of its secondary structure.