Cloning, Expression and Characterization of Glutathione S-Transferase from Euphausia superba

A novel glutathione S-transferase gene, with an open reading frame (ORF) of 663 bp, was cloned from Euphausia superba and successfully expressed in Escherichia coli. The purified protein (EsGST), with a molecular weight of approximately 24 kDa, was obtained via Ni²⁺-NTA affinity chromatography. EsGST demonstrated catalytic efficiency (k_cat/K_m) values of 106.36 mM^-1 s^-1 for glutathione (GSH) and 39.11 mM^-1 s^-1 for 1-chloro-2,4-dinitrobenzene (CDNB). The enzyme's optimum reaction temperature was 20 °C, with an optimum pH of 8.0. K⁺, Na⁺, and DTT ions enhanced its catalytic activity, whereas Fe²⁺, Fe³⁺, Al³⁺, Zn²⁺, Co²⁺, Cu²⁺, EDTA, and SDS inhibited it. Compared to the wild-type, the activities of the EsGST-Y105W and EsGST-R118H mutants increased by 1.6-fold and 1.78-fold, respectively. Unlike other glutathione S-transferases, which have optimal temperatures above 25 °C, the EsGST-Y105W mutant demonstrated enhanced cold performance, with its optimal temperature reduced from 20 to 10 °C.