Cloning and expression of a tomato glutathione S- transferase (GST) in Escherichia coli
Glutathione S- transferases (GSTs) fulfill a diverse range of functions in an organism. In a previous study, a putative glutathione S-transferase gene (ShGSTU1) from a wild-type tomato, Solanum habrochaites G1.1560, was identified to be a key gene in pathogen resistant response against powdery mildew in tomato. In this study, ShGSTU1 was cloned into plasmid pET-28a, efficiently expressed in Escherichia coli upon isopropyl-β-D-1-thiogalactopyronoside (IPTG) induction, purified with Ni2+ affinity chromatography and biochemically characterized. The results show that the optimal conditions for the expression of recombinant ShGSTU1 in E. coli were growth under 37°C, and 4-h IPTG induction with 1 mM concentration. About 18.93 mg ShGSTU1 was recovered from 1 g wet bacteria. The recombinant ShGSTU1 exhibited enzymatic activity with specific activity 0.625 U/mg. These results might provide a significant foundation for the later research on the mechanism of ShGSTU1 in tomato resistance to powdery mildew.
Key words: Tomato, glutathione S-transferase, expression, purification, enzyme activity.