PRODUCTION OF ORYZACYSTATIN-I AND ORYZACYSTATIN-II IN ESCHERICHIA-COLI USING THE GLUTATHIONE-S-TRANSFERASE GENE FUSION SYSTEM

In recent years, several studies have demonstrated the potential of pr oteinase inhibitors for the biocontrol of insect pests. For such a con trol approach, however, strategies must be developed for the simple an d efficient production of active inhibitors. In this study, oryzacysta tins I (OCI) and II (OCII) were produced in Escherichia coli using the glutathione S-transferase (GST) gene fusion system. Both inhibitors w ere produced in large amounts as fusion products (similar to 100 mg/L E. coli culture) and were apparently stable when accumulated in bacter ial cells. Enzyme specificities and inhibition constants of the fusion proteins were similar to those determined for free inhibitors and tho se reported for naturally occurring inhibitors isolated from rice, pre venting the necessity of using an expensive and time-consuming cleavag e step for the obtention of active OCs. The GST system thus appears ap propriate for simple and efficient large-scale production of the two c ysteine proteinase inhibitors.