Production of recombinant form of rice Glutaredoxin in Escherichia coli

Glutaredoxins (Grxs) are low-molecular-mass proteins with two cysteins in their active site which are involved in reversible reduction of disulfide bonds. The reduction of Grxs is catalyzed by glutathioen reductase via glithathione. Plants contain several isoforms of Grx. In this study we aim to heterologously express OsGrx9 in E.coli. To this end the gene encoding OsGrx9 was cloned in pET28a. The new construct pET28a-OsGrx9 was transformed to Rosetta (DE3). After inducing T7 promoter with IPTG, proper amount of recombinant proteins were produced in soluble fraction. The recombinant proteins were purified by affinity chromatography and was analyzed for its ability to reduce Insulin – an electron receptor substrate for Grxs. For this reaction we used DTT or GSH as reducing agent. The results show that OsGrx9 is able to reduce insulin in vitro and the rate of insulin reduction is pH-dependent.