Interaction of recombinant form of NADPH-dependent thioredoxin reductase from rice with thioredoxin from two plant and bacterial sources

Maintaining redox homeostasis in the cell is critical for different cellular metabolisms and signal transduction pathways. In plants different molecular mechanisms are involved in maintaining cellular redox homeostasis. NADP/thioredoxin system, consisting of NADPH, NADP-thioredoxin reductase (NTR) and thioredoxin plays important role as electron donor to disulfide bonds in many cellular proteins. In this study, the gene encoding one of NTR isoform from rice, namely OsNTRB was cloned in pET28a as fusion with His6-tag and transferred to Roseta (DE3), a strain of Escherichia coli. Considerable amount of His- OsNTRB was produced after induction of bacteria culture with IPTG and purified using affinity chromatography. Heterologous expression and purification of recombinant form of OsNTRB enabled us to study the interaction of this protein with thioredoxin from barley (HvTrxh1) and E. coli (Ec Trx). The results showed that recombinant form of OsNTRB is active and can reduce both HvTrxh1 and EcTrx in vitro. However the rates of reaction with these two Trx were significantly different.