Comparison of Reduction Activity of three Rice (Oryza sativa) Thioredoxin Isoforms in Reaction with Insulin

Document Type : Research Paper

Authors

Abstract

Thioredoxins (Trxs) are ubiquitous, low-molecular-mass proteins with two cysteins in their active site WC(G/P)PC which are involved in reversible reduction of disulfide bonds. Plants contain several forms of Trxs. Thioredoxin f, m, x and y are found in the chloroplast, Trxo is localized in mitochondria and Trxh is typically cytosolic. Trxh isoforms play important roles in plants development. In this study, we describe isolation and cloning of three cDNAs encoding different Trx h isoforms, stated OsTrx1, OsTrx20 and OsTrx23. Three Trx h were heterologously expressed in Rosetta (DE3) - a strain of Escherichia coli. After inducing T7 promoter with IPTG, proper amount of recombinant proteins were produced. The recombinant proteins were purified by affinity chromatography. The recombinant Trxh isoforms were analyzed for their ability to reduce Insulin – an electron receptor substrate for Trxs- and DTT, as a primary reducing agent. All three isoforms were active, but the rate of reduction varied among different reactions. The existence of observed variations can reveal different functions for thioredoxins in plant system.
 

Keywords


: Rice, Thioredoxin, Cloning, Heterologous protein expression, Insulin, Reduction activity.  

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