Cloning, heterologous expression and characterization of three thioredoxin h isoforms (OsTrx1, OsTrx20 and OsTrx23) from rice

Abstract

Thioredoxin h (Trx h) is a major cytoplamic and mitochondrial disulfide reductase. In plants, Trx h isoforms are encoded by a multigenic family of genes. The multiplicity of these isoforms raises the question of their functional specificity. In this study, we describe isolation and cloning of three cDNAs encoding different Trx h isoforms, namely OsTrx1, OsTrx20 and OsTrx23. Three Trx h were heterologously expressed in Escherichia coli and their activities were compared using DTT-dependent insulin assay. OsTrx23 and OsTrx1 demonstrated highest (0.05 Δ650/min) and lowest (0.016 Δ650/min) activity, respectively. In contrast to OsTrx1 and OsTrx20 isoforms, OsTrx23 was efficiently reduced by NADPH-dependent thioredoxin reductase from barley (HvNTR2). The gene expression of three Trx h was analyzed in one, two and three-week old rice seedlings. The transcripts of OsTrx23 with high intensity and OsTrx20 with low intensity were expressed in both root and shoot, whereas OsTrx1 was only expressed in root. All of three isoforms were appeared partially dimerized under non-reducing conditions suggesting that disulfide bridges were responsible for dimerization.