Isolation, identification and sequence analysis of a thioredoxin h gene, a member of subgroup III of h-type Trxs from grape (Vitis vinifera L. cv. Askari)

Abstract

Thioredoxins (Trxs) are small ubiquitous proteins which play a regulatory role in a variety of cellular processes. In contrast to other organisms, plants have a great number of Trx types, consisting of six well-defined groups: f, m, x, and y in chloroplasts, o in mitochondria, and h mainly in cytosol. A full-length cDNA, designated VvCxxS2, encoding Trx h polypeptide was isolated and cloned from grape (Vitis vinifera L. cv. Askari) berries organ by reverse transcription polymerase chain reaction (RT-PCR). The cDNA was 381bp nucleotides in length with a deduced amino acid of 126 residues, possessing a WCIPS active site, which belongs to the subgroup III of h-type Trxs based on phylogenetic analysis. The calculated molecular mass and the predicted isoelectric point of the deduced polypeptide are 14.25kDa and 4.68, respectively. Nucleotide sequence analysis of genomic DNA fragment of VvCxxS2 gene revealed that this gene possesses two introns at positions identical to the previously sequenced Trx h genes. A modeling analysis indicated that VvCxxS2 shares a common structure with other Trxs, and is preferably reduced by Grx rather than NADPH-dependent thioredoxin reductase (NTR). The deduced protein sequence showed a high similarity to Trx h from other plants, in particular from castor bean (Ricinus communis), Betula pendula and sweet orange (Citrus sinensis). Semiquantitative RT-PCR experiments indicated that the transcripts of VvCxxS2 gene are present in all plant organs and different developmental stages. In addition, the higher expression of the VvCxxS2 gene was observed in berry organ as compared to the other organs.