Plant Biotechnology Reports

The Korean Society of Plant Biotechnology (한국식물생명공학회)
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Characterization of dihydroflavonol 4-reductase cDNA in tea [Camellia sinensis (L.) O. Kuntze]

  • Singh, Kashmir (Biotechnology Division, Institute of Himalayan Bioresource Technology) ;
  • Kumar, Sanjay (Biotechnology Division, Institute of Himalayan Bioresource Technology) ;
  • Yadav, Sudesh Kumar (Biotechnology Division, Institute of Himalayan Bioresource Technology) ;
  • Ahuja, Paramvir Singh (Biotechnology Division, Institute of Himalayan Bioresource Technology)
  • Received : 2008.03.19
  • Accepted : 2008.11.17
  • Published : 2009.02.28
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Abstract

Tea leaves are major source of catechins—antioxidant flavonoids. Dihydroflavonol 4-reductase (DFR, EC 1.1.1.219) is one of the important enzymes that catalyzes the reduction of dihydroflavonols to leucoanthocyanins, a key ''late'' step in the biosynthesis of catechins. This manuscript reports characterization of DFR from tea (CsDFR) that comprised 1,413 bp full-length cDNA with ORF of 1,044 bp (115-1,158) and encoding a protein of 347 amino acids. Sequence comparison of CsDFR with earlier reported DFR sequences in a database indicated conservation of 69-87% among amino acid residues. In silico analysis revealed CsDFR to be a membrane-localized protein with a domain (between 16 and 218 amino acids) resembling the NAD-dependent epimerase/dehydratase family. The theoretical molecular weight and isoelectric point of the deduced amino sequence of CsDFR were 38.67 kDa and 6.22, respectively. Upon expression of CsDFR in E. coli, recombinant protein was found to be functional and showed specific activity of 42.85 nmol $min^{-1}$ mg $protein^{-1}$. Expression of CsDFR was maximum in younger rather than older leaves. Expression was down-regulated in response to drought stress and abscisic acid, unaffected by gibberellic acid treatment, but up-regulated in response to wounding, with concomitant modulation of catechins content. This is the first report of functionality of recombinant CsDFR and its expression in tea.

Keywords

Acknowledgement

Grant : Using functional genomics in plants: development and use of technologies for gene discovery and expression modulation - niche pathway engineering in tea

Supported by : Council of Scientific and Industrial Research (CSIR)

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