Plant Diversity ›› 2010, Vol. 32 ›› Issue (01): 32-40.DOI: 10.3724/SP.J.1143.2010.09182

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Prokaryotic Expression and Bioinformatics Analysis of Cytosolic Malate Dehydrogenase from Camellia sinensis(Theaceae)

CHEN LuLu1, ZHAI YuJia1, WANG Peng 1, Wang Hui1,2, HAO JiaSheng1,2, ZHU GuoPing1,2   

  1. 1 The Key Laboratory of Molecular Evolution and Biodiversity, College of Life Sciences, Anhui Normal University, Wuhu 241000, China; 2Anhui Key Laboratory of Biotic Environment and Ecological Security, Wuhu 241000, China
  • Received:2009-09-14 Online:2010-02-25 Published:2010-02-25
  • Contact: ZHU GuoPing

Abstract: The complete gene of cytosolic malate dehydrogenase (cMDH) from Camellia sinensis, called CscMDH, was obtained by RTPCR and rapid amplification of cDNA ends (GenBank accession number GQ845406). This gene was 1 235 bp in length, encoding a protein of 332 amino acids with the putative molecular weight of 355 kD. The Ecoli Rosetta (DE3) harboring pGEXMDH was induced by 05 mmol·L1 IPTG at 32℃ for 3 hours, and a 615 kD glutathione Stransferase (GST)fused MDH was obtained in soluble form. The results of NCBIBLAST revealed that CscMDH shared 88%-93% of amino acid sequence identity with other cMDH from different higher plants. According to the multiple sequence alignment based on the threedimensional structure of protein, CscMDH was predicted to be a dimer with thirteen βsheet and thirteen αhelix of each subunit. CscMDH contains typical fingerprint sequence (G12AAGQIG18) as all MDHs. The amino acid D43 in CscMDH is conserved in all NADMDHs. CscMDH also has some conserved sequence units homologous to other NADMDHs, such as NAD+ binding sites, catalytic motif and substrate binding sites. Moreover, CscMDH contains six Cys which are highly conserved in all plant NADcMDHs. Therefore, CscMDH was inferred to be NADdependent cMDH. The present study may provide the fundament for the further functional characterization of CscMDH.

Key words: Camellia sinensis

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