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Molecular Characterization of a dehydroascorbate reductase from Pinus bungeana

Journal of Integrative Plant Biology 2009年第11期51卷 993-1001页

作者： Hai-Ling Yang Ying-Ru Zhao Cai-Ling Wang Zhi-Ling Yang Qing-Yin Zeng Hai Lu College of Life Sciences and Biotechnology Beijing Forestry University Beijing 100083 China State Key Laboratory of Systematic and Evolutionary Botany Institute of Botany the Chinese Academy of Sciences Beijing 100093 China

dehydroascorbate reductase （DHAR） plays a critical role in the ascorbate-glutathione recycling reaction for most higher plants. To date, studies on DHAR in higher plants have focused largely on Arabidopsis and agricultural plants, and there is virtually no information on the molecular characteristics of DHAR in gymnosperms. The present study reports the cloning and characteristics of a DHAR （PbDHAR） from a pine, Pinus bungeana Zucc. ex Endl. The PbDHAR gene encodes a protein of 215 amino acid residues with a calculated molecular mass of 24.26 kDa. The predicted 3-D structure of PbDHAR showed a typical glutathione S-transferase fold. Reverse transcripUon-polymerase chain reaction revealed that the PbDHAR was a constitutive expression gene in P. bungeana. The expression level of PbDHAR mRNA in P. bungeana seedlings did not show significant change under high temperature stress. The recombinant PbDHAR was overexpressed in Escherichia coil following purification with affinity chromatography. The recombinant PbDHAR exhibited enzymatic activity （19.84 ***/min per mg） and high affinity （a Krn of 0.08 mM） towards the substrates dehydroascorbate （DHA）. Moreover, the recombinant PbDHAR was a thermostable enzyme, and retained 77% of its initial activity at 55℃. The present study is the first to provide a detailed molecular characterization of the DHAR in P. bungeana.

关键词： cloning dehydroascorbate reductase enzyme activity gymnosperm.

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An increased Vitamin C content accompanied by an enhanced recycling pathway

An increased Vitamin C content accompanied by an enhanced re...

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中华中医药学会第九届中药鉴定学术会议

作者： Lei Zhang,Zinan Wang,Yang Xia,Wansheng Chen,Kexuan Tang 1 Department of Pharmacognosy,School of Pharmacy,Second Military Medical University, Shanghai,200433,P.R.China 2 State Key Laboratory of Genetic Engineering,School of Life Science, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center,Fudan University,Shanghai 200433,P.R.China 3 Department of pharmacy,Changzheng Hospital,Second Military Medical University, Shanghai 200003,P.R.China

Vitamin C(L-ascorbic acid,AsA) has important antioxidant and metabolic functions in both plants and animals. Once used,ascorbic acid can be regenerated from its oxidized form in a reaction catalyzed by dehydroascorbate reductase (DHAR).A DHAR cDNA from Arabidopsis thaliana was isolated and expressed in *** to examine whether overexpression of DHAR in plants would increase the level of ascorbic acid through improved ascorbate *** transgenic seedlings,DHAR expression was increased up to 1.5 to 5.4 fold,which enhanced foliar ascorbic acid levels 2- to 4.25-fold and ratio of AsA/DHA about 3- to *** addition,the level of glutathione,the reductant used by DHAR,also increased as did its redox *** is suggested that the vitamin C content of plants can be increased through enhanced ascorbate recycling.

关键词： ascorbic acid dehydroascorbate reductase redox state glutathione metabolic engineering

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