Characterization of a New (R)-Hydroxynitrile Lyase from the Japanese Apricot Prunus mume and cDNA Cloning and Secretory Expression of One of the Isozymes in Pichia pastoris

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概要

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• PmHNL, a hydroxynitrile lyase from Japanese apricot ume (Prunus mume) seed was purified to homogeneity by ammonium sulfate fractionation and chromatographic steps. The purified enzyme was a monomer with molecular mass of 58 kDa. It was a flavoprotein similar to other hydroxynitrile lyases of the Rosaceae family. It was active over a broad temperature, and pH range. The N-terminal amino acid sequence (20 amino acids) was identical with that of the enzyme from almond (Prunus dulcis). Based on the N-terminal sequence of the purified enzyme and the conserved amino acid sequences of the enzymes from Pr. dulcis, inverse PCR method was used for cloning of a putative PmHNL (PmHNL2) gene from a Pr. mume seedling. Then the cDNA for the enzyme was cloned. The deduced amino acid sequence was found to be highly similar (95%) to that of an enzyme from Pr. serotina, isozyme 2. The recombinant Pichia pastoris transformed with the PmHNL2 gene secreted an active enzyme in glycosylated form.

• 2011-02-23

著者

• Sakai Yasuyoshi

  Division Of Applied Life Sciences Graduate School Of Agriculture Kyoto University

• Yurimoto Hiroya

  Division Of Applied Life Sciences Graduate School Of Agriculture Kyoto University

• Asano Y

  National Inst. Genetics Shizuoka

• Fukuta Yasuhisa

  Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University

• Komeda Hidenobu

  Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University

• KATO YASUO

  Biotechnology Research Center, Faculty of Engineering, Toyama Prefectural University

• Asano Yasuhisa

  Biotechnology Res. Center And Dep. Of Biotechnology Toyama Prefectural Univ.

• Komeda Hidenobu

  Biotechnology Res. Center And Dep. Of Biotechnology Toyama Prefectural Univ.

• Komeda Hidenobu

  Biotechnology Research Center Toyama Prefectural University

• Nanda Samiko

  Biotechnology Research Center And Department Of Biotechnology Toyama Prefectural University

• Fukuta Yasuhisa

  Biotechnology Research Center And Department Of Biotechnology Toyama Prefectural University

• Kato Yasuo

  Biotechnology Research Center And Department Of Biotechnology Toyama Prefectural University

• Nanda Samik

  Biotechnology Research Center and Department of Biotechnology, Toyama Prefectural University

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