Hyperexpression of the Gene for a Bacillus α-Amylase in Bacillus subtilis Cells : Enzymatic Properties and Crystallization of the Recombinant Enzyme

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概要

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• We have constructed a new excretion vector, pHSP64,to develop a hyperexcretion system for Bacillus subtilis [Sumitomo et al., Biosci. Biotech. Biochem., 59,2172-2175 (1995)]. The structural gene for a novel liquefying semi-alkaline α-amylase from the alkaliphilic Bacillus sp. KSM-1378 was amplified by PCR. It was cloned into a SalI-SmaI site of pHSP64 and the recombinant plasmid obtained was introduced into B. subtilis. The transformed B. subtilis hyperproduced the α-amylase activity extracellularly, corresponding to approximately 1.0g (5×10^6 units) per liter of an optimized liquid culture. The recombinant enzyme was purified to homogeneity by a simple purification procedure with very high yield. No significant differences in physicochemical and catalytic properties were observed between the recombinant enzyme and the native enzyme produced by Bacillus sp. KSM-1378. The enzymatic properties of the recombinant enzyme were further examined with respect to the responses to various metal ions. The recombinant enzyme could easily be crystallized at room temperature within one day in a buffered solution of 10% (w/v) ammonium sulfate (pH 6.5).

• 社団法人日本農芸化学会の論文
• 1998-09-23

著者

• Hatada Yuji

  Tochigi Research Laboratories Of Kao Corporation

• Ozawa T

  Univ. Tsukuba Ibaraki Jpn

• Hayashi Y

  Research Institute For Food Science Kyoto University

• Hagihara H

  Tochigi Research Laboratories Kao Corporation

• Ito S

  Extremobiosphere Research Center Of Japan Agency For Marine-earth Science And Technology (jamstec):c

• ITO Susumu

  Tochigi Research laboratories, Kao Corporation

• Araki Hiroyuki

  Tochigi Research Laboratories, Kao Corporation

• Hagihara Hiroshi

  Tochigi Research Laboratories, Kao Corporation

• Tsujino Yukiharu

  Tochigi Research Laboratories, Kao Corporation

• Ozaki Katsuya

  Tochigi Research Laboratories, Kao Corporation

• Kobayashi Tohru

  Extremobiosphere Research Center of Japan Agency for Marine-Earth Science and Technology (JAMSTEC)

• IGARASHI Kazuaki

  Tochigi Research Laboratories of Kao Corporation

• KOBAYASHI Tohru

  Tochigi Research Laboratories of Kao Corporation

• IKAWA-KITAYAMA Kaori

  Tochigi Research Laboratories of Kao Corporation

• HAKAMADA Yoshihiro

  Tochigi Research Laboratories of Kao Corporation

• OZAWA Tadahiro

  Tochigi Research Laboratories of Kao Corporation

• HAYASHI Yasuhiro

  Tochigi Research Laboratories of Kao Corporation

• IKAWA Kaori

  Tochigi Research Laboratories of Kao Corporation

• Kobayashi T

  Extremobiosphere Research Center Of Japan Agency For Marine-earth Science And Technology (jamstec)

• Igarashi Kaori

  Graduate School Of Human Life Sciences Showa Women's University

• Ozawa T

  Tochigi Res. Lab. Kao Corp. Tochigi Jpn

• Igarashi Kazuei

  Graduate School Of Pharmaceutical Sciences Chiba University:amine Pharma Research Institute Innovati

• Igarashi K

  Department Of Applied Chemistry And Bioengineering Graduate School Of Engineering Osaka City Univers

• Ito S

  National Industrial Research Institute

• Ozaki Katsuya

  Tochigi Research Laboratories Of Kao Corporation

• Hayashi Y

  Department Of Medicinal Chemistry Center For Frontier Research In Medicinal Science Kyoto Pharmaceut

• Tsujino Y

  Tochigi Research Laboratories Kao Corporation

• Araki Hiroyuki

  Tochigi Research Laboratories Kao Corporation

• Sugimoto Isamu

  Division Of Pathological Biochemistry Department Of Biomedical Sciences School Of Life Sciences Facu

• Araki H

  Tochigi Research Laboratories Kao Corporation

• Igarashi Kazuhiko

  Graduate School Of Pharmaceutical Sciences Chiba University:amine Pharma Research Institute Innovati

• Kobayashi Tohru

  Tochigi Research Laboratories Kao Corporation

• Ito Susumu

  Tochigi Research Laboratories Kao Corporation

• Hagihara Hiroshi

  Tochigi Research Laboratories Kao Corporation

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