Molecular Cloning and Characterization of a Thermostable Carboxylesterase from an Archaeon, Sulfolobus shibatae DSM5389 : Non-Linear Kinetic Behavior of a Hormone-Sensitive Lipase Family Enzyme(ENZYMOLOGY, PROTEIN ENGINEERING, AND ENZYME TECHNOLOGY)

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概要

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• A gene coding for an esterase (SshEstI, 915 bp in length) of the thermoacidophilic archaeon Sulfolobus shibatae DSM5389 was cloned, sequenced, and overexpressed in Escherichia coli JM109 cells as a soluble, catalytically active protein. The deduced amino acid sequence of SshEstI was consistent with a protein containing 305 amino acid residues with a molecular mass of 33 kDa. Sequence comparison studies indicated that SshEstI could be a member of the hormone-sensitive lipase family, in that it had the highest sequence similarity to esterases from Sulfolobus solfataricus (90% identity) and Archaeoglobus fulgidus (42%) and a lipase from Pseudomonas sp. B11-1 (38%). The recombinant enzyme was highly thermostable and retained more than 70% of its initial activity after incubation at 90℃ and pH 7.0 for 30 min. The recombinant enzyme catalyzed the hydrolysis of P-nitrophenyl (p-NP) esters with C_2-C_<16> acyl chains but not the hydrolysis of triacylglycerides such as tributyrin and triolein. The enzymatic hydrolysis of p-NP acetate proceeded in a linear manner with time, whereas that of p-NP esters with acyl chains of C_5 or longer showed a biphasic profile, where a rapid release of p-nitrophenol (〜3 min) was followed by a slow, sustained release. These non-linear kinetics may be explained in terms of a very slow, presteadystate burst phenomenon of p-nitrophenol release or a hysteretic behavior of SshEstI with these substrates.

• 社団法人日本生物工学会の論文
• 2004-12-25

著者

• Nishino Tokuzo

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• Nakayama Toru

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• Nishino T

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Nishino Tokuzo

  Dep. Of Biochemistry And Engineering Graduate School Of Engineering Tohoku Univ. Aoba-yama Sendai Mi

• Hemmi H

  Tohoku Univ. Miyagi Jpn

• Hemmi Hisashi

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Hemmi Hisashi

  Department Of Biochemistry And Engineering Tohoku University

• Hemmi Hisashi

  Dep. Of Biochemistry And Engineering Graduate School Of Engineering Tohoku Univ. Aoba-yama Sendai Mi

• EJIMA KOKI

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• LIU JIAN

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• OSHIMA YASUHIRO

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• HIROOKA KAZUTAKE

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• SHIMANUKI SHOKO

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• YOKOTA YUKAKO

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• OHSHIMA Yasuhiro

  Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University

• Nakayama Toru

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Oshima Yasuhiro

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Hamada Hiroaki

  Department Of Life Science Faculty Of Science Okayama University Of Science

• Nishino Tokuzo

  Department Of Biochemistry And Engineering Faculty Of Engineering Tohoku University

• Ohshima Yasuhiro

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Liu J

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Liu Jian

  Department Of Anesthesiology Plastic Surgery Hospital Chinese Academy Of Medical Sciences And Peking

• Hemmi Hisashi

  Department Of Applied Molecular Biosciences Graduate School Of Bioagricultural Sciences Nagoya Unive

• Ejima Koki

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Yokota Yukako

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Hirooka Kazutake

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• Shimanuki Shoko

  Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University

• NAKAYAMA Toru

  Department of Applied Microbial Technology, The Kumamoto Institute of Technology:(Present office)Institute for Fundamental Research, Suntory Ltd.

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