Separation of Functional Domains for the α-1,4 and α-1,6 Hydrolytic Activities of a Bacillus Amylopullulanase by Limited Proteolysis with Papain
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概要
- 論文の詳細を見る
An amylopullulanase (APase) from alkalophilic Bacillus sp. KSM-1378 hydrolyzes both α-1,6 linkages in pullulan and α-1,4 linkages in other polysaccharides, each being maximally active at an alkaline pH, to generate oligosaccharides. We analyzed proteolytic fragments that were produced by exposing pure APase to various proteases, to identify its catalytic domain(s). The intact, pure 210-kDa APase was partially digested with papain for a short time, yielding simultaneously two smaller non-overlapping active fragments, designated amylose-hydrolyzing fragment (AHF114, 114kDa) and pullulan-hydrolyzing fragment (PHF102,102kDa). The two truncated protein fragments, each containing a single catalytic domain, were purified to homogeneity. The purified AHF114 and PHF102 had similar enzymatic properties to the amylase and pullulanase activities, respectively, of intact APase. The partial amino-terminal sequences of APase and AHF114 were both Glu-Thr-Gly-Asp-Lys-Arg-Ile-Glu-Phe-Ser-Tyr-Glu-Arg-Pro and that of PHF102 was Thr-Val-Pro-Leu-Ala-Leu-Val-Ser-Gly-Glu-Val-Leu-Ser-Asp-Lys-Leu. These results were direct evidence that the α-1,6 and α-1,4 hydrolytic activities were associated with two different active sites in this novel enzyme. Our alkaline APase is obviously a "biheaded enzyme".
- 社団法人日本農芸化学会の論文
- 1996-04-23
著者
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Hagihara H
Tochigi Research Laboratories Kao Corporation
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Ito S
Extremobiosphere Research Center Of Japan Agency For Marine-earth Science And Technology (jamstec):c
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ITO Susumu
Tochigi Research laboratories, Kao Corporation
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Hagihara Hiroshi
Tochigi Research Laboratories, Kao Corporation
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Kobayashi Tohru
Extremobiosphere Research Center of Japan Agency for Marine-Earth Science and Technology (JAMSTEC)
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IGARASHI Kazuaki
Tochigi Research Laboratories of Kao Corporation
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KOBAYASHI Tohru
Tochigi Research Laboratories of Kao Corporation
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IKAWA-KITAYAMA Kaori
Tochigi Research Laboratories of Kao Corporation
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SAWADA Kazuhisa
Tochigi Research Laboratories of Kao Corporation
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SAITO Kazuhiro
Tochigi Research Laboratories of Kao Corporation
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ARA Katsutoshi
Tochigi Research Laboratories of Kao Corporation
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Kawai S
Tochigi Research Laboratories Kao Corporation
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Kobayashi T
Extremobiosphere Research Center Of Japan Agency For Marine-earth Science And Technology (jamstec)
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Igarashi Kaori
Graduate School Of Human Life Sciences Showa Women's University
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Igarashi Kazuei
Graduate School Of Pharmaceutical Sciences Chiba University:amine Pharma Research Institute Innovati
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Kawai Shinya
Graduate School Of Engineering Tokyo University Of Agriculture And Technology
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Igarashi K
Department Of Applied Chemistry And Bioengineering Graduate School Of Engineering Osaka City Univers
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Ito S
National Industrial Research Institute
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Sugimoto Isamu
Division Of Pathological Biochemistry Department Of Biomedical Sciences School Of Life Sciences Facu
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Igarashi Kazuhiko
Graduate School Of Pharmaceutical Sciences Chiba University:amine Pharma Research Institute Innovati
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Kobayashi Tohru
Tochigi Research Laboratories Kao Corporation
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Ito Susumu
Tochigi Research Laboratories Kao Corporation
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Hagihara Hiroshi
Tochigi Research Laboratories Kao Corporation
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