Bacterial expression and characterization of starfish phospholipase A2

Kishimura, Hideki; Ojima, Takao; Hayashi, Kenji; Nishita, Kiyoyoshi

Bacterial expression and characterization of starfish phospholipase A2

Kishimura, Hideki; Ojima, Takao; Hayashi, Kenji et al.

Comparative Biochemistry and Physiology Part B Biochemistry and Molecular Biology, 2001, 128(3), 565-573

Permalink : https://hdl.handle.net/2115/16939

このアイテムのアクセス数:1,502件（2026-03-19 07:12 集計）

閲覧可能ファイル

ファイル	フォーマット	サイズ	閲覧回数	説明
CBPB128-3	pdf	577 KB	1,340

論文情報

ファイル出力 EndNote Basic出力 Mendeley出力

アクセス権	open access
URI	https://hdl.handle.net/2115/16939
タイトル	Bacterial expression and characterization of starfish phospholipase A2
著者	Kishimura, Hideki ; Ojima, Takao ; Hayashi, Kenji ; Nishita, Kiyoyoshi
e-Rad_Researcher 50204855 著者名 Kishimura, Hideki
e-Rad_Researcher 30160865 著者名 Ojima, Takao
著者名 Hayashi, Kenji
著者名 Nishita, Kiyoyoshi
言語	英語
キーワード	Asterina pectinifera
	Bacterial expression
	Group I
	Phospholipase A2
	Starfish
	Substrate specificity
	Surface loop
	β-wing
発行日	2001-03
出版者	Elsevier Science Inc.
収録物名	Comparative Biochemistry and Physiology Part B Biochemistry and Molecular Biology
巻(号)	128(3)
ページ	565-573
抄録	Phospholipase A2 (PLA2) from the pyloric ceca of the starfish Asterina pectinifera showed high specific activity and characteristic substrate specificity, compared with commercially available PLA2 from porcine pancreas. To investigate enzymatic properties of the starfish PLA2 in further detail, we constructed a bacterial expression system for the enzyme. The starfish PLA2 cDNA isolated previously (Kishimura et al., 2000b. cDNA cloning and sequencing of phospholipase A2 from the pyloric ceca of the starfish Asterina pectinifera. Comp. Biochem. Physiol. 126B, 579-586) was inserted into the expression plasmid pET-16b and the PLA2 protein was expressed in Escherichia coli BL21 (DE3) by induction with isopropyl-β-(–)-thiogalactopyranoside. The recombinant PLA2 produced as inclusion bodies was dissociated with 8 M urea and 10 mM 2-mercaptoethanol and renatured by dialyzing against 10 mM Tris–HCl buffer (pH 8.0). Renatured PLA2 was purified by subsequent column chromatographies on DEAE–cellulose (DE-52) and Sephadex G-50. Although an N-terminal Ser in the native starfish PLA2 was replaced by an Ala in the recombinant PLA2, the recombinant enzyme showed essentially the same properties as did the native PLA2 with respect to specific activity, substrate specificity, optimum pH and temperature, and Ca2+ requirement.
資源タイプ	学術雑誌論文
出版タイプ	Accepted Manuscript
PISSN	1096-4959
関連情報	PMID 11250552
関連情報	URI http://www.sciencedirect.com/science/journal/10964959
関連情報 (isVersionOf)	出版社版DOI https://doi.org/10.1016/S1096-4959(00)00349-3