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Specificity of the medaka enteropeptidase serine protease and its usefulness as a biotechnological tool for fusion-protein cleavage

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タイトル: Specificity of the medaka enteropeptidase serine protease and its usefulness as a biotechnological tool for fusion-protein cleavage
著者: Ogiwara, Katsueki 著作を一覧する
Takahashi, Takayuki 著作を一覧する
キーワード: enteropeptidase
cleavage specificity
biotechnology tool
medaka fish
発行日: 2007年 4月24日
出版者: The National Academy of Sciences of the United States of America
誌名: Proceedings of the National Academy of Sciencesof the United States of America
巻: 104
号: 17
開始ページ: 7021
終了ページ: 7026
出版社 DOI: 10.1073/pnas.0610447104
抄録: We cloned two distinct cDNAs for enteropeptidase (EP) from the intestine of the medaka, Oryzias latipes, which is a small freshwater teleost. The mRNAs code for EP-1 (1036 residues) and EP-2 (1043 residues), both of which have a unique, conserved domain structure of the N-terminal heavy chain and C-terminal catalytic serine protease light chain. When compared with mammalian EP serine proteases, the medaka enzyme exhibited extremely low amidolytic activity for small synthetic peptide substrates. Twelve mutated forms of the medaka EP protease were produced by site-directed mutagenesis. Among them, one mutant protease, E173A, was found to have considerably reduced nonspecific hydrolytic activities both for synthetic and protein substrates without serious reduction of its Asp-Asp-Asp-Asp-Lys (D4K)-cleavage activity. For the cleavage of fusion proteins containing a D4K-cleavage site, the medaka EP proteases were shown to have advantages over their mammalian counterparts. Based on our present data, we propose that the E173A mutant is the most appropriate protease to specifically cleave proteins containing the D4K cleavage sequence.
資料タイプ: article (author version)
URI: http://hdl.handle.net/2115/30134
出現コレクション:雑誌発表論文等 (Peer-reviewed Journal Articles, etc)

提供者: 高橋 孝行

 

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