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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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Please use this identifier to cite or link to this item:http://hdl.handle.net/2115/30134

Title: Specificity of the medaka enteropeptidase serine protease and its usefulness as a biotechnological tool for fusion-protein cleavage
Authors: Ogiwara, Katsueki Browse this author
Takahashi, Takayuki Browse this author →KAKEN DB
Keywords: enteropeptidase
cleavage specificity
biotechnology tool
medaka fish
Issue Date: 24-Apr-2007
Publisher: The National Academy of Sciences of the United States of America
Journal Title: Proceedings of the National Academy of Sciencesof the United States of America
Volume: 104
Issue: 17
Start Page: 7021
End Page: 7026
Publisher DOI: 10.1073/pnas.0610447104
PMID: 17438297
Abstract: 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.
Type: article (author version)
URI: http://hdl.handle.net/2115/30134
Appears in Collections:生命科学院・先端生命科学研究院 (Graduate School of Life Science / Faculty of Advanced Life Science) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)

Submitter: 高橋 孝行

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