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Determination of three amino acids causing alteration of proteolytic activities of staphylococcal glutamyl endopeptidases.

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Title: Determination of three amino acids causing alteration of proteolytic activities of staphylococcal glutamyl endopeptidases.
Authors: Nemoto, Takayuki K / Ono, Toshio / Shimoyama, Yu / Kimura, Shigenobu / Ohara-Nemoto, Yuko
Issue Date: Mar-2009
Publisher: Walter de Gruyter GmbH & Co. KG
Citation: Biological chemistry, 390(3), pp.277-285; 2009
Abstract: Abstract Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus warneri secrete glutamyl endopeptidases, designated GluV8, GluSE and GluSW, respectively. The order of their protease activities was GluSE<GluSW<<GluV8. The present study investigated the mechanism that causes these differences. Expression of chimeric proteins between GluV8 and GluSE revealed that the difference was primarily attributed to amino acids at residues 170-195, which defined the intrinsic protease activity, and additionally to residues 119-169, which affected the proteolysis sensitivity. Among nine substitutions present in residues 170-195 of the three proteases, the substitutions at positions 185, 188 and 189 were responsible for the changes in their activities; and the combination of W185, V188 and P189, which naturally occurred on GluV8, exerted the highest protease activity. Among them, W185 and P189 were indispensable for the full activity; but V188 could be replaced by hydrophobic amino acids. These three amino acid residues appeared to create a substrate-binding pocket together with the catalytic triad and the N-terminal V1, and therefore, defined the K(m) values of the proteases. This study also describes the way to produce a chimeric form of GluSE and GluV8 that was resistant to proteolysis, and therefore, possessed activity 4-fold higher than that of the wild-type recombinant GluV8.
Keywords: Glutamyl endopeptidase / Staphylococcus aureus / Staphylococcus epidermidis / Substrate-binding pocket / V8 protease
URI: http://hdl.handle.net/10069/23192
ISSN: 14316730
DOI: 10.1515/BC.2009.027
PubMed ID: 19090719
Relational Links: www.degruyter.com/journals/bc
Rights: Walter de Gruyter.
Type: Journal Article
Text Version: author
Appears in Collections:Articles in academic journal

Citable URI : http://hdl.handle.net/10069/23192

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