Determination of three amino acids causing alteration of proteolytic activities of staphylococcal glutamyl endopeptidases
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Takayuki K. Nemoto
Abstract
Staphylococcus aureus, Staphylococcus epidermidis, and Staphylococcus warneri secrete glutamyl endopeptidases, designated GluV8, GluSE, and GluSW, respectively. The order of their protease activities is GluSE<GluSW<<GluV8. In the present study, we investigated the mechanism that causes these differences. Expression of chimeric proteins between GluV8 and GluSE revealed that the difference is primarily attributed to amino acid residues 170–195, which define the intrinsic protease activity, and additionally to residues 119–169, which affect the proteolytic 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 occurs in GluV8, exerts the highest protease activity. W185 and P189 were indispensable for full activity, but V188 could be replaced by hydrophobic amino acids. These three amino acid residues appear to create a substrate-binding pocket together with the catalytic triad and the N-terminal V1, and therefore define the Km values of the proteases. We also describe a method to produce a chimeric form of GluSE and GluV8 that is resistant to proteolysis, and therefore possesses 4-fold higher activity than the wild-type recombinant GluV8.
©2009 by Walter de Gruyter Berlin New York
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Articles in the same Issue
- Review
- Glutathione dysregulation and the etiology and progression of human diseases
- Protein Structure and Function
- Interaction of the porcine reproductive and respiratory syndrome virus nucleocapsid protein with the inhibitor of MyoD family-a domain-containing protein
- Role of catalytic and non-catalytic subsite residues in ribonuclease activity of human eosinophil-derived neurotoxin
- Cell Biology and Signaling
- Expression and localization of atypical PKC isoforms in liver parenchymal cells
- Regulation of phosphoenolpyruvate carboxylase in PVYNTN-infected tobacco plants
- The influence of linoleic and linolenic acid on the activity and intracellular localisation of phospholipase D in COS-1 cells
- Proteolysis
- Two aspartic proteinases secreted by the pathogenic yeast Candida parapsilosis differ in expression pattern and catalytic properties
- The assembly and activation of kinin-forming systems on the surface of human U-937 macrophage-like cells
- Determination of three amino acids causing alteration of proteolytic activities of staphylococcal glutamyl endopeptidases
