The dimerization of Staphylococcus aureus sortase A on cell membrane

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The dimerization of Staphylococcus aureus sortase A on cell membrane

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Title: The dimerization of Staphylococcus aureus sortase A on cell membrane
Author: Zhu, Jie, 1980-
Abstract: Staphylococcus aureus sortase A (SrtA) transpeptidase is a prominent membrane bound virulence factor in gram-positive bacteria, which organizes the peptidoglycan cell wall of the organism. Here, we report the first direct observation of the self-association behavior of SrtA. Formation of a SrtA dimer is highly selective in vitro in E. coli and in vivo on the S. aureus cell membrane. Quantitative analysis of protein binding affinity indicated a moderate association between two SrtA molecules with an apparent K[subscript d] of about 55 [micrometres] in vitro. Furthermore, to address the importance of dimerization for enzyme function, site-directed mutagenesis on potential target residues was performed to generate monomer only SrtA mutant proteins to completely disrupt dimer formation both in vitro and in vivo. Finally, an in vivo activity assay was performed to evaluate the function of SrtA wild type protein as well as its monomer only mutants. Our data demonstrated that S. aureus cells expressing mutant SrtA in a monomer only form are more successful at invading human epithelial cells than those expressing wild type SrtA in dimer-monomer equilibrium. It suggested that the monomeric form of SrtA is more active than the dimeric enzyme. We also demonstrated the uniqueness of SrtA dimerization by identifying that at least one other sortase family protein, SrtB only exists in monomer form. SrtA dimerization may have significant implications for understanding its biological function at both the cellular and molecular levels, which will lead to the development of new anti-infective therapies against gram-positive pathogens.
Department: Cellular and Molecular Biology
Subject: Staphylococcus aureus Sortase Membrane protein Protein-protein interaction SrtA Dimeric enzymes
URI: http://hdl.handle.net/2152/17327
Date: 2010-05

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