Analyzing the catalytic mechanism of protein tyrosine phosphatase PtpB from Staphylococcus aureus through site-directed mutagenesis

Abstract

Protein tyrosine phosphatase B (PtpB) from Staphylococcus aureus, MRSA 252, is a low molecular weight protein tyrosine phosphatase involved in its pathogenicity. PtpB has been modeled in silico and site-directed mutagenesis performed to ascertain the importance of active site residues Cys8, Arg14, Ser15 and Asp120 in its catalytic mechanism. Kinetic characterization of wild-type and the mutant PtpBs, C8S, R14A, S15T, S15A, D120A, D120E, D120N revealed the reaction mechanism followed by this LMWPTPase. The mutations caused major changes in the local environment resulting in significant decrease of its catalytic activity. Inhibition kinetics for the wild-type enzyme was performed with maleimide and maleimidobutyric acid.