The protein tyrosine phosphatase (PTP) SHP-1 plays an important role in both immune regulation and oncogenesis. This enzyme is part of a broader family of PTPs that all play important regulatory roles in vivo. Common to these enzymes is a highly conserved aspartic acid (D421 in SHP-1) that acts as an acid/base catalyst during the PTP-catalyzed reaction. This residue is located on a mobile loop, the WPD-loop, the dynamic behavior of which is intimately connected to the catalytic activity. The SHP-1 WPD-loop variants H422Q, E427A, and S418A have been kinetically characterized and compared to those of the wild-type (WT) enzyme. These variants exhibit limiting magnitudes of k cat ranging from 43 to 77% of the WT enzyme. However, their pH profiles are significantly broadened in the basic pH range. As a result, above pH 6, the E427A and S418A variants have turnover numbers notably higher than those of WT SHP-1. Molecular modeling results indicate that the shifted pH dependencies result primarily from changes in solvation and hydrogen-bonding networks that affect the pK a of the D421 residue, explaining the changes in pH-rate profiles for k cat on the basic side. In contrast, a previous study of a noncatalytic residue variant of the PTP YopH, which also exhibited changes in pH dependency, showed that the catalytic change arose from mutation-induced changes in conformational equilibria of the WPD-loop. This finding and the present study show the existence of distinct strategies for nature to tune the activity of PTPs in particular environments through controlling the pH dependency of catalysis.
© 2024 The Authors. Published by American Chemical Society.