For each system, we evluated the root mean square displacement (RMSD) of both the Cα-atoms of just the WPD loop of the different SHP-1 variants (Figure ) as well as the root mean square fluctuations of these atoms across the full enzyme scaffold (Figure ). Based on this data, there are no statistically significant differences between wild-type SHP-1 and the variants as determined by pairwise t-tests conducted at each residue index followed by a Benjamin-Hochberg correction (apart from small local differences in flexibility in residue A513 E427 and P428 between the wildtype and E427A variant). This is in contrast to our prior studies of PTP1B and YopH, where a single point swap between T177G (PTP1B) and G352T (YopH) on the WPD-loop both significantly impacted loop dynamics and the pH-dependency of catalysis. For comparison, the PTP1B F182Q point mutation similarly has no significant impact on loop dynamics, but in this case the pH-dependency of catalysis is unaffected. We have successfully used the empirical valence bond (EVB) approach to model chemistry in both wild-type PTP1B and YopH and variants. In the present case, while the H422Q, E427A and S418A mutations all impact the rates of catalysis by the SHP-1 catalytic domain, from a thermodynamic perspective, the difference in activation free energy between the variants is very small (within 0.5 kcal mol -1 change compared to wild-type), and thus EVB (or other) simulations of the chemical step of catalysis are unlikely to capture the origins of these small energy differences. Nevertheless, our EVB simulations ( resolution, the differences in calculated activation free energies between the variants is minimal, in agreement with the pH-independent limiting kcat values (Table ). Interestingly, examination of the number of water molecules within 3.5Å of the side chain of D421 (measured as the distance between the Cγ-atom of the D421 side-chain and the oxygen atoms of the surrounding water molecules) during our simulations of the phosphoenzyme intermediate with a closed WPD-loop (Figure and Table ) suggest subtle differences in solvation of the aspartic acid that could rationalize the differences in pKa of this residue. Further, the mean P-Onuc distance during our simulations of wild-type SHP1 and variants in the phosphoenzyme intermediate state is very subtly shifted to a tighter P-O distance, which would be expected to be favorable for catalysis, although this difference is within the standard deviation of the calculations (Figure and Table ). .