Cluster analysis of hydration waters around the active sites of bacterial alanine racemase using a 2‐ns MD simulation

Structural data produced by a 2‐ns molecular dynamics (MD) simulation on Geobacillus alanine racemase (AlaR; PDB: 1SFT) was used to study hydration around the two AlaR active sites. AlaR is a crucial enzyme for bacterial cell wall biosynthesis. It has been shown previously that the potency of an inhibitor can be increased by incorporating a functional group or atom that displaces hydration sites close to the substrate binding pocket of its target enzyme. The complete linkage algorithm was used for cluster analysis of the active site water positions from 126 solvent configurations sampled at regular intervals from the 2‐ns MD simulation. Crystal waters in the 1SFT X‐ray structure occupy most of the tightly bound water sites that were discovered. We show here that tightly bound water sites can be identified by cluster analysis of MD‐generated coordinates starting with data supplied by a single X‐ray structure, and we predict a highly conserved hydration site close to the carboxyl oxygen of L‐Ala substrate. This approach holds promise for accelerating the drug design process. We also discuss an analysis of the well‐known notion of residence time and introduce a new measure called retention time. © 2007 Wiley Periodicals, Inc. Biopolymers 89: 210–219, 2008.