Identifying key electrostatic interactions in Rhizomucor miehei lipase: the influence of solvent dielectric

The conformational change associated with the interfacial activation of Rhi zomucor miehei lipase involves the displacement of an m-helical lid (residu es 82-96) away from the active site on moving from water (high dielectric) to lipid (low dielectric). The presence of two media of very different diel ectric properties suggests that electrostatic inter-actions play an importa nt role in this process. We have used linearized Poisson-Boltzmann calculat ions to examine the key electrostatic interactions which contribute to lid stability in the closed and open states. It is the two charged residues of the lid, Arg86 and Asp91, that form the strongest electrostatic interaction s with the rest of the protein. We identify key residues whose interactions with the lid are significantly perturbed by the change in the dielectric o f the medium: Asp61, Arg80, Lys109, Glu117 and the active-site residues Asp 203 and Asp256, all of which lie within approximately 20 Angstrom of the li d. We suggest that these residues are good candidates for site-specific mut ation studies, which could help elucidate their role in the lipase activati on mechanism.