Theoretical calculations on the acidity of the active site in aspartic proteinases.

Semiempirical minimal neglect of differential overlap-self-consistent field calculations, corrected and modified for multiple hydrogen-bonding interactions, were applied to models of the active site of aspartic proteinases (AP). The propensities of the two active-site aspartates to ionize were compared under the influence of various neighboring residues and of water molecules. Asp-32 and Asp-215 in three aspartic proteinases (endothiapepsin, Rhizopus pepsin, and penicillopepsin) are found to be basically asymmetric, Asp-32 being preferentially (by 2-3 kcal) ionized with respect to Asp-215. In penicillopepsin, this asymmetry is compensated by effects of surrounding residues. In our largest model for the active site, which includes such other residues, near equality is found for the ionizing tendency of Asp-32 and Asp-215. The pK difference is rationalized in terms of first and second ionizations of the full active-site model. Its ionization enthalpies correlate well with those of other small organic diacids. This "gas-phase" approach to AP active-site interactions represents the main possible contributions to the acidity of the active site.