Computational analysis of binding of P1 variants to trypsin

The binding of P1 variants of bovine pancreatic trypsin inhibitor (BPTI) to trypsin has been investigated by means of molecular dynamics simulations. The specific interaction formed between the amino acid at the primary bindi ng (P1) position of the binding loop of BPTI and the specificity pocket of trypsin was estimated by use of the linear interaction energy (LTE) method. Calculations for 13 of the naturally occurring amino acids at the P1 posit ion were carried out, and the results obtained were found to correlate well with the experimental binding free energies. The LIE calculations rank the majority of the 13 variants correctly according to the experimental associ ation energies and the mean error between calculated and experimental bindi ng free energies is only 0.38 kcal/mole, excluding the Glu and Asp variants , which Me associated with some uncertainties regarding protonation and the possible presence of counter-ions. The three-dimensional structures of the complex with three of the P1 variants (Asn, Tyr, and Ser) included in this study have not at present been solved by any experimental techniques and, therefore, were modeled on the basis of experimental data from pi variants of similar size. Average structures were calculated from the MD simulations , from which specific interactions explaining the broad variation in associ ation energies were identified. The present study also shows that explicit treatment of the complex water-mediated hydrogen bonding network at the pro tein-protein interface is of crucial importance for obtaining reliable bind ing free energies, The successful reproduction of relative binding energies shows that this type of methodology can be very useful as an aid in ration al design and redesign of biologically active macromolecules.