Free energy perturbation and molecular dynamics calculations of copper binding to azurin.

Free energy perturbation/mol. dynamics simulations have been carried out on copper/azurin systems calcg. the binding affinities of copper (II) ion to azurin either in the native or in the unfolded state. In order to test the validity of the strategy adopted for the calcns. and to establish what force field is suitable for these kinds of calcns., three different force fields, AMBER, CVFF, and CFF, have been alternatively used for the calcns. and the results have been compared with exptl. data obtained by spectroscopic titrns. of copper (II)/azurin solns. and denaturation expts. Our findings have pointed out that only CFF gives satisfactory results, thus providing a reliable tool for copper binding simulations in copper protein.