MOLECULAR-BASIS OF RESISTANCE TO HIV-1 PROTEASE INHIBITION - A PLAUSIBLE HYPOTHESIS

The binding thermodynamics of the HIV-1 protease inhibitor acetyl peps tatin and the substrate Val-Ser-Gln-Asn-Tyr-Pro-Ile-Val-Gln, correspon ding to one of the cleavage sites in the gag, gag-pol polyproteins, ha ve been measured by direct microcalorimetric analysis. The results ind icate that the binding of the peptide substrate or peptide inhibitor i s entropically driven; i.e., it is characterized by an unfavorable ent halpy and a favorable entropy change, in agreement with a structure-ba sed thermodynamic analysis based upon an empirical parameterization of the energetics. Dissection of the binding enthalpy indicates that the intrinsic interactions are favorable and that the unfavorable enthalp y originates from the energy cost of rearranging the flap region in th e protease molecule. In addition, the binding is coupled to a negative heat capacity change. The dominant binding force is the increase in s olvent entropy that accompanies the burial of a significant hydrophobi c surface. Comparison of the binding energetics obtained for the subst rate with that obtained for synthetic nonpeptide inhibitors indicates that the major difference is in the magnitude of the conformational en tropy change. In solution, the peptide substrate has a higher flexibil ity than the synthetic inhibitors and therefore suffers a higher confo rmational entropy loss upon binding. This higher entropy loss accounts for the lower binding affinity of the substrate. On the other hand, d ue to its higher flexibility, the peptide substrate is more amenable t o adapt to backbone rearrangements or subtle conformational changes in duced by mutations in the protease. The synthetic inhibitors are less flexible, and their capacity to adapt is more restricted. The expected result is a more pronounced effect of mutations on the binding affini ty of the synthetic inhibitors. On the basis of the thermodynamic diff erences in the mode of binding of substrate and synthetic inhibitors, it appears that a key factor to understanding resistance is given by t he relative balance of the different forces that contribute to the bin ding free energy and, in particular, the balance between conformationa l and solvation entropy.