A theoretical investigation of tight-binding thermolysin inhibitors.

A tight-binding thermolysin inhibitor, Cbz-Phe-psi[PO2NH]-Leu-Ala (ZFpLA, Ki = 0.068 nM), and its analogs, ZRp(O)LA (R = Ala, Leu or Phe) have been studied using the finite difference solution to the linearized Poisson-Boltzmann equation (FDPB) and solvation entropy correction (SEC). The binding energy difference between conformationally different thermolysin inhibitors ZFpLA and ZGpLL is estimated using three approaches. Two of approaches use the X-ray structures of ZFpLA-thermolysin and ZGpLL-thermolysin structures. The third one uses both X-ray structures to calculate binding energy differences from ZFpLA and ZGpLL to a hypothetical intermediate MepLA. All the results are qualitatively correct with one closely reproducing the experimental value. The enhancement of the ZFpLA binding is attributed largely to the solvation entropy or "hydrophobic force". The binding mode of the ZGpLR N-terminal moiety appears to be electrostatically unfavorable. Reducing the polarity of that moiety is predicted to enhance binding affinity. The binding trends due to the hydrophobic variation of ZRp(O)LA are calculated within 1 kcal/mol of the experimental values. Increasing lipophilicity of a ligand favors the binding due to the difference of surface area change between the free state and the bound state. The analysis of energetic components shows that these trends are not specific for the binding of phosphorus-containing inhibitors but are generally true for protein-ligand interactions. The electrostatic calculation does not support the involvement of the second protonation of ZFpLA in binding. Therefore, reexamining the second protonation of ZFpLA or seeking further experimental support seems appropriate. The structural sensitivity of the FDPB calculation was assessed by using ligand and receptor structures from different X-ray studies of thermolysin. The small deviations (< 0.3 A) in the receptor structures do not cause significant changes in electrostatic binding energy if there is no structural change in modified regions.