Control of metalloprotein reduction potential: The role of electrostatic and solvation effects probed on plastocyanin mutants

The changes in the thermodynamics of Cu(II) reduction for spinach plastocya nin induced by point mutations altering the electrostatic potential in prox imity of the copper center were determined through variable temperature dir ect electrochemistry experiments. In particular, the functionally important surface residues Leu12 and Gln88 were replaced with charged and polar resi dues, and Asn38 was substituted with Asp. The mutational variations of the reduction enthalpy and entropy were analyzed with a QSPR (quantitative stru cture-property relationships) approach, employing global and local theoreti cal descriptors defined and computed on the three-dimensional protein struc ture. The correlations found are informative on how electrostatic and solva tion effects control the E degrees' values in this species through the comb ined effects on the reduction enthalpy and entropy. The changes in reductio n enthalpy can be justified with electrostatic considerations. Most notably , enthalpy-entropy compensation phenomena play a significant role: the entr opic effects due to the insertion of charged residues determine E degrees' changes that are invariably opposite to those induced by the concomitant en thalpic effects. Therefore, the resulting E degrees' changes are small or e ven opposite to those expected on simple electrostatic grounds. The mutatio nal variation in the reduction entropy appears to be linked to the hydrogen bonding donor/acceptor character of the northern part of the protein, abov e the metal site, and to the electrostatic potential distribution around th e copper site. Both properties influence the reduction-induced reorganizati on of the water molecules on the protein surface in the same region.