Electrostatic roles in electron transfer from [NiFe] hydrogenase to cytochrome c 3 from Desulfovibrio vulgaris Miyazaki F.

Electrostatic interactions between proteins are key factors that govern the association and reaction rate. We spectroscopically determine the second-order reaction rate constant (k) of electron transfer from [NiFe] hydrogenase (H ₂ ase) to cytochrome (cyt) c ₃ at various ionic strengths (I). The k value decreases with I. To analyze the results, we develop a semi-analytical formula for I dependence of k based on the assumptions that molecules are spherical and the reaction proceeds via a transition state. Fitting of the formula to the experimental data reveals that the interaction occurs in limited regions with opposite charges and with radii much smaller than those estimated from crystal structures. This suggests that local charges in H ₂ ase and cyt c ₃ play important roles in the reaction. Although the crystallographic data indicate a positive electrostatic potential over almost the entire surface of the proteins, there exists a small region with negative potential on H ₂ ase at which the electron transfer from H ₂ ase to cyt c ₃ may occur. This local negative potential region is identical to the hypothetical interaction sphere predicted by the analysis. Furthermore, I dependence of k is predicted by the Adaptive Poisson-Boltzmann Solver considering all charges of the amino acids in the proteins and the configuration of H ₂ ase/cyt c ₃ complex. The calculation reproduces the experimental results except at extremely low I. These results indicate that the stabilization derived from the local electrostatic interaction in the H ₂ ase/cyt c ₃ complex overcomes the destabilization derived from the electrostatic repulsion of the overall positive charge of both proteins.
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