GATING OF PHOTOINDUCED ELECTRON-TRANSFER FROM ZINC CYTOCHROME-C AND TIN CYTOCHROME-C TO PLASTOCYANIN - EFFECTS OF SOLUTION VISCOSITY ON REARRANGEMENT OF THE METALLOPROTEIN COMPLEX

Oxidative quenching by pc(II) of the triplet excited states Zn-3(cyt) and Sn-3(cyt) is studied by laser flash photolysis. The quenching is biphasic at low ionic strength. The faster component corresponds to the unimolecular electron-transfer reactions within the electrostatic complexes 3M(cyt)/pc(II), in which M is Zn(II) or Sn(IV); these reactions are the main subject of this study. The slower component corresponds to the bimolecular reactions between the unassociated 3M(cyt) and pc(II). The relative amplitudes of the two components depend on ionic strength and pc(II) concentration in a predictable way. The unimolecular reaction is studied in solutions whose viscosity is adjusted with glycerol, ethylene glycol, and D-glucose. In the electrostatic complex the positively charged (basic) patch around the exposed heme edge in cyt abuts the negatively charged (acidic) patch remote from the copper atom in pc. In the covalent complex this orientation is reinforced by noninvasive, tight cross-links. The intracomplex rate constant in the electrostatic complexes 3M(cyt)/pc(II), which are flexible, does not depend on ionic strength (in the range from 2.5 to 20 mM) but decreases smoothly from (2.5 +/- 0.4) X 10(5) s-1 when M is Zn(II) and from (1.9 +/- 0.4) X 10(5) s-1 when M is Sn(IV) as the viscosity is raised. The intracomplex rate constant in the covalent complex Zn-3(cyt)/pc(II), which is rigid, is (2.4 +/- 0.2) X 10(4) s-1 and is invariant with viscosity. The viscosity effects on the electrostatic complexes are fitted well to a simple mechanism involving two conformations of the electrostatic complex-the initial one that optimizes docking but not the intracomplex electron-transfer reaction and the rearranged one that optimizes this reaction. The fitted rate constant within the initial conformation of the electrostatic complex equals the observed rate constant within the covalent complex. At very high viscosity the rate constant within the electrostatic complex converges down to the rate constant within the covalent complex. These facts show that viscous solvents and cross-links act similarly in stabilizing and capturing the initial conformation of the electrostatic complex. The two limiting conformations of 3M(Cyt)/pc(II) interconvert by a process, probably conformational fluctuation, for which the fitted rate constants are 2.2 X 10(5) and 1.8 X 10(5) s-1 When M is Zn(II) and Sn(IV), respectively. Because the small difference between these values probably is insignificant, conformational fluctuation does not seem to depend on the protein overall charge as long as the surface charge is unaltered. The photoinduced reactions in Zn3(cyt)/pc(II) and Sn3(cyt)/pc(II), with the respective driving forces of 1.2 and 0.8 eV, are gated because the electron-transfer step in the rearranged complexes is faster than the rearrangement. The thermal reaction in cyt(II)/pc(II), with a driving force of 0.10 eV, is not gated because the electron-transfer step (1300+/-200 s-1) is slower than this rearrangement. Similar electron-transfer reactions between proteins can be gated or not gated, depending on the driving force.