INTRAMOLECULAR ELECTRON-TRANSFER REACTIONS OF CYTOCHROME B(5) COVALENTLY BONDED TO RUTHENIUM(II) POLYPYRIDINE COMPLEXES - REORGANIZATIONAL ENERGY AND PRESSURE EFFECTS

A series of ruthenium polypyridine complexes have been covalently atta ched to Cys73 of the T73C variant of synthetic rat liver cytochrome bg Photoexcitation of the ruthenium complex results in transient electro n transfer from the excited ruthenium complex, Ru(II), to the Fe(III) center of the protein followed by a rapid electron-transfer reaction from Fe(II) to Ru(III). The observed rate constants for the excited-st ate reaction ranged from 3.4 x 10(7) to 5 x 10(4) s(-1) and from 4.3 x 10(7) to 4.5 x 10(5) s(-1) for the back reaction. The free energy dep endence of the rate constants is best described by a reorganizational energy = 1 eV and an electron tunneling matrix element H-AB = 0.35 cm( -1) for electron transfer between the heme iron and the ruthenium comp lex. The reorganizational energy is the same as that obtained with the T65C variant labeled at Cys65 with the same series of ruthenium compl exes, H-AB, however, is 30% larger in the T73C derivatives. The pressu re dependence of the rate constants of two derivatives of cytochrome b (5) have been determined in order to investigate the sensitivity of th e electronic coupling to pressure dependent van der Waals contacts bet ween the redox centers. The first derivative was made by labeling the T65C variant with Ru((CH3)(2)bpy)(2)(CH(3)bpyCH(2-))(2+) and contains a well defined 12 bond covalent link between the redox centers. The T6 5C derivative shows no pressure dependence over the range of 0.1-200 M Pa. The second derivative was made by labeling the T73C variant with t he same complex. In this case, the electronic coupling between the red ox centers appears to involve a van der Waals contact between the hydr ogens on the gamma-carbon of Ser71 and a porphyrin methyl group. This derivative shows a very small negative volume of activation.