KINETIC STUDIES ON 1-1 ELECTRON-TRANSFER REACTIONS INVOLVING BLUE COPPER PROTEINS .2. PROTONATION EFFECTS AND DIFFERENT BINDING-SITES IN THE OXIDATION OF PARSLEY PLASTOCYANIN WITH CO(4,7-DPSPHEN)33-, FE(CN)63-, AND CO(PHEN)33+

Strong protcin-complex association (K) prior to electron transfer (ket) is observed in the oxidation of parsley plastocyanin, PCu1, with the 4, 7-di(phenvl-4ʹ-sulfonate)-1, 10-phenanthroline complex. Co(4, 7-DPSphen)33-. At 25 °C, I = 0.10 M (NaCl). K = 4600 M-1, πH° = -4.2 kcal mol-1, πS° = 2.7 cal K-1mol-1, ket= 0.041 s-1, πH‡cl= 13.2 kcal mol-1. πS‡et= -20.6 cal -1mol-1. Rate constants are independent of pH in the range 5.2-7.5 investigated. This contrasts with the strong pH dependence observed with Co(phen)33+ and Fe(CN)63~ as oxidants (p/Ca— 6). Addition of redox-inactive Cr-(phen)33+(which forms a 1: 1 adduct with the protein) blocks reaction with Co(phen)33+, consistent with a single binding site being utilized by this oxidant. Rate constants for the Fe(CN)63-oxidation of PCu1at pH 7.0 are unaffected by the presence of Co(4.7-DPSphen)33-which is !:1 associated (up to 70%) with the protein. It is concluded that Co(4, 7-DPSphen)33-and Fe(CN)63-use different binding sites on the protein. Moreover, for the reaction of Co(phen)33+with PCu1it has been shown that on decreasing the pH from 7.5 to 5.5 K decreases significantly, whereas fcclincreases slightly. For this reaction the decrease in overall rate constants to zero at pH ca. 5 is therefore due to K→0. Such an effect suggests that H+modifies the binding site on the PCu1. A similar change is not observed with PCu1. These results indicate thatdifferent binding sites have different reduction potentials. © 1979, American Chemical Society. All rights reserved.