PHOTOINDUCED ELECTRON-TRANSFER REACTION IN A TERNARY-SYSTEM INVOLVING ZINC CYTOCHROME-C AND PLASTOCYANIN - INTERPLAY OF MONOPOLAR AND DIPOLAR ELECTROSTATIC INTERACTIONS BETWEEN METALLOPROTEINS

A carbodiimide promotes noninvasive cross-linking between amino groups surrounding the exposed heme edge in zinc cytochrome c and carboxylic groups in the acidic patch in plastocyanin. Eight derivatives of the covalent complex Zncyt/pc(I), which have similar structures but different overall charges because of different numbers and locations of N-acylurea groups, are separated by cation-exchange chromatography. Kinetics of electron transfer from the diprotein complex in the triplet excited state, 3Zncyt/pc(I), to free cupriplastocyanin at pH 7.0 and various ionic strengths is studied by laser flash spectroscopy. This reaction is purely bimolecular for all eight N-acylurea derivatives of the diprotein complex. The overall charges of the derivatives 1 and 2 at pH 7.0 are -2 and 0, respectively; both of them, however, have very large dipole moments of 410-480 D. The rate constants for their reactions with cupriplastocyanin, whose charge at pH 7.0 is -8 and whose dipole moment is 362 D, are determined over the range of ionic strengths from 2.5 mM to 3.00 M. The observed dependence of the rate constants on ionic strength cannot be explained in terms of net charges (monopole-monopole interactions) alone, but it can be fitted quantitatively with a theory that recognizes also monopole-dipole and dipole-dipole interactions [van Leeuwen, J. W. (1983) Biochim. Biophys. Acta 743, 408]. At ionic strengths up to ca. 10 mM monopole-monopole interactions predominate and Bronsted-Debye-Huckel theory applies. At higher ionic strengths the monopole-dipole and dipole-dipole interactions predominate, Bronsted-Debye-Huckel theory yields incorrect net charges, and all three types of electrostatic interactions must be recognized. Our previous study [Zhou, J. S., & Kostic, N. M. (1991) J. Am. Chem. Soc. 113, 7040] showed that cupriplastocyanin uses its acidic patch near Tyr 83 for interaction with free 3Zncyt. Many fittings of the kinetic results to van Leeuwen theory, with reasonable parameters, are all consistent with the notion that cupriplastocyanin uses its hydrophobic patch near His 87 for interactions with 3ZnCyt in the complex 3Zncyt/pc(I). The covalently attached cuproplastocyanin largely neutralizes the positively charged patch and largely shields the exposed heme edge in zinc cytochrome c. Therefore, the 3Zncyt moiety of the diprotein complex is not significantly attracted to the acidic patch in free cupriplastocyanin, which is connected to the copper site by relatively inefficient electron-transfer paths. This moiety reacts at the hydrophobic patch in free cupriplastocyanin, the origin of efficient electron-transfer paths to the copper site.