Electron Transfer Reactions of Chemically Modified Plastocyanin with P700 and Cytochrome f. Importance of Local Charges1

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Abstract
Chemically modified spinach plastocyanin, in which negatively charged carboxyl residues are replaced with positively charged amino residues, has been prepared. Four distinct species of chemically modified plastocyanin, having 1 to 4 mol of modified carboxyl residue per mol of plastocyanin, could be separated by ion exchange chromatography on DEAE-Sephacel. The rate of electron transfer from reduced cytochrome f to oxidized singly substituted plastocyanin was only 30% of that of the native unmodified plastocyanin, and the reaction rate decreased further with increasing number of modified carboxyl residues. These results indicate the importance of electrostatic interactions between the negative charges on plastocyanin and the positive charges on cytochrome f in this reaction. Since the overall net charge of cytochrome f is negative at neutral pH, the positive charges on cytochrome f involved in the reaction should be localized ones. On the other hand, the rates of electron transfer from reduced singly and doubly substituted plastocyanin to photooxidized P700 in the P700-chlorophyll a protein complex were similar to that of native plastocyanin, which suggests that these carboxyl residues have only a minor role in the electron transfer to P700. Although divalent cation is essential for the electron transfer from native plastocyanin to P700 at neutral pH, the triply substituted plastocyanin could donate electrons to P700 even without MgCl 2 , and the rate of this reaction reached the maximum at a low concentration of MgCl 2 (2 . These results suggest the presence of different reaction site domains, at least partially, on the plastocyanin for the electron transfer to P700 and for the electron transfer from cytochrome f . The results are discussed in relation to the electron input to and output from plastocyanin in vivo .