Comparative voltammetric studies on Anabaena variabilis plastocyanin (positively charged at neutral pH) and spinach and poplar plastocyanins (negatively charged at neutral pH) have been undertaken at an edge-plane graphite electrode as a function of ionic strength, pH, and Mg2+ concentration at 3 °C. The aim was to provide a more detailed understanding of the influence of the electrode-protein (solution) interfacial characteristics, as well as the variation of the formal potential with both the nature of the plastocyanin species and the pH. As might be expected, some of the interfacial properties associated with the positive charge on A. variabilis plastocyanin are the opposite of those observed with the negatively charged plastocyanins. For example, the linear diffusion component of the mass transport process for A. variabilis plastocyanin under the conditions of cyclic voltammetry is decreased and the radial diffusion component is increased by the addition of Mg2+, whereas the reverse occurs with poplar and spinach plastocyanins. The voltammetrically determined reversible potentials for A. variabilis plastocyanin are considerably less positive than those for spinach and poplar plastocyanins, in agreement with values calculated from chemically based redox studies. Ionic strength effects, as determined by addition of NaCIO4 over the concentration range 0.005-0.20 M, are negligible for all three proteins. The addition of Mg2+ causes a significant shift in the reversible potential toward more positive values for spinach and poplar plastocyanin but only a small positive shift for A. variabilis plastocyanin. The difference is attributed to a specific binding effect. The addition of Mg2+ also dramatically alters the pH dependence of the reversible potential, indicating that the equilibrium between the protonated and unprotonated forms of reduced plastocyanin is modified by binding of Mg2+ to the protein. It is concluded that the effects of biologically relevant redox-inactive cations such as Mg2+ or Ca2+ have to be considered carefully in studies of the redox chemistry of metalloproteins.
|Number of pages||10|
|Publication status||Published - 1 Dec 1996|