Plastocyanin (PC) and its physiological reaction partner cytochrome (cyt) f form a complex which is electrostatically stabilized by interactions between complementary localized charges. We have measured the kinetics of intracomplex electron transfer between several reduced cytochromes and PC using laser flash photolysis. With spinach cyt f and spinach PC, we obtain first-order rate constants, kforward = 2780 s-1 and kreverse = 1050 s-1 for the reversible reaction and a complex dissociation constant of about 23 μM at an ionic strength (I) of 5 mM. The observed rate constant increases by a factor of 2 between I = 5 and 40 mM and then decreases monotonically at higher ionic strengths. This indicates that the complex is not completely dissociated until I=150 mM and that the proteins within the electrostatically most stable complex are not optimally oriented for electron transfer. Similar results were obtained with turnip cyt f and spinach PC, although in this case intracomplex electron transfer is about 4 times as fast. Horse cyt c also forms an electrostatically stabilized complex with PC, and yields a limiting rate constant for intracomplex electron transfer (1750 s-1) and a dissociation constant (10 μM) comparable to those for spinach cyt f. The ionic strength dependence shows that the complex is more readily dissociated (complete at I = 25 mM) than is that of cyt f and that rearrangement is not required for optimal electron transfer. Addition of polylysine results in 10-fold inhibition of the rate of electron transfer. Pseudomonas cyt c-551 is an acidic cytochrome which does not form a complex with PC. The second-order rate constant for reaction with PC at I = 5 mM is about 2 orders of magnitude smaller than for cyt f or cyt c, and the effect of increasing ionic strength on the cyt c-551 reaction is to monotonically increase the rate constant. Addition of polylysine also results in an increase in the rate constant, which is comparable in magnitude to that caused by ionic strength. Chlorobium cyt c-555 is a basic protein like horse cyt c, but the kinetic results indicate that it does not form a complex with PC, although the second-order rate constant is similar to those obtained with the other basic cytochromes. The effect of ionic strength is to monotonically decrease the rate constant in a manner consistent with simple electrostatic theory. For all of the cytochromes, the rate constants extrapolated to infinite ionic strength are comparable in magnitude. However, protein-protein electrontransfer rate constants should correlate with the thermodynamic driving force if the distances and orientations are held constant. The fact that they do not suggests that these factors vary for the different cytochromes in their reactions with PC.
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