PRIMARY ELECTRON-TRANSFER IN MEMBRANE-BOUND REACTION CENTERS WITH MUTATIONS AT THE M210 POSITION

The kinetics of primary electron transfer in membrane-bound Rhodobacte r sphaeroides reaction centers (RCs) were measured on both wild type ( WT) and site-directed mutant RC's bearing mutations at the tyrosine M2 10 position. The tyrosine was replaced by histidine (H), phenylalanine (F), leucine (L), or tryptophan (W). A mutant with histidine at both the M210 and symmetry-related L181 positions (YM210H/FL181H) was also examined. Rates of primary charge separation were determined by both s ingle and multiple wavelength pump-probe techniques. The time constant s for the decay of stimulated emission in the membrane-bound mutant RC 's were approximately 27 ps (F), 36 ps (L), 72 ps (W), 5.8 ps (H), and 4.2 ps (HH), compared with 4.6 ps in WT membrane-bound RC's. For all RC's, the decay of stimulated emission was found to be multiexponentia l, demonstrating that this phenomenon is not a consequence of the remo val of the RC from the membrane. The source of the multiexponential de cay of the primary donor excited state was examined, leading to the co nclusion that a distribution in the driving force (AG) for electron tr ansfer cannot be the sole parameter that determines the multiexponenti al character. Further measurements on membrane-bound mutant RC's showe d that chemical prereduction of the acceptor quinones resulted in a si gnificant slowing of the rate of primary charge separation. This was m ost marked in those mutants in which the rate of charge separation had already been slowed down as a result of mutagenesis at the M210 posit ion. The phenomenon is discussed in terms of the Coulombic interaction between Q(A)(-) and the other pigments involved in electron transfer and the influence of this interaction on the driving force for charge separation.