Primary electron transfer in reaction centers of YM210L and YM210L/HL168L mutants of <Emphasis Type="Italic">Rhodobacter sphaeroides</Emphasis>

The role of tyrosine M210 in charge separation and stabilization of separated charges was studied by analyzing of the femtosecond oscillations in the kinetics of decay of stimulated emission from P* and of a population of the primary charge separated state P⁺B_A⁻ in YM210L and YM210L/HL168L mutant reaction centers (RCs) of Rhodobacter sphaeroides in comparison with those in native Rba. sphaeroides RCs. In the mutant RCs, TyrM210 was replaced by Leu. The HL168L mutation placed the redox potential of the P⁺/P pair 123 mV below that of native RCs, thus creating a theoretical possibility of P⁺B_A⁻ stabilization. Kinetics of P* decay at 940 nm of both mutants show a significant slowing of the primary charge separation reaction in comparison with native RCs. Distinct damped oscillations in these kinetics with main frequency bands in the range of 90–150 cm⁻¹ reflect mostly nuclear motions inside the dimer P. Formation of a very small absorption band of B_A⁻ at 1020 nm is registered in RCs of both mutants. The formation of the B_A⁻ band is accompanied by damped oscillations with main frequencies from ∼10 to ∼150 cm⁻¹. Only a partial stabilization of the P⁺B_A⁻ state is seen in the YM210L/HL168L mutant in the form of a small non-oscillating background of the 1020-nm kinetics. A similar charge stabilization is absent in the YM210L mutant. A model of oscillatory reorientation of the OH-group of TyrM210 in the electric fields of P⁺ and B_A⁻ is proposed to explain rapid stabilization of the P⁺B_A⁻ state in native RCs. Small oscillatory components at ∼330–380 cm⁻¹ in the 1020-nm kinetics of native RCs are assumed to reflect this reorientation. We conclude that the absence of TyrM210 probably cannot be compensated by lowering of the P⁺B_A⁻ free energy that is expected for the double YM210L/HL168L mutant. An oscillatory motion of the HOH55 water molecule under the influence of P⁺ and B_A⁻ is assumed to be another potential contributor to the mechanism of P⁺B_A⁻ stabilization.