Structure of the charge separated state P865+QA- in the photosynthetic reaction centers of Rhodobacter sphaeroides by quantum beat oscillations and high-field electron paramagnetic resonance:: Evidence for light-induced QA- reorientation

The structure of the secondary radical pair, P-865(+),Q(A)(-), in fully deuterated and Zn-substituted reaction centers (RCs) of the purple bacterium Rhodobacter sphaeroides R-26 has been determined by high-time resolution and high-field electron paramagnetic resonance (EPR). A computer analysis of quantum beat oscillations, observed in a two-dimensional Q-band (34 GHz) EPR experiment, provides the orientation of the various magnetic tensors of P(865)(+)Q(A)(-) with respect to a magnetic reference frame. The orientation of the g-tensor of P-865(+), in an external reference system is adapted from a single-crystal W-band (95 GHz) EPR study [KIette, R.; T6rring, J. T.; Plato, M.; Mobius, K.; Bonigk, B.; Lubitz, W. J Phys. Chem. 1993, 97, 2015-2020]. Thus, we obtain the three-dimensional structure of the charge separated state P-865(+) Q(A)(-) on a nanosecond time scale after light-induced charge separation. Comparison with crystallographic data reveals that the position of the quinone is essentially the same as that in the X-ray structure. However, the head group of Q(A)(-) has undergone a 60 degrees rotation in the ring plane relative to its orientation in the crystal structure. Analysis suggests that the two different Q(A) Conformations are functionally relevant states which control the electron-transfer kinetics from Q(A)(-) to the secondary quinone acceptor Q(B). It appears that the rate-limiting step of this reaction is a reorientation of Q(A)(-) in its binding pocket upon light-induced reduction. The new kinetic model accounts for striking observations by Kleinfeld et al. who reported that electron transfer from Q(A)(-) to Q(B) proceeds in RCs cooled to cryogenic temperature under illumination but does not proceed in RCs cooled in the dark [Kleinfeld, D.; Okamura, M. Y.; Feher, G. Biochemistry 1984, 23, 5780-5786].