LIGHT-INDUCED PROTON UPTAKE BY PHOTOSYNTHETIC REACTION CENTERS FROM RHODOBACTER-SPHAEROIDES R-26.1 .2. PROTONATION OF THE STATE DQ(A)Q(B)(2-)

Proton uptake associated with the two-electron reduction of Q(B) was investigated in reaction centers (RCs) from Rhodobacter sphaeroides R-26.1 using pH-sensitive dyes. An uptake of two protons was observed at pH less-than-or-equal-to 7.5, consistent with the formation of the dihydroquinone Q(B)H-2. At higher pH, the proton uptake decreased with an apparent pK(a) of approx. 8.5, i.e., to 1.5 H+/2 e- at pH 8.5. A molecular model is presented in which the apparent pK(a) is due to the protonation of either the carbonyl oxygen on Q(B) or of an amino acid residue near Q(B) (e.g., His-L190). Experimental evidence in favor of the protonation of the oxygen is discussed. The kinetics of the electron transfer from Q(A)-Q(B)- to Q(A)Q(B)2- and the associated proton uptake were compared at several pH values and temperatures. At pH 8.5 (21.5-degrees-C) the rate constants for the proton uptake and electron transfer are the same within the precision of the measurement. At lower pH, the proton uptake rate constant is smaller than that for electron transfer. The difference between the rate constants is temperature dependent, i.e., it varies from 12 +/- 4% at 21.5-degrees-C (pH 7.5) to 28 +/- 4% at 4.0-degrees-C (pH 7.5). We show that the kinetics can be explained by a previously proposed model (Paddock, M.L., McPherson, P.H., Feher, G. and Okamura, M.Y. (1990) Proc. Natl. Acad. Sci. USA 87, 6803-6807) in which the uptake of two protons by doubly reduced Q(B) occurs sequentially, one concomitant with and the other after electron transfer.