Light-induced proton uptake by photosynthetic reaction centers from Rhodobacter sphaeroides R-26.1. II. Protonation of the state DQAQB2-

Abstract

Proton uptake associated with the two-electron reduction of QB 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 < or = 7.5, consistent with the formation of the dihydroquinone QBH2. At higher pH, the proton uptake decreased with an apparent pKa 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 pKa is due to the protonation of either the carbonyl oxygen on QB or of an amino acid residue near QB (e.g., His-L190). Experimental evidence in favor of the protonation of the oxygen is discussed. The kinetics of the electron transfer from QA-QB- to QAQB2- 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 QB occurs sequentially, one concomitant with and the other after electron transfer.