STRUCTURAL AND FUNCTIONAL CONSEQUENCES OF A GLU L212-]LYS MUTATION IN THE Q(B) BINDING-SITE OF THE PHOTOSYNTHETIC REACTION-CENTER OF RHODOPSEUDOMONAS-VIRIDIS

The properties of the quinone acceptor complex in the photosynthetic reaction center of the atrazine-resistant Rhodopseudomonas viridis mutant A2 (Glu L212 --> Lys) were studied by EPR spectroscopy and by photoelectric measurements. The EPR signal attributed to the semiquinone-iron (Q(B)(-)Fe(2+)) was significantly different from wild type and resembled that found in PS II. Essentially normal oscillations of Q(B)(-)Fe(2+) were observed upon flash illumination. The kinetics of the first and the second electron transfer from Q(A) to Q(B) were characterized by a photoelectric double-flash method. Compared to wild type, the rate of the first electron transfer in the large majority of reaction centers was decreased drastically from k(1) = (18 mu s)(-1) in the wild type to (70 ms)(-1) in the mutant, whereas the second electron transfer was only slightly slowed down with a rate of k(2) = (260 mu s)(-1) compared to (65 mu s)(-1) in wild type (pH 7). When the pH was raised above 10, in a major fraction of the reaction centers a fast kinetics of the first electron transfer, like that in wild type, reappeared. The experimental results are interpreted as an effect of the positive charge on the lysine causing a significant structural change of the QB binding pocket and a strongly diminished affinity for ubiquinone. The slow Q(A)(-) --> Q(B) electron transfer kinetics are thus attributed to ubiquinone binding, which is rate limiting. The possible role of the residue Glu L212, which is conserved in all purple bacteria, in electron and proton transfer to Q(B) is discussed.