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

The properties of the quinone acceptor complex in the photosynthetic r eaction center of the atrazine-resistant Rhodopseudomonas viridis muta nt A2 (Glu L212 --> Lys) were studied by EPR spectroscopy and by photo electric measurements. The EPR signal attributed to the semiquinone-ir on (Q(B)(-)Fe(2+)) was significantly different from wild type and rese mbled that found in PS II. Essentially normal oscillations of Q(B)(-)F e(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 ra te of the first electron transfer in the large majority of reaction ce nters 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 transfe r was only slightly slowed down with a rate of k(2) = (260 mu s)(-1) c ompared to (65 mu s)(-1) in wild type (pH 7). When the pH was raised a bove 10, in a major fraction of the reaction centers a fast kinetics o f the first electron transfer, like that in wild type, reappeared. The experimental results are interpreted as an effect of the positive cha rge on the lysine causing a significant structural change of the QB bi nding pocket and a strongly diminished affinity for ubiquinone. The sl ow Q(A)(-) --> Q(B) electron transfer kinetics are thus attributed to ubiquinone binding, which is rate limiting. The possible role of the r esidue Glu L212, which is conserved in all purple bacteria, in electro n and proton transfer to Q(B) is discussed.