Absorption changes induced by the binding of triazines to the Q(B) pocket in reaction centers of Rhodobacter capsulatus

Inhibitors which block electron transfer from the primary (Q(A)) to the sec ondary (Q(B)) quinone of the bacterial reaction center are competing with t he pool ubiquinones for binding at the QB pocket. Due to the much greater s tability of the semiquinone state Q(B)(-) compared with fully oxidized or r educed quinone, a displacement of the inhibitors takes place after one flas h from state Q(A)(-)I to State Q(A)Q(B)(-). This process can be monitored f rom near-IR absorption changes which reflect local absorption shifts specif ic to Q(A)(-) and Q(B)(-). An anomalous behavior was observed when using tr iazines in chromatophores of R. capsulatus: the IR absorption change reflec ting the formation of Q(B)(-) after one flash was absent. A normal transien t decay of this signal was, however, triggered by a second flash, followed by a rapid return to the baseline. We show that this phenomenon is due to a n absorption change induced by inhibitor binding (thus present in the dark baseline), with a spectrum close to that of Q(B)(-), so that the Q(B)(-) ch anges are canceled out during the inhibitor displacement process. On the se cond flash, one monitors the destruction of the semiquinone, leading transi ently to the Q(A)Q(B) state, followed by inhibitor rebinding. This allows a direct measurement of the binding kinetics. This behavior was observed bot h in chromatophores and in isolated reaction centers from R. capsulatus, bu t not in R. sphaeroides.