Retardation of proton transfer caused by binding of the transition metal ion to the bacterial reaction center is due to pKa shifts of key protonatable residues.
Transition metal ions bind to the reaction center (RC) protein of the photosynthetic bacterium Rhodobacter sphaeroides and slow the light-induced electron and proton transfer to the secondary quinone, Q(B). We studied the properties of the metal ion-RC complex by measuring the pH dependence of the dissociation constant and the stoichiometry of proton release upon ligand formation. We investigated the mechanism of inhibition by measuring the stoichiometry and kinetics of flash-induced proton binding, the transfer of (first and second) electrons to Q(B), and the rate of steady-state turnover of the RC in the absence and presence of Cd(2+) and Ni(2+) on a wide pH range. The following results were obtained. (1) The complexation of transition metal ions Cd(2+) and Ni(2+) with the bacterial RC showed strong pH dependence. This observation was explained by different (pH-dependent) states of the metal-ligand cluster: the complex formation was strong when the ligand (Asp and His residues) was deprotonated and was much weaker if the ligand was partly (or fully) protonated. A direct consequence of the model was the pH-dependent proton release upon complexation. (2) The retardation of transfer of electrons and protons to Q(B) was also strongly pH-dependent. The effect was large in the neutral pH range and decreased toward the acidic and alkaline pH values. (3) Steady-state turnover measurements indicated that the rate of the second proton transfer was much less inhibited than that of the first one, which became the rate-limiting step in continuous turnover of the RC. (4) Sodium azide partly recovered the proton transfer rate. The effect is not due to removal of the bound metal ion by azide but probably by formation of a proton-transporting azide network similarly as water molecules may build up proton pathways. (5) We argue that the inhibition comes mainly from pK(a) shifts of key protonatable residues that control the proton transfer along the H-bond network to Q(B). The electrostatic interaction between the metal ion and these residues may result in acidic pK(a) shifts between 1.5 and 2.0 that account for the observed retardation of the electron and proton transfer.
Gerencsér
L
L
Department of Biophysics, University of Szeged, P.O. Box 655, 6701 Szeged, Hungary.
Maróti
P
P
D001386:Azides / Q000378:metabolism; D016227:Benzoquinones / Q000378:metabolism; D001665:Binding Sites; D002413:Cations, Divalent / Q000378:metabolism; D004579:Electron Transport; D007700:Kinetics; D008024:Ligands; D008670:Metals / Q000378:metabolism*; D008956:Models, Chemical; D010782:Photolysis; D045322:Photosynthetic Reaction Center Complex Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011522:Protons; D055672:Static Electricity
D001386:Azides; D016227:Benzoquinones; D002413:Cations, Divalent; D008024:Ligands; D008670:Metals; D045322:Photosynthetic Reaction Center Complex Proteins; D011522:Protons; C004532:quinone