Proton transfer from the bulk to the bound ubiquinone Q(B) of the reaction center in chromatophores of Rhodobacter sphaeroides: retarded conveyance by neutral water.
The mechanism of proton transfer from the bulk into the membrane protein interior was studied. The light-induced reduction of a bound ubiquinone molecule Q(B) by the photosynthetic reaction center is accompanied by proton trapping. We used kinetic spectroscopy to measure (i) the electron transfer to Q(B) (at 450 nm), (ii) the electrogenic proton delivery from the surface to the Q(B) site (by electrochromic carotenoid response at 524 nm), and (iii) the disappearance of protons from the bulk solution (by pH indicators). The electron transfer to Q(B)(-) and the proton-related electrogenesis proceeded with the same time constant of approximately 100 microseconds (at pH 6.2), whereas the alkalinization in the bulk was distinctly delayed (tau approximately 400 microseconds). We investigated the latter reaction as a function of the pH indicator concentration, the added pH buffers, and the temperature. The results led us to the following conclusions: (i) proton transfer from the surface-located acidic groups into the Q(B) site followed the reduction of Q(B) without measurable delay; (ii) the reprotonation of these surface groups by pH indicators and hydronium ions was impeded, supposedly, because of their slow diffusion in the surface water layer; and (iii) as a result, the protons were slowly donated by neutral water to refill the proton vacancies at the surface. It is conceivable that the same mechanism accounts for the delayed relaxation of the surface pH changes into the bulk observed previously with bacteriorhodopsin membranes and thylakoids. Concerning the coupling between proton pumps in bioenergetic membranes, our results imply a tendency for the transient confinement of protons at the membrane surface.
Gopta
O A
OA
Division of Biophysics, University of Osnabrück, D-49069 Osnabrück, Germany.
Cherepanov
D A
DA
Junge
W
W
Mulkidjanian
A Y
AY
D017136:Ion Transport; D007700:Kinetics; D011522:Protons; D012242:Rhodobacter sphaeroides / Q000737:chemistry*; D013816:Thermodynamics; D014451:Ubiquinone / Q000737:chemistry*; D014867:Water / Q000737:chemistry
D011522:Protons; D014867:Water; D014451:Ubiquinone