Electron transfer between cytochrome c(2) and the tetraheme cytochrome c in Rhodopseudomonas viridis

Kinetics of electron transfer from soluble cytochrome c(2) to the tetraheme cytochrome c have been measured in isolated reaction centers and in membra ne fragments of the photosynthetic purple bacterium Rhodopseudomonas viridi s by time-resolved flash absorption spectroscopy. Absorbance changes kineti cs in the region of cytochrome alpha-bands (540-560 nm) were measured at 21 degrees C under redox conditions where the two high-potential hemes (c-559 and c-556) of the tetraheme cytochrome were chemically reduced. After flas h excitation, the heme c-559 donates an electron to the special pair of bac teriochlorophylls and is then re-reduced by heme c-556. The data show that oxidized heme c-556 is subsequently re-reduced by electron transfer from re duced cytochrome c(2) present in the solution. The rate of this reaction ha s a non-linear dependence on the concentration of cytochrome c(2), suggesti ng a (minimal) two-step mechanism involving the formation of a complex betw een cytochrome c(2) and the reaction center, followed by intracomplex elect ron transfer. To explain the monophasic character of the reaction kinetics, we propose a collisional mechanism where the lifetime of the temporary com plex is short compared to electron transfer. The limit of the halftime of t he bimolecular process when extrapolated to high concentrations of cytochro me c(2) is 60 +/- 20 mu s. There is a large ionic strength effect on the ki netics of electron transfer from cytochrome c(2) to heme c-556, The pseudof irst-order rate constant decreases from 1.1 x 10(7) M-1 s(-1) to 1.3 x 10(6 ) M-1 s(-1) when the ionic strength is increased from 1 to 1000 mM. The max imum rate (1.1 x 10(7) M-1 s(-1)) was obtained at about 1 mM ionic strength . This dependence of the rate on ionic strength suggests that attractive el ectrostatic interactions contribute to the binding of cytochrome c(2) with the tetraheme cytochrome. On the basis of our data and of previous molecula r modelling, it is proposed that cytochrome c(2) docks close to the low-pot ential heme c-554 and reduces heme c-556 via c-554.