Complete Tracking of Proton Flow in Thylakoids the Unit Conductance of CF0 is Greater than 10 fs
We investigated the proton conductance of the channel portion of chlo-roplast ATP synthase (CF0) in thylakoids which were depleted from the soluble portion of the ATP synthase (CF1) by EDTA treatment. Proton pumps were stimulated by short flashes of light. Proton flux through CF0 was measured spectrophotometrically in three different ways: as proton efflux from the lumen (via neutral red), charge flow across the membrane (via electrochrom-ism) and proton influx into the medium (via phenol red). Hence we completely tracked the protons on their way from the lumen through CF0 into the medium.
A first treatment with EDTA removed up to 12% of total CF1 without increasing the proton permeability of the membranes. A 2nd EDTA treatment removed further 20% of CF1 and increased the proton permeability of membranes by 3 orders of magnitude. In control thylakoids a pH transient decayed with a relaxation time of 20–30 s. After the 2nd EDTA wash the decay time was 85 ms if the electric potential difference was shunted (e.g. by added gramicidin) and 7 ms if it was present. We calculated a lower limit of the protonic unit conductance of CF0 under the assumption that all exposed CF0 were proton conducting. It was 10 fS, corresponding to the passage of 6200 protons/s per CF0 (at 100 mV electric driving force) and by orders of magnitude higher than so far reported for any F0 channel.
KeywordsElectron Transport Chain Thylakoid Membrane Proton Conductance Proton Flux Electric Potential Difference
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