Electrogenic steps of light-driven proton transport in ESR, a retinal protein from Exiguobacterium sibiricum

• ESR is a H+ pump with Lys in the position of the H+ donor to the retinal Schiff base.
• The kinetics of light-induced membrane potential generation by ESR was examined.
• The rates and the electrogenic distances of proton transfer steps were obtained.
• They indicate different mechanism of H + uptake and release in ESR, compared to BR.

A retinal protein from Exiguobacterium sibiricum (ESR) functions as a light-driven proton pump. Unlike other proton pumps, it contains Lys96 instead of a usual carboxylic residue in the internal proton donor site. Nevertheless, the reprotonation of the Schiff base occurs fast, indicating that Lys96 facilitates proton transfer from the bulk. In this study we examined kinetics of light-induced transmembrane electrical potential difference, ΔΨ, generated in proteoliposomes reconstituted with ESR. We show that total magnitude of ΔΨ is comparable to that produced by bacteriorhodopsin but its kinetic components and their pH dependence are substantially different. The results are in agreement with the earlier finding that proton uptake precedes reprotonation of the Schiff base in ESR, suggesting that Lys96 is unprotonated in the initial state and gains a proton transiently in the photocycle. The electrogenic phases and the photocycle transitions related to proton transfer from the bulk to the Schiff base are pH dependent. At neutral pH, they occur with τ 0.5 ms and 4.5 ms. At alkaline pH, the fast component ceases and Schiff base reprotonation slows. At pH 8.4, a spectrally silent electrogenic component with τ 0.25 ms is detected, which can be attributed to proton transfer from the bulk to Lys96. At pH 5.1, the amplitude of ΔΨ decreases 10 fold, reflecting a decreased yield and rate of proton transfer, apparently from protonation of the acceptor (Asp85-His57 pair) in the initial state. The features of the photoelectric potential generation correlate with the ESR structure and proposed mechanism of proton transfer.