Driving forces of proton-pumping rhodopsins.

Proton-pumping rhodopsins are light-driven proton transporters that have been discovered from various microbiota. They are categorized into two groups: outward-directed and inward-directed proton pumps. Although the directions of transport are opposite, they are active proton transporters that create an H⁺ gradient across a membrane. Here, we aimed to study the driving force of the proton-pumping rhodopsins and the effect of ΔΨ and ΔpH on their pumping functions. We systematically characterized the H⁺ transport properties of nine different rhodopsins, six outward-directed H⁺ pumps and three inward-directed pumps, by patch-clamp measurements after expressing them in mammalian cells. The driving force of each pump was estimated from the slope of the current-voltage relations (I-V plot). Notably, among the tested rhodopsins, we found a large variation in driving forces, ranging from 83 to 399 mV. The driving force and decay rate of each pump current exhibited a good correlation. We determined driving forces under various pHs. pH dependency was less than predicted by the Nernst potential in most of the rhodopsins. Our study demonstrates that the H⁺-pumping rhodopsins from different organisms exhibit various pumping properties in terms of driving force, kinetics, and pH dependency, which could be evolutionarily derived from adaptations to their environments.