Weak Organic Acid Effect of Bacterial Light-Driven Proton-Pumping Rhodopsin

Microbial rhodopsins are photoreceptor proteins that utilize light to elicit various biological functions. The best-studied microbial rhodopsins are outward proton (H⁺)-pumping rhodopsins, which transport H⁺ from the cytoplasmic to the extracellular side. Recently, the weak organic acid (WOA) effect, specifically the enhancement of pumping activity by WOAs such as acetic acid and indole-3-acetic acid (IAA), was discovered in outward H⁺-pumping rhodopsins from fungi. However, it remains unclear whether the WOA effect exists in nonfungal H⁺-pumping rhodopsins. Here, we revealed that the H⁺-pumping activity of a bacterial outward H⁺ pump rhodopsin, PspR, from the rhizobacterium Pseudomonas putida, is also enhanced by extracellular acetic acid and IAA. Using transient absorption measurements on purified PspR protein, we found that extracellular WOAs accelerate cytoplasmic H⁺ uptake and extracellular H⁺ release from a protonated counterion during its photocycle. Furthermore, acetic acid applied on the cytoplasmic side has an inhibitory effect on the H⁺ pump activity of PspR, which is less significant for IAA and can be mitigated by increasing the H⁺ concentration or introducing a cytoplasmic donor residue. These findings on the WOA effect in a bacterial rhodopsin provide new insights into the physiological function of outward H⁺-pumping rhodopsins in bacteria, particularly in their interaction with plants.