Coexistence of light-driven Na and H transport in a microbial rhodopsin from Nonlabens dokdonensis.

Ion pumping microbial rhodopsins are photochemically active membrane proteins, converting light energy into ion-motive-force for ATP synthesis. Nonlabens dokdonensis rhodopsin 2 (NdR2), was recently identified as a light-driven Na pump. However, few functional studies on NdR2 have been conducted to elucidate its mechanism of ion transport. By reconstituting NdR2 into liposomes, we proved that NdR2 functions as a light-driven Na/H pump. As Na concentration increased, the dominant H pump activity switched to the Na pump activity at neutral pH. The inversion of pH change by the addition of CCCP at low Na further suggested that the transport of Na and H should coexist in NdR2. By increasing H concentration, the affinity for Na lowered, which was indicated by an increase in K from ~31mM at pH ~7.5, to ~74mM at pH ~6.5. These results demonstrated that Na transport competed with H transport in NdR2, which was confirmed by the dominant H pump activity at pH ~5.7. Kinetic experiments using pyranine uncovered a transient H uptake, followed by an H release at the millisecond time scale in both Na and K solutions. Therefore, these NdR2 results may provide functional and kinetic insights into the ion transport mechanism in light-driven Na pumps.