RESUMO
Microbial rhodopsins are photoreceptors containing the retinal Schiff base chromophore and are ubiquitous among microorganisms. The Schiff base configuration of the chromophore, 15-anti (CâN trans) or 15-syn (CâN cis), is structurally important for their functions, such as membrane ion transport, because this configuration dictates the orientation of the positively charged NH group that interacts with substrate ions. The 15-anti/syn configuration is thus essential for elucidating the ion-transport mechanisms in microbial rhodopsins. Here, we identified the Schiff base configuration during the photoreaction of a sodium pumping rhodopsin from Indibacter alkaliphilus using Raman spectroscopy. We found that the unique configurational change from the 13-cis, 15-anti to all-trans, 15-syn form occurs between the photointermediates termed O1 and O2, which accomplish the Na+ uptake and release, respectively. This isomerization is considered to give rise to the highly irreversible O1 â O2 step that is crucial for unidirectional Na+ transport.
Assuntos
Rodopsina , Bases de Schiff , Rodopsina/química , Bases de Schiff/química , Íons , Transporte de Íons , Rodopsinas Microbianas , Sódio/químicaRESUMO
Sodium-pumping rhodopsins (NaRs) are membrane transporters that utilize light energy to pump Na+ across the cellular membrane. Within the NaRs, the retinal Schiff base chromophore absorbs light, and a photochemically induced transient state, referred to as the "O intermediate", performs both the uptake and release of Na+. However, the structure of the O intermediate remains unclear. Here, we used time-resolved cryo-Raman spectroscopy under preresonance conditions to study the structure of the retinal chromophore in the O intermediate of an NaR from the bacterium Indibacter alkaliphilus. We observed two O intermediates, termed O1 and O2, having distinct chromophore structures. We show O1 displays a distorted 13-cis chromophore, while O2 contains a distorted all-trans structure. This finding indicated that the uptake and release of Na+ are achieved not by a single O intermediate but by two sequential O intermediates that are toggled via isomerization of the retinal chromophore. These results provide crucial structural insight into the unidirectional Na+ transport mediated by the chromophore-binding pocket of NaRs.