Time-resolved detection of light-induced conformational changes of heliorhodopsin

Heliorhodopsins (HeRs) are a new category of rhodopsins. They exist as a dimer and exhibit a characteristic inverted topology. HeRs bind all-trans-retinal as a chromophore in the dark, and its isomerization to the 13-cis form by light illumination leads to a photocyclic reaction involving several photo-intermediates: K, L, M, and O. In this study, the kinetics of conformational changes of HeR from Thermoplasmatales archaeon SG8-52-1 (TaHeR) were studied by the transient grating (TG) and circular dichroism (CD) methods. The TG method reveals that the diffusion coefficient (D) does not change until the O formation suggesting no significant conformation change at the surface of the protein during the early steps of the reaction. Subsequently, D decreases upon the O formation. Although two time constants (202 mu s and 2.6 ms) are observed for the conversion from the M to O by the absorption detection, D decreases only at the first step (202 mu s). Light-induced unfolding of helical structure is detected by the CD method. To examine the contribution of a characteristic helix in the intracellular loop 1 (ICL1 helix), Tyr93 on the ICL1 helix was replaced by Gly (Y93G), and the reaction of this mutant was also investigated. It was found that this replacement partially suppresses the D-change, although the CD-change is almost the same as that of the wild type. These results are interpreted in terms of different sensitivities of TG and CD methods, that is, D is sensitive to the structure of the solvent-exposed surface and selectively observes the conformational change in the ICL1 region. It is suggested that the structure of hydrophilic residues in the ICL1 helix is changed during this process.

Automated summary

Heliorhodopsins (HeRs) are a new category of rhodopsins, which exist as dimers and have an inverted topology. This study investigated the conformational changes of HeRs using the transient grating (TG) and circular dichroism (CD) methods. The results showed that there were no significant conformation changes on the protein's surface during the early steps of the reaction, but a decrease in diffusion coefficient (D) was observed after the formation of the O intermediate. The contribution of the ICL1 helix in the conformational changes was examined. The study suggests that the hydrophilic residues in the ICL1 helix undergo structural changes during the process. Rating: 8/10