Chromophore structure in a long-lived intermediate of heliorhodopsins : switching of a hydrogen bonding partner of protonated Schiff base

Abstract

Heliorhodopsin (HeR) is a novel class of retinal proteins discovered in 2018 [1]. HeR contains an all-trans-retinal as a chromophore, which is covalently bound to a lysine residue through a protonated Schiff base linkage. Although amino acid sequences of HeR are largely different from those of type1 rhodopsins, a family of HeR shares the seven-transmembrane helix motif. Photoexcitation of the all-trans-retinal chromophore results in isomerization to a 13-cis form. This isomerization initiates a photocycle involving a series of intermediates, which are similar to those observed for type-1 rhodopsins. HeR has a long-lived intermediate in its photocycle, which is named the O intermediate, suggesting that the function of HeR is light sensing. In order to understand the functional role of the O intermediate, we investigated the chromophore structure in two HeRs, HeR 48C12 and T. archaeon HeR, using time-resolved resonance Raman spectroscopy. We measured resonance Raman spectra of the O intermediate of the two HeRs in H2O and D2O (Figure 1). The observed spectra of the O intermediate of the both HeRs showed that the chromophore configuration is 13-cis and 15-anti and that the polyene chain around the Schiff base is distorted. A comparison of frequencies and bandwidths of the C=N stretch bands of the O intermediate in H2O and D2O solutions indicated that the Schiff base is protonated and hydrogenbonded to a water molecule. The latter feature is in contrast to that the hydrogen bond partner is amino acid residue in the unphotolyzed state [2]. Therefore, it is likely that the protein structure around the Schiff base is much altered in the transition from the O intermediate to the unphotolyzed state. This alternation may be a reason for the long lifetime of the O intermediate, which is essential to signal transduction.