Function of Tyr185 in Stabilizing the Isomerization Equilibrium of the Retinal Chromophore in the Bacteriorhodopsin Ground State

Abstract

Bacteriorhodopsin (bR), a member of the microbial rhodopsin family's seven trans-membrane receptors, functions as a light-driven proton pump for light energy capture and conversion in Halobacterium Salinarum. In the dark-adapted (DA) state, bR contains two thermally interconvertible retinal isomers: 13-cis, 15-syn retinal (bRcis), and all-trans, 15-anti retinal (bRtrans) with a molar ratio of 6:4 in the purple membrane. Absorption of a photon causes photoisomerization of the chromophore from the all-trans to the 13-cis, 15-anti configuration and triggers a series of structural rearrangements of the protein that initiates a vectorial translocation of a proton out of the cell.The function of Tyrosine 185 (Tyr185) in bR photoreaction mechanism has long been a topic of discussion, and been suggested to play an important role in stabilizing the pentagonal hydrogen-bonded network on the extracellular side of the protonated Schiff base during the photoreaction. However, little is to know on coupling of Tyr185 with the protonated Schiff base with respect to the thermal isomerization of the retinal chromophore at the bR ground state. Heteronuclear single molecular manipulation, combined with homonuclear correlation experiments, UV spectroscopy, light-induced transient absorption change measurements, molecular dynamic simulations and mutagenesis, has been established as a powerful tool for providing insight into the conformation and coupling of Tyr185 with the retinal chromophore at the bR ground state in the purple membrane. Our results show that the retinal chromophore in 13-cis and 15-syn configurations has more accumulated charge and, is more twisted and rigid than that in the all-trans configuration. Moreover, the cis-trans thermal isomerization equilibrium with the molar ratio of 6:4 at the ground state is stabilized by coupling with Tyr185.