Dynamic Regulation of the 7TM Receptor Bacteriorhodopsin Function under Alkaline pH Conditions

Abstract

Bacteriorhodopsin (bR), a light-driven proton pump for light energy capture in Halobacterium Salinarum, has been studied by multidisciplinary approaches to reveal the molecular machinery of proton transfer and photocycle. In order to study the functionally related structural change in the M intermediate state, different media with a high pH value have been used to extend the lifetime of the M state. However, not much attention have been paid to the conformational changes that occur around the retinal binding pocket and expand further to the overall structure in the ground state. In this study, dynamic regulation of bR function under alkaline pH conditions has been investigated by magic-angle spinning solid-state NMR through chemical shifts, torsion angle and C-H bond length measurements on reconstituted [10,11,14,15-13C4]-retinal-labelled bR purple membrane samples. Combined with UV spectroscopy, light-induced transient absorption changes and molecular dynamic simulations, possible perturbation mechanism from the retinal binding pocket to the overall structure have been discussed. Our results show that C14 of the retinal chromophore is more sensitive to the environmental change than any other carbon sites due to its adjacent position to the C13 pivot. Elongation and attenuation of the M state may be caused by the shift of trans-cis thermal equilibrium to a more populated cis configuration in the dark-adapted purple membrane. Deprotonation of the alkaline residues on the helices C, D, F and G may result in the reformation of the H-bond network on both proton release and uptake channels and further the cause of elongation and attenuation of the M and N states, and vanish of the O state in the bR photocycle.