Structures of the archaerhodopsin-3 transporter reveal that disordering of internal water networks underpins receptor sensitization

Juan F Bada Juarez,Anthony Watts,Igor Schapiro,Tristan O C Kwan,Suliman Adam,Pierre-Emmanuel Milhiet,Javier Vinals,Kin Kuan Hoi,Peter J Judge,Hsin-Yung Yen,James Birch,Anthony Vial,Carol V Robinson,Isabel Moraes,Danny Axford

doi:10.1038/s41467-020-20596-0

Abstract

Many transmembrane receptors have a desensitized state, in which they are unable to respond to external stimuli. The family of microbial rhodopsin proteins includes one such group of receptors, whose inactive or dark-adapted (DA) state is established in the prolonged absence of light. Here, we present high-resolution crystal structures of the ground (light-adapted) and DA states of Archaerhodopsin-3 (AR3), solved to 1.1 Å and 1.3 Å resolution respectively. We observe significant differences between the two states in the dynamics of water molecules that are coupled via H-bonds to the retinal Schiff Base. Supporting QM/MM calculations reveal how the DA state permits a thermodynamic equilibrium between retinal isomers to be established, and how this same change is prevented in the ground state in the absence of light. We suggest that the different arrangement of internal water networks in AR3 is responsible for the faster photocycle kinetics compared to homologs.

Highlights

Many transmembrane receptors have a desensitized state, in which they are unable to respond to external stimuli
AR3 is suitable for these applications, since the protein has been suggested to have faster photocycle kinetics than many of its homologs[13] (including bacteriorhodopsin from Halobacterium salinarum), the current produced by recombinant AR3 expressed in Xenopus oocytes has been measured as comparable to that of bR14
Using quantum mechanical/molecular mechanical (QM/MM) approaches, our study explores the differences between the two states in both the activation energy barrier for conversion between cis and trans retinal, and the equilibrium position between the two isomers

Summary

Introduction

Many transmembrane receptors have a desensitized state, in which they are unable to respond to external stimuli. They enable cells to sense and to respond to their environment by undergoing conformational changes on ligand binding or light absorption In addition to their active and resting states, several receptor proteins have a desensitized or inactive form, in which their responsiveness to external stimuli is reduced. Development of AR3 mutants (commonly termed Arch in the optogenetics field) has been hampered both by the absence of high-resolution structural information[15,16,17,18] and by a lack of understanding of the mechanisms of receptor desensitization[19,20,21,22] It is not a direct homolog of the Class A GPCR rhodopsin, AR3 undergoes a similar, highly ordered sequence of conformational changes (known collectively as the photocycle) after the ground state of the protein is stimulated by a photon of appropriate wavelength. It is a light-induced change in the isomerization state of the retinal, detected by the surrounding receptor, that initiates the progression around the photocycle[23,24,25]

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Conclusion