Retinal-Based Anion Pump from the Cyanobacterium Tolypothrix campylonemoides

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Tatyana I. Rokitskaya^1,a*, Aleksey A. Alekseev², Fedor M. Tsybrov², Sergej M. Bukhalovich², Yuri N. Antonenko^1,b*, and Valentin I. Gordeliy^3,c*

¹Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119991 Moscow, Russia

²Moscow Institute of Physics and Technology, 141701 Dolgoprudny, Moscow Region, Russia

³Institut de Biologie Structurale (IBS), Université Grenoble Alpes, CEA, CNRS, Grenoble, France

^* To whom correspondence should be addressed.

Received June 30, 2023; Revised September 15, 2023; Accepted September 15, 2023

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In this work, TcaR rhodopsin from the cyanobacterium Tolypothrix campylonemoides was characterized. Analysis of the amino acid sequence of TcaR revealed that this protein possesses a TSD motif that differs by only one amino acid from the TSA motif of the known halorhodopsin chloride pump. The TcaR protein was expressed in E. coli, purified, and incorporated into proteoliposomes and nanodiscs. Functional activity was measured by electric current generation through the planar bilayer lipid membranes (BLMs) with proteoliposomes adsorbed on one side of the membrane surface, as well as by fluorescence using the voltage-dependent dye oxonol VI. We have shown that TcaR rhodopsin functions as a powerful anion pump. Our results show that the novel microbial anion transporter, TcaR, deserves deeper investigation and may be of interest both for fundamental studies of membrane proteins and as a tool for optogenetics.

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KEY WORDS: microbial rhodopsin, photosensitive ion pump, optogenetics, proteoliposomes, bilayer lipid membrane

DOI: 10.1134/S0006297923100127

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