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1.
Photochem Photobiol Sci ; 22(8): 1809-1823, 2023 Aug.
Article in English | MEDLINE | ID: mdl-37036621

ABSTRACT

A putative xanthorhodopsin-encoding gene, XR34, was found in the genome of the moderately halophilic gammaproteobacterium Salinivibrio socompensis S34, isolated from modern stromatolites found on the shore of Laguna Socompa (3570 m), Argentina Puna. XR-encoding genes were clustered together with genes encoding X-carotene, retinal (vitamin-A aldehyde), and carotenoid biosynthesis enzymes while the carotene ketolase gene critical for the salinixanthin antenna compound was absent. To identify its functional behavior, we herein overexpressed and characterized this intriguing microbial rhodopsin. Recombinant XR34 showed all the salient features of canonical microbial rhodopsin and covalently bound retinal as a functional chromophore with λmax = 561 nm (εmax ca. 60,000 M-1 cm-1). Two canonical counterions with pK values of around 6 and 3 were identified by pH titration of the recombinant protein. With a recovery time of approximately half an hour in the dark, XR34 shows light-dark adaptation shifting the absorption maximum from 551 to 561 nm. Laser-flash induced photochemistry at pH 9 (deprotonated primary counterion) identified a photocycle starting with a K-like intermediate, followed by an M-state (λmax ca. 400 nm, deprotonated Schiff base), and a final long wavelength-absorbing N- or O-like intermediate before returning to the parental 561 nm-state. Initiating the photocycle at pH 5 (protonated counterion) yields only bathochromic intermediates, due to the lacking capacity of the counterion to accept the Schiff base proton. Illumination of the membrane-embedded protein yielded a capacitive transport current. The presence of the M-intermediate under these conditions was demonstrated by a blue light-induced shunt process.


Subject(s)
Bacteriorhodopsins , Schiff Bases , Schiff Bases/chemistry , Carotenoids/metabolism , Retinaldehyde/chemistry , Rhodopsins, Microbial/genetics , Rhodopsins, Microbial/chemistry , Rhodopsins, Microbial/metabolism , Hydrogen-Ion Concentration
2.
Bioessays ; 44(10): e2100210, 2022 Oct.
Article in English | MEDLINE | ID: mdl-35986436

ABSTRACT

The osmotic activity produced by internal, non-permeable, anionic nucleic acids and metabolites causes a persistent and life-threatening cell swelling, or cellular edema, produced by the Gibbs-Donnan effect. This evolutionary-critical osmotic challenge must have been resolved by LUCA or its ancestors, but we lack a cell-physiology look into the biophysical constraints to the solutions. Like mycoplasma, early cells conceivably preserved their volume with Cl- , Na+ , and K+ -channels, Na+ /H+ -exchangers, and a light-dependent bacteriorhodopsin-like H+ -pump. Here, I simulated protocells having these ionic-permeabilities and inhabiting an oceanic pond before the Great-Oxygenation-Event. Protocells showed better volume control and stable resting potentials at lower external pH and higher temperatures, favoring a certain type of extremophile life. Prevention of Na+ -influx at night, with low bacteriorhodopsin activity, required deep shutdown of highly voltage-sensitive Na+ -channels and extremely selective K+ -channels, two conserved features essential for modern neuronal encoding. The Gibbs-Donnan effect universality implies that extraterrestrial cells, if they exist, may reveal similar volume-controlling mechanisms.


Subject(s)
Bacteriorhodopsins , Nucleic Acids , Cell Physiological Phenomena , Membrane Potentials/physiology , Sodium/metabolism
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