Evidence for a sulfhydryl group near the retinal-binding site of halorhodopsin.
Amino acid analysis of the halorhodopsin chromoprotein shows that this protein contains a cysteine residue. Such a residue is absent in bacteriorhodopsin. Low concentrations (micromolar) of HgCl2 inhibit light-dependent chloride transport by halorhodopsin in envelope vesicles prepared from Halobacterium halobium strain L-33 and increase the Km for chloride. The decay rate of the flash-induced absorption change of halorhodopsin, measured at 570 nm, is considerably slowed by HgCl2, and this effect is reversed at higher concentrations of chloride. In addition, the magnitude of the absorption changes is diminished by HgCl2. These effects of the mercurial are also seen with the purified, solubilized chromoprotein. Upon addition of HgCl2 to the chromoprotein at low chloride concentrations in the dark, a decrease of absorption at 580 nm and an increase at 380 nm occur, as well as a blue shift of the chromophore by about 20 nm. Sustained illumination of halorhodopsin results in a 410 nm photoproduct. The reconversion of this species to 580 nm in the dark is strongly inhibited by HgCl2. These results show that a thiol group is essential for the stability of the halorhodopsin chromophore and for its photochemical reactions and suggest that this group is in the vicinity of both the retinal Schiff's base and the chloride-binding site.
D000596:Amino Acids / Q000032:analysis; D001436:Bacteriorhodopsins / Q000378:metabolism*; D001665:Binding Sites; D002338:Carotenoids / Q000378:metabolism*; D003624:Darkness; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D006217:Halobacterium / Q000378:metabolism; D025602:Halorhodopsins; D007700:Kinetics; D008027:Light; D008627:Mercuric Chloride; D008628:Mercury / Q000494:pharmacology; D011485:Protein Binding; D012172:Retinaldehyde / Q000378:metabolism*; D012176:Retinoids / Q000378:metabolism*; D013053:Spectrophotometry; D013438:Sulfhydryl Compounds / Q000032:analysis
D000596:Amino Acids; D025602:Halorhodopsins; D012176:Retinoids; D013438:Sulfhydryl Compounds; D002338:Carotenoids; D001436:Bacteriorhodopsins; D008627:Mercuric Chloride; D004228:Dithionitrobenzoic Acid; D008628:Mercury; D012172:Retinaldehyde