Effects of arginine modification on the photocycle of halorhodopsin.
Exhaustive reaction with phenylglyoxal removed 9 of the 12 arginine and 1 of the 2 lysine residues in detergent-solubilized halorhodopsin, without affecting the chromophore. The consequences of this extensive removal of positive charges on various chloride-binding equilibria and the photochemistry were evaluated. No significant effects were seen on the affinity of Site I to chloride and on the increase in the pKa of Schiff-base deprotonation, which is caused by the chloride binding at this site. No significant effects were seen on the affinity of Site II to chloride, either. However, the photocycle of the pigment was affected. Kinetic modeling of the observed changes in flash-induced absorption changes suggests that the modification increases the affinity of the main halorhodopsin photointermediate to chloride by about fourfold. If chloride translocation involves release of chloride from this intermediate during the transport cycle, the result might explain the observed partial inhibitory effects on chloride transport. Plausible models of chloride translocation include reversible binding of the anion by positively charged groups, strategically arranged in the protein. The results indicate that two of the three spectroscopically observable chloride-dependent equilibria do not depend on a large number of positively charged residues in the protein. To the extent that the unaffected equilibria represent association and dissociation which occur during chloride translocation, at least part of the chloride translocation might be accomplished with the participation of only a few positively charged residues.
D001120:Arginine / Q000037:antagonists & inhibitors / Q000502:physiology*; D001436:Bacteriorhodopsins / Q000502:physiology*; D001665:Binding Sites; D002338:Carotenoids / Q000502:physiology*; D002712:Chlorides / Q000502:physiology; D025602:Halorhodopsins; D007473:Ion Channels / Q000187:drug effects; D007700:Kinetics; D008027:Light; D010658:Phenylglyoxal / Q000494:pharmacology; D013057:Spectrum Analysis; D013997:Time Factors
D002712:Chlorides; D025602:Halorhodopsins; D007473:Ion Channels; D002338:Carotenoids; D001436:Bacteriorhodopsins; D001120:Arginine; D010658:Phenylglyoxal