Light-driven chloride ion transport by halorhodopsin from Natronobacterium pharaonis. 2. Chloride release and uptake,...

The photocycle of the light-driven chloride pump, N. pharaonis halorhodopsin, is described by the scheme HR-->K--><==>L<==>N<==>O<==>HR'-->HR. From the chloride dependencies of the rate constants in this model we identify the N-->O and O-->HR' reactions as the steps where chloride release and uptake occur, respectively, during the transport. The dependencies of the rate constants on temperature describe a thermodynamic cycle in which enthalpy-entropy conversion occurs in the O-->HR' reaction. The dependencies of the rate constants on hydrostatic pressure indicate that a substantial volume decrease occurs at the L-->N reaction, a result of a large-scale conformational change. This is the opposite of the volume increase in the photocycle of the proton pump, bacteriorhodopsin, that is implicated in the access change of the active site during the transport and the passage of a proton from the cytoplasmic surface to the active site. The results together suggest a chloride transport mechanism, in which the equivalents of all the ion transfer steps in bacteriorhodopsin occur but in the reverse sense, so as to cause the extracellular-to-cytoplasmic translocation of a chloride ion instead of the cytoplasmic-to-extracellular transport of a proton.