MECHANISM OF CHLORIDE TRANSPORT BY HALORHODOPSIN

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We will explore the mechanism by which halorhodopsin functions as a light-driven electrogenic chloride pump. This chloride-transport system, found in the cytoplasmic membrane of halobacteria, is a retinal pigment whose chromophore exhibits flash-induced absorbance changes dependent on the presence of chloride. Furthermore, chloride, as well as other anions, apparently influences the pK of the retinal Schiff's base. Studying these phenomena in more detail, and using specific labels to modify selected amino acid residues, we will describe the groups involved in the binding of chloride, the number of chloride-binding sites and their relationship to one another, and the role of protonation/deprotonation of groups in the chloride translocation. Special attention will be paid to the possibility that halorhodopsin contains a sulfhydryl group with an influence on the photochemistry of the pigment. Although the purified chromoprotein (apparent molecular weight 21 kDa) of halorhodopsin binds chloride, and shows the chloride-dependent effects observed with the intact system, we will search for and identify any other, so far unknown, components of halorhodopsin. In this study we expect to establish the basis for a first model of the molecular mechanism of how the photochemical changes are related to chloride translocation against an electrochemical gradient. Such a model should contribute to the understanding of ionic pumps in general.