The carbohydrate moiety of rhodopsin: Lectin-binding, chemical modification and fluorescence studies

Abstract

Rhodopsin contains a covalently attached carbohydrate moiety consisting of three N-acetylglucosamine and three mannose residues (Heller and Lawrence, 1970). We summarize here some of our recent studies of the carbohydrate moiety of bovine rhodopsin. (1) Concanavalin A (Con A) and wheat germ agglutinin bind to carbohydrate residues on dise membranes. The stoichiometry of binding was 1 Con A monomer per retinal, suggesting that the carbohydrate moiety of rhodopsin is located on the surface of disc membranes. Rhodopsin can be separated from some other constituents of the rod outer segment by affinity chromatography on a Con A-agarose column. (2) The carbohydrate unit of rhodopsin was oxidized with periodate. The 500 nm absorption band and the regenerability of this band after bleaching were not markedly altered by periodate oxidation. Thus, an intact carbohydrate unit does not appear to be essential for the chromophoric properties of rhodopsin or for regenerability. (3) The carbohydrate moiety of rhodopsin was oxidized with periodate and then labeled with a fluorescent hydrazide, such as methyl anthranilate hydrazide or fluorescein carbohydrazide. The efficiency of energy transfer from these fluorescent hydrazides to 11- cis retinal revealed that the distance between the carbohydrate group and 11- cis retinal is greater than 50 Å.