Membrane damage caused by irradiation of fluorescent concanavalin A.

Abstract

Visible light irradiation of fluoresceinated concanavalin A (f-Con A) bound to the outside of resealed erythrocyte membranes caused crosslinking of up to 50% of the membrane proteins. Crosslinking was absent in controls where equivalent amounts of f-Con A were added to the membranes but prevented from binding by the presence of 10 mM .alpha.-methylmannoside. The photodamage was not accompanied by a change in the membrane permeability barrier or membrane shape. Although fluorescein bleaching accompanies the formation of protein aggregates, the amount of aggregated protein was not simply a function of the number of fluoresceins bleached. The percentage of aggregated protein decreased when the same dose of light was given in a shorter time. Although certain antioxidants and free-radical scavengers had no detected effect on the crosslinking, reducing agents such as cysteamine and reduced glutathione either blocked or reversed the protein crosslinking. The mechanism of photoinduced oxidation and the implications of these results for fluorescence studies of cell membranes are discussed.