Effect of specific carboxyl modifications on the blue acid species and O650 photocycle intermediate of bacteriorhodopsin.

Carboxyl groups were specifically modified in purple membranes using hydrophobic or hydrophilic activating reagents and endogenous or exogenous nucleophiles. Modification of surface residues using a negatively charged nucleophile had no effect on blue-species formation, whereas surface or internal carboxyl modification using an uncharged nucleophile prevented blue-species formation. The carboxyl-modified samples showed poor correlation between blue-species and O650 intermediate. These results indicate that formation of the blue-species requires negative surface charge and is not associated with the O650 intermediate of bacteriorhodopsin, as had been previously postulated.