THE EFFECT OF AZIDE ON THE PHOTOCYCLE OF BACTERIORHODOPSIN

Azide has previously been shown to have an effect on the photocycle of bacteriorhodopsin mutants. In appropriate mutants, both the proton re lease from the Schiff base and reprotonation are accelerated. However, no effect on native bacteriorhodopsin has been found. In this work, w e show that protonated azide influences the photocycle of native bacte riorhodopsin, although to a lesser extent than in certain mutants. Sev eral transitions are influenced by protonated azide; however, the over all effect is dominated by the modification of a single step. The M to N transition, i.e. reprotonation of the Schiff base, is accelerated b y azide. As a consequence of this kinetic effect, the concentrations o f several intermediates are influenced. The interpretation of the kine tic effect of azide involves an alternative parallel reaction path fro m the M form to the ground state in native bacteriorhodopsin. It is co ncluded that there are two routes of Schiff base reprotonation: via me diation by Asp-96 (in this case, the molecule returns to the ground st ate through the N intermediate; this is the generally accepted scenari o); via Schiff base reprotonation directly from the external medium (i n this case, the molecule returns from another M form to the ground st ate in one step). (C) 1997 Elsevier Science S.A.