Electric-field effects in dry films of D85N and D85,96N mutant bacteriorhodopsin

Abstract

In the D85N mutant of the protein bacteriorhodopsin (BR), the Schiff base, by which the retinal chromophore is bound to the protein, exhibits an abnormally low proton affinity (p K a≈8.9). Recent experiments on thin films of this protein have shown that this causes the protonation state of the Schiff base, and thus the visible absorption spectrum, to be sensitive to external electric fields. In this paper, we explore the dependence of this effect on parameters such as pH, humidity, and film thickness. The results of these experiments point to the importance of water molecules bound in the acceptor part of the proton channel as sources and donors in field-induced proton-transfer reactions. We describe additional results obtained with the D85,96N mutant, which also exhibits a low Schiff-base p K. The similar behavior of the two mutants under applied electric fields at high pH implies that the residue Asp-96 plays no role in field-induced Schiff-base protonation.