Bacteriorhodopsin molecular dynamics: photobleaching and chemical cross-linking

Abstract

The conformational capabilities of the in situ transmembrane protein bacteriorhodopsin (bR) were studied by mid-IR linear dichroism of purple membrane (PM) films. Illumination of bR, with a Schiff-base bound retinylidene prosthetic group, is usually accompanied by the vectorial translocation of H/sup +/ across the membrane bilayer. In the presence of hydroxylamine HCl however, illumination hydrolyzes the Schiff-base linkage between bR and its retinylidene chromophore (photobleaching). Analysis of the mid-IR linear dichroism of PM indicates: (1) the /spl alpha/-helical segments in bR are oriented nearly parallel to the PM normal; (2) after photobleaching the PM, the /spl alpha/-helical segments of bR are tilted 24/spl deg/ away from the PM normal; (3) after cross-linking PM with dimethyl adipimidate the /spl alpha/-helical segments of bR are tilted 9/spl deg/ away from the PM normal; (4) after cross-linking and photobleaching, the /spl alpha/-helical segments of bR are tilted 30/spl deg/ away from the PM normal.<>