Direct observation of protein motion during the photochemical reaction cycle of bacteriorhodopsin

Abstract

Platinum-coated, conductive atomic force microscope cantilevers were used to deposit electrophoretically purple membranes from Halobacterium salinarum on the bottom part of the cantilevers. By illuminating the bacteriorhodopsin-containing purple membranes, the protein goes through its photochemical reaction cycle, during which a conformational change happens in the protein, changing its shape and size. The size change of the protein acts upon the cantilever by causing its deflection, which can be monitored by the detection system of the atomic force microscope. The shape of the signal, the action spectrum of the deflection amplitude, and the blue light inhibition of the deflection all prove that the origin of the signal is the conformational change arising in the bacteriorhodopsin during the photocycle. From the size of the signal, the magnitude of the protein motion could be estimated. Using polarized light, the orientation of the motion was determined, relative to the transition moment of the retinal.