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Abstract

Bacteriorhodopsin (BR) is a retinal-binding protein in the purple membrane of Halobacteria where it acts as a light-driven proton pump. The quantum yield, γB, of the primary photoreaction of BR which initiates a cyclic reaction of the chromophore was measured. A liquid suspension of purple membranes was flown across a cw laser beam where the BR chromophore was partially photolyzed. After the sample had left the photolysis beam it was probed by recording resonance Raman spectra in the region of the strong and characteristic bands of the retinal chromophores (1500–1600 cm−1). The ethylenic band of the parent species at 1529 cm−1 could be separated with good accuracy from those of its photoproducts by fitting procedures. In this way the relative intensity decrease of the 1529 cm−1 band was monitored as a function of the power of the photolysis beam. From the quantitative relation between these two quantities which besides γB still contains the intensity profile of the photolysis laser, the velocity of the flowing sample and the extinction coefficient of the chromophore, the quantum yield could be deduced. From photolysis experiments at 502, 514, 568 and 647 nm, a mean value of 0.67 was obtained for γB. Combining this value with literature data for the quantum yield of proton release from purple membranes it was concluded that on the average not more than one proton is translocated across the membrane per each photochemical reaction cycle of BR. Our value of 0.67 for γB is of the same magnitude as the quantum yield for the primary photochemical reaction of the retinal chromophore in rhodopsin which initiates the visual process in photoreceptor cells of vertebrates.