The O intermediate of the photocycle of bacteriorhodopsin (BR) was studied by absorption kinetic measurements at different actinic light densities. With increasing exciting flash intensity, the relative yield of O slightly increases, while that of Mf strongly decreases at the expense of Ms. Kinetic calculations and the optical anisotropy of O show that O can be formed only from Mf although Mf and O have different light intensity dependences. In order to resolve the apparent contradiction, a phenomenologically new cooperative regulatory mechanism seems to be necessary.
