STUDY OF THE PHOTOCYCLE AND CHARGE MOTIONS OF THE BACTERIORHODOPSIN MUTANT D96N

Absorption kinetic and electric measurements were performed on oriente d purple membranes of D96N bacteriorhodopsin mutant embedded in polyac rylamide gel and the kinetic parameters of the photointermediates dete rmined. The rate constants, obtained from fits to time-dependent conce ntrations, were used to calculate the relative electrogenicity of the intermediates. The signals were analyzed on the basis of different pho tocycle models. The preferred model is the sequential one with reversi ble reaction. To improve the quality of the fits the necessity of intr oducing a second L intermediate arose. We also attempted to interpret our data in the view of reversible reactions containing two parallel p hotocycles, but the pH dependencies of the rate constants and electrog enicities favored the model containing sequential reversible transitio ns. A fast equilibrium for the L2 <-- --> M1 transition and a strong p H dependence of the M2 electrogenicity was found, indicating that the M1 to M2 transition involves complex charge motions, as is expected in a conformational change of the protein.