TEMPERATURE AND PH SENSITIVITY OF THE O640 INTERMEDIATE OF THE BACTERIORHODOPSIN PHOTOCYCLE

The temperature and pH dependencies of the O640 intermediate of the photocycle of bacteriorhodopsin (bR) were investigated by flash photolysis and T-jump experiments. The maximal concentration of the O640 intermediate was found to be dependent on the temperature, which is described by a sigmoidal relationship. With increasing pH the midpoint of the sigmoidal curves shifts to higher temperatures. The Van't Hoff equation provides enthalpy and entropy values of the observed states. These results indicate that, in the investigated temperature (0-60-degrees-C) and pH (pH 4.0-10.0) range, the sequence of the principal intermediates in the pathway "M-N-O-bR" does not change. The observations of the O640 intermediate at pH < 8.0 and of the N550 intermediate at pH > 8.0 are most probably due only to changes of the intrinsic rate constants of the bR photocycle, not to a different mechanism.