Circular dichroism and photocycle kinetics of partially detergent solubilized and partially retinal regenerated bacteriorhodopsin.

The circular dichroism (CD) spectra and the kinetics of the M(412) formation have been determined and compared for bacteriorhodopsin (bR) partially delipidated by the addition of Triton X-100 and partially reconstituted by the addition of retinal to apoprotein at pH 6.8 and 22 degrees C. As the degree of delipidation increases or the fraction of reconstitution decreases, the following observations are made: (a) the shape of the visible CD band changes from biphasic, as found in bR, to a single monomer type band; (b) the CD spectra and the deprotonation rate constants change in a similar way; (c) the relative amplitudes of the components of the deprotonation kinetics do not change upon reconstitution, but change greatly upon delipidation. These results lead to the conclusion that the CD band shape as well as the deprotonation rate constants are sensitive to one type of perturbation, which is linked to the retinal structure within the protein environment, whereas the relative amplitudes of the components of the deprotonation kinetics are sensitive to another type, which may be linked to acid-base equilibria of the amino acid side groups within the active site.