REPLACEMENT EFFECTS OF NEUTRAL AMINO-ACID-RESIDUES OF DIFFERENT MOLECULAR VOLUMES IN THE RETINAL BINDING CAVITY OF BACTERIORHODOPSIN ON THEDYNAMICS OF ITS PRIMARY PROCESS

We have determined the rate and quantum yield of retinal photoisomeriz ation, the spectra of the primary transients, and the energy stored in the K intermediate in the photocycle of some bacteriorhodopsin mutant s (V49A, A53G, and W182F) in which residue replacements are found to c hange the Schiff base deprotonation kinetics (and thus the protein-ret inal interaction). Because of their change in the local volume resulti ng from these individual replacements, these substitutions perturb the proton donor-acceptor relative orientation change and thus the Schiff base deprotonation kinetics. These replacements are thus expected to change the charge distribution around the retinal, which controls its photoisomerization dynamics. Subpicosecond transient spectroscopy as w ell as photoacoustic technique are used to determine the retinal photo isomerization rate, quantum yield, and the energy stored in the K-inte rmediate for these mutants. The results are compared with those obtain ed for wild-type bacteriorhodopsin and other mutants in which charged residues in the cavity are replaced by neutral ones. In some of the mu tants the rate of photoisomerization is changed, but in none is the qu antum yield or the energy stored in the K intermediate altered from th at in the wild type. These results are discussed in terms of the shape s of the potential energy surfaces of the excited and ground states of retinal in the perpendicular configuration within the protein and the stabilization of the positive charge in the ground and the excited st ate of the electronic system of retinal.