Stopped-flow fluorescence studies on saccharide binding to lysozyme.

The binding of the beta-1-4-linked trimer of N-acetyl-D-glucosamine to hen egg-white lysozyme was studied by rapid-reaction-kinetic methods with tryptophyl fluorescence observation of the transients. It was found that discrete segments of the fluorescence-difference spectrum from this reaction were perturbed at different time-points during the binding process. The results were interpretated as the formation of the initial complex, the fast phase of the reaction, perturbing the environment of tryptophan-62 and a subsequent and slower rearrangement of the initial complex perturbing the environment of tryptophan-108. At pH 4.4, the release of protons from aspartate-101 occurred during the rearrangement step of the binding reaction. A model for the reaction is presented (E, enzyme; L, ligand): (see article) The association of this ligand with lysozyme may be visualized in three-dimensional terms as initial complex-formation across the top of the active-site cleft followed by a diving motion of the ligand into the cleft.