Determination of binding constants for N-acetyl-D-glucosamine oligomers with lysozyme

A method to determine intrinsic binding constants of lysozyme with substrate analogues such as N-acetyl-D-glucosamine dimer and trimer is proposed. The method is based on the competitive interaction of an anionic azo dye with substrate analogues for lysozyme. There are two binding sites for substrate analogues and dyes, respectively, on lysozyme. One binding mode of the substrate analogues to subsites D-F on lysozyme was non-competitive, and another binding mode to subsites A-C was competitive with the dye. From the binding constants obtained it is suggested that the binding of the substrate analogues to subsite D on lysozyme is weaker than the binding to the other subsites.