The binding of monosaccharide inhibitors to hen egg-white lysozyme by proton magnetic resonance at 270 MHz and analysis by ring-current calculations

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Abstract
Studies of the binding of the 4 sugars .alpha.- and .beta.-N-acetyl-D-glucosamine (GlcNAc) and its .alpha.- and .beta.-methyl glycosides to hen egg-white lysozyme (EC 3.2.1.17) by means of high-resolution 1H NMR at 270 MHz are reported. The details of the binding analyses are described in an Appendix. The results show that the sugars bind independently to more than 1 site in lysozyme. The apparent fully bound chemical shifts of the inhibitor proton signals show that, although the major binding modes are generally similar for the 4 sugars, the binding of .alpha.GlcNAc is distinct from that of .alpha.MeGlcNAc and .beta.MeClcNAc. The binding of .beta.GlcNAc is intermediate in character between these 2 modes. The observed shift changes of the inhibitor signals are correlated with the crystal structures of lysozyme-inhibitor complexes by the use of Johnson-Bovey ring-current calculations. Together with consideration of the chemical-shift anisotropy of the GlcNAc amide group, these suggest that the GlcNAc-binding sites in solution are in subsites C and E. The calculations show also that the indole rings of Trp-62 and Trp-63 rotate towards subsite C on the binding of GlcNAc; Trp-108 moves away slightly. These findings indicate a difference between the solution and tetragonal crystal forms of lysozyme-GlcNAc and lysozymes-.beta.MeGlcNAc complexes. In the crystal structure, binding of acetamido monosaccharides is only observed in subsite C and binding in subsite E is prevented by crystal packing.