Proton nuclear magnetic resonance studies of the binding of sialosides to intact influenza virus.

The dissociation constants for binding of sialic acid derivatives to the hemagglutinin on intact influenza virus were determined using nuclear magnetic resonance (NMR) spectroscopy. The dissociation constants determined with whole virus are similar to, but slightly higher than, those determined with BHA (hemagglutinin released from virus by treatment with the protease bromelain; Sauter et al., 1989, Biochemistry 28, 8388-8396), indicating that the sialic acid binding site is not significantly altered when hemagglutinin is released from virus. Binding was quantified by observing the concentration-dependent broadening of the sialoside resonances in the presence of X-31 virus or alternatively by observing the effect of the sialoside on the resonances of a competitive "reporter" ligand. The glycosidic substituent attached to the sialic acid makes relatively little difference in the affinity of the sialoside for virus: alpha(2,6)-sialyllactose (KD = 2.7 mM) binds only slightly more tightly than alpha(2,3)-sialyllactose (KD = 3.5 mM). However, inversion of the glycosidic center produces a dramatic change in affinity: the dissociation constant for the alpha-methyl glycoside of sialic acid is 4.2 mM, but not binding is observed with the beta-methyl glycoside.