Characterization of the carbohydrate binding specificity and kinetic parameters of lectins by using surface plasmon resonance

An accurate, rapid, and sensitive method for characterizing the carbohydrat e binding properties of lectins using a BIAcore apparatus and the detection method of surface plasmon resonance is described. As a model study, the si alic acid binding lectins from Sambucus nigra and Maackia amurensis, which are specific for the epitopes Neu5Ac(alpha 2-6)Gal and Neu5Ac(alpha 2-3)Gal , respectively, were chosen as suitable candidates. Two systems, one for th e analysis of oligosaccharides and the other for glycoproteins, were develo ped after a rigorous analysis and evaluation of such parameters as binding conditions, buffers, and regeneration conditions. The systems take into acc ount nonspecific binding, using the respective denatured lectin as negative blank, and avoid loss of activity: regeneration of the surface using eithe r 10 mM NaOAc (pH 4.3) buffer (oligosaccharide system) or 20 mM HCl (glycop rotein system). The specificity of the lectins is well illustrated, while t he kinetics parameters are shown to be sensitive to subtle changes in the r ecognized epitopes, and to be affected by steric hindrance. Surface plasmon resonance is a suitable technique for the analysis and characterization of lectins. (C) 1999 Academic Press.