TOPOGRAPHY OF THE COMBINING REGION OF A THOMSEN-FRIEDENREICH-ANTIGEN-SPECIFIC LECTIN JACALIN (ARTOCARPUS-INTEGRIFOLIA AGGLUTININ) - A THERMODYNAMIC AND CIRCULAR-DICHROISM SPECTROSCOPIC STUDY

Therodynamic analysis of carbohydrate binding by Artocarpus integrifolia (jackfruit) agglutinin (jacalin) shows that, among monosaccharides, MeαGalNAc (methyl-α-N-acetylgalactosamine) is the strongest binding ligand. Despite its strong affinity for MeαGalNAc and MeαGal, the lectin binds very poorly when Gal and GalNAc are in α-linkage with other sugars such as in A- and B-blood-group trisaccharides, Galα1-3Gal and Galα1-4Gal. These binding properties are explained by considering the thermodynamic parameters in conjunction with the minimum energy conformations of these sugars. It binds to Galβ1-3GalNAcαMe with 2800-fold stronger affinity over Galβ1-3GalNAcβMe. It does not bind to asialo-G(M1) (monosialoganglioside) oligosaccharide. Moreover, it binds to Galβ1-3GalNAcαSer, the authentic T (Thomsen-Friedenreich)-antigen, with about 2.5-fold greater affinity as compared with Galβ1-3GalNAc. Asialoglycophorin A was found to be about 169333 times stronger an inhibitor than Galβ1-3GalNAcαSer. The present study thus reveals the exquisite specificity of A. integrifolia lectin for the T-antigen. Appreciable binding of disaccharides Glcβ1-3GalNAc and GlcNAcβ1-3Gal and the very poor binding of β-linked disaccharides, which instead of Gal and GalNAc contain other sugars at the reducing end, underscore the important contribution made by Gal and GalNAc at the reducing end for recognition by the lectin. The ligand-structure-dependent alterations of the c.d. spectrum in the tertiary structural region of the protein allows the placement of various sugar units in the combining region of the lectin. These studies suggest that the primary subsite (subsite A) can accommodate only Gal or GalNAc or α-linked Gal or GalNAc, whereas the secondary subsite (subsite B) can associate either with GalNAcβMe or GalβMe. Considering these factors a likely arrangement for various disaccharides in the binding site of the lectin is proposed. Its exquisite specificity for the authentic T-antigen, Galβ1-3GalNAcαSer, together with its virtual non-binding to A- and B-blood-group antigens, Galβ1-3GalNAcβMe and asialo-G(M1) should make A. integrifolia lectin a valuable probe for monitoring the expression of T-antigen on cell surfaces.