Isolation, macromolecular properties, and combining site of a chito-oligosaccharide-specific lectin from the exudate of ridge gourd (Luffa acutangula).
A lectin specific for chito-oligosaccharides from the exudate of ridge gourd (Luffa acutangula) fruits has been purified to homogeneity by affinity chromatography. The lectin has a molecular weight of 48,000, an S(0)20,w of 4.06 S and a Stokes radius of 2.9 nm. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis, a single band corresponding to Mr of 24,000 was observed both in the presence and absence of beta-mercaptoethanol. The subunits in this dimeric lectin are, therefore, held together solely by noncovalent interactions. The lectin is not a glycoprotein, and secondary structure analysis by CD measurements showed 31% alpha-helix. The hemagglutinating activity of L. acutangula agglutinin was not inhibited by any of the monosaccharides tested. Among the disaccharides only di-N-acetylchitobiose was inhibitory. The inhibitory potency of chito-oligosaccharides increased dramatically with their size up to penta-N-acetylchitopentaose. The lectin has two binding sites for saccharides. The affinity of chito-oligosaccharides for L. acutangula lectin, as monitored by titrating the changes in the near UV-CD spectra and intrinsic fluorescence, increased strikingly with the number of GlcNAc units in them. The values of delta G, delta H, and delta S for the binding process showed a pronounced dependence on the size of the chito-oligosaccharides, indicating that the binding of higher oligomers is progressively more favored thermodynamically than di-N-acetylchitobiose. The thermodynamic data are consistent with an extended binding site in this lectin, which accommodates a tetrasaccharide.
D000117:Acetylglucosamine; D002686:Chitin; D006384:Hemagglutination; D037102:Lectins / Q000302:isolation & purification*; D046911:Macromolecular Substances; D008970:Molecular Weight; D009844:Oligosaccharides; D037121:Plant Lectins; D011487:Protein Conformation; D013329:Structure-Activity Relationship
D037102:Lectins; D046911:Macromolecular Substances; D009844:Oligosaccharides; D037121:Plant Lectins; C050365:chito-oligosaccharide-specific Luffa-lectin; D002686:Chitin; D000117:Acetylglucosamine