Prediction and comparison of the secondary structure of legume lectins

Secondary structure prediction for the 4 legume lectins: Concanavalin A, soybean agglutinin, favabean lectin and lentil lectin, was done by the method of Chou and Fasman. This prediction shows that these four lectins fall into a structurally distinct class of proteins, containing high amounts of β-sheet and β-turns. There is a notable similarity in the gross structure of these proteins; all four of them contain about 40-50% of β-sheet, 35-45 % β-turn and 0-10% of α-helix. When the secondary structure of corresponding residues in each pair of these lectins was compared, there was a striking similarity in the Concanavalin A-soybean agglutinin and favabean lectin-lentil lectin pairs, and considerably less similarity in the other pairs, suggesting that these legume lectins have probably evolved in a divergent manner from a common ancestor. A comparison of the predicted potential β-turn sites also supports the hypothesis of divergent evolution in this class of lectins

Arrangement of Subunits in Peanut Lectin

X-ray intensity data from the native orthorhombic crystals of peanut lectin have been collected using oscillation photography. Rotatioq function studies using data up to a resolution of $4.5 \AA$ indicate that the four subunits in the molecule, which constitute the asymmetric unit in the crystals, are related to one another by three mutually perpendicular noncrystallographic 2-fold axes. Chemical cross-linking experiments ins olution followed by sodium dodecyl sulfate gel electrophoresis, carried out in parallel, suggest that thereis more than one type of intersubunit approach in the molecule. Rotation function and cross-linking studies thus show that the tetrameric molecule of pleactniunt is a dimer of a dimer. The two monomers in a dimer are related by 2a- fold axis. The two dimers are in turn related by another 2-fold axis perpendicular to the one that relates the two monomers in the dimer, endowing the inolecule with 222 $(D_2)$ symmetry

Analysis of Saccharide Binding to Artocarpus integrifolia Lectin Reveals Specific Recognition of T-antigen (\beta D-Gal(1\rightarrow 3)GalNAc

The binding of Artocarpus integrifolia lectin to N-dansylgalactosamine (where dansyl is 5-dimethylaminonaphthalene-1-sulfonyl) leads to a 100\% increase in dansyl fluorescence with a concomitant blue shift in the emission maximum by 10 nm. This binding is carbohydrate-specific and has an association constant of 1.74 \times $10^4 M^{-1}$ at $20^oC$. The lectin has two binding sites for N-dansylgalactosamine. The values of -\Delta H and -\Delta S for the bindinogf N-dansylgalactosamine are in the range of values reported for severa lectin monosaccharide interactions, indicating an absence of non-polar interaction of the dansylmoiety of the sugar with the combining region of the protein. Dissociation of the bound N-dansylgalactosamine from its complex with the lectin and cosequent change in its fluorescence on addition of nonfluorescent sugars allowed evaluation of the association constant for competing ligands. The thermodynamic parameters for the binding of monosaccharides suggest that the OH groups at C-2, C-3, C-4, and C-6 in the D-galactose configuration are important loci for interaction with the lectin. The acetamido group at C-2 of 2-acetamido-2-deoxygalactopyranose and a methoxyl group at C-1 of methyl-\gamma -Dgalactopyranoside are presumably also involved in binding through nonpolar and van der Waals' interactions. The T-antigenic disaccharide Gal\beta 1\rightarrow 3GalNAc binds very strongly to the lectinw hen compared with methyl-\beta -D-galactopyranoside, the \beta ( 1\rightarrow 3)-linked disaccharides such as Gal\beta 1\rightarrow 3GalNAc, and the \beta (1\rightarrow 4)- linked disaccharides, N-acetyllactosamine andla ctose. The major stabilizing force for the avid binding of Tantigenic disaccharide appears to be a favorable enthalpic contribution. The combining site of the lectin is, therefore, extended. These data takent ogether suggest that the Artocarpus lectin is specific toward the Thomsen-Friedenreich (T) antigen. There are subtle differences in the overaltlo pography of its combining site when compared with that of peanut (Arachis hypogaea)agglutinin.The results of stopped flow spectrometry for the binding of N-dansylgalactosamine to the Artocarpus lectin are consistent with a simple single-step bimolecular association and unimolecular dissociation rate processes. The value of $K_{+1}$ and $K_{-1}$, at $21 ^oC$ are 8.1 X $10^5 M^{-1} s^{-1}$ and 50 $s^{-1}$, respectively. The activation parameters indicate an enthalpy-controlled association process

Saccharide binding to three Gal/GalNAc specific lectins: Fluorescence, spectroscopic and stopped-flow kinetic studies

Fluorescence and stopped-flow spectrophotometric studies on three plant lectins fromPsophocarpus tetragonolobus (winged bean),Glycine max (soybean) andArtocarpus integrifolia (jack fruit) have been studied usingN-dansylgalactosamine as a fluorescent ligand. The best monosaccharide for the winged bean agglutinin I (WBA I) and soybean (SBA) is Me-agrGalNAc and for jack fruit agglutinin (JFA) is Me-agrGal. Examination of the percentage enhancement and association constants (1.51×106, 6.56×106 and 4.17×105 M–1 for SBA, WBA I and JFA, respectively) suggests that the combining regions of the lectins SBA and WBA I are apolar whereas that of JFA is polar. Thermodynamic parameters obtained for the binding of several monosaccharides to these lectins are enthalpically favourable. The binding of monosaccharides to these lectins suggests that the-OH groups at C-1, C-2, C-4 and C-6 in thed-galactose configuration are important loci for interaction with these lectins. An important finding is that the JFA binds specifically to Galß1-3GaINAc with much higher affinity than the other disaccharides which are structurally and topographically similar.The results of stopped-flow spectrometry on the binding ofN-dansylgalactosamine to these lectins are consistent with a bimolecular single step mechanism. The association rate constants (2.4×105, 1.3×104, and 11.7×105 M–1 sec–1 for SBA, WBA I and JFA, respectively) obtained are several orders of magnitude slower than the ones expected for diffusion controlled reactions. The dissociation rate constants (0.2, 3.2×10–2, 83.3 sec–1 for SBA, WBA I and JFA, respectively) obtained for the dissociation ofN-dansylgalactosamine from its lectin complex are slowest for SBA and WBA I when compared with any other lectin-ligand dissociation process

Saccharide binding to three Gal/GalNAc specific lectins: Fluorescence, spectroscopic and stopped-flow kinetic studies

Fluorescence and stopped-flow spectrophotometric studies on three plant lectins fromPsophocarpus tetragonolobus (winged bean),Glycine max (soybean) andArtocarpus integrifolia (jack fruit) have been studied usingN-dansylgalactosamine as a fluorescent ligand. The best monosaccharide for the winged bean agglutinin I (WBA I) and soybean (SBA) is Me-αGalNAc and for jack fruit agglutinin (JFA) is Me-αGal. Examination of the percentage enhancement and association constants (1.51×10⁶, 6.56×10⁶ and 4.17×10⁵ M⁻¹ for SBA, WBA I and JFA, respectively) suggests that the combining regions of the lectins SBA and WBA I are apolar whereas that of JFA is polar. Thermodynamic parameters obtained for the binding of several monosaccharides to these lectins are enthalpically favourable. The binding of monosaccharides to these lectins suggests that the-OH groups at C-1, C-2, C-4 and C-6 in thed-galactose configuration are important loci for interaction with these lectins. An important finding is that the JFA binds specifically to Galß1-3GaINAc with much higher affinity than the other disaccharides which are structurally and topographically similar. The results of stopped-flow spectrometry on the binding ofN-dansylgalactosamine to these lectins are consistent with a bimolecular single step mechanism. The association rate constants (2.4×10⁵, 1.3×10⁴, and 11.7×10⁵ M⁻¹ sec⁻¹ for SBA, WBA I and JFA, respectively) obtained are several orders of magnitude slower than the ones expected for diffusion controlled reactions. The dissociation rate constants (0.2, 3.2×10⁻², 83.3 sec⁻¹ for SBA, WBA I and JFA, respectively) obtained for the dissociation ofN-dansylgalactosamine from its lectin complex are slowest for SBA and WBA I when compared with any other lectin-ligand dissociation process