Thermodynamic and kinetic studies on saccharide binding to soya-bean agglutinin.
The fluorescence of N-dansylgalactosamine [N-(5-dimethylaminonaphthalene-1-sulphonyl)galactosamine] was enhanced 11-fold with a 25 nm blue-shift in the emission maximum upon binding to soya-bean agglutinin (SBA). This change was used to determine the association constants and thermodynamic parameters for this interaction. The association constant of 1.51 X 10(6) M-1 at 20 degrees C indicated a very strong binding, which is mainly due to a relatively small entropy value, as revealed by the thermodynamic parameters: delta G = -34.7 kJ X mol-1, delta H = -37.9 kJ X mol-1 and delta S = -10.9 J X mol-1 X K-1. The specific binding of this sugar to SBA shows that the lectin can accommodate a large hydrophobic substituent on the C-2 of galactose. Binding of non-fluorescent ligands, studied by monitoring the fluorescence changes when they are added to a mixture of SBA and N-dansylgalactosamine, indicates that a hydrophobic substituent at the anomeric position increases the affinity of the interaction. The C-6 hydroxy group also stabilizes the binding considerably. Kinetics of binding of N-dansylgalactosamine to SBA studied by stopped-flow spectrofluorimetry are consistent with a single-step mechanism and yielded k+1 = 2.4 X 10(5) M-1 X s-1 and k-1 = 0.2 s-1 at 20 degrees C. The activation parameters indicate an enthalpicly controlled association process.
D000116:Acetylgalactosamine / Q000378:metabolism; D050260:Carbohydrate Metabolism; D003619:Dansyl Compounds; D005456:Fluorescent Dyes; D005688:Galactosamine / Q000031:analogs & derivatives; D007700:Kinetics; D037102:Lectins / Q000378:metabolism*; D037121:Plant Lectins; D030262:Soybean Proteins; D013025:Glycine max / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics; D010835:Phytohemagglutinins
D000116:Acetylgalactosamine; D003619:Dansyl Compounds; D005456:Fluorescent Dyes; D005688:Galactosamine; D037102:Lectins; D037121:Plant Lectins; D030262:Soybean Proteins; C028824:soybean lectin; D010835:Phytohemagglutinins; C041456:dansylgalactosamine