Evaluation of the stoichiometry and energetics of carbohydrate binding to Ricinus communis agglutinin: a calorimetric study.
High-sensitivity isothermal titration calorimetry has been used to investigate the thermodynamics of binding of Ricinus communis agglutinin to galactose, lactose and their derivatives in the temperature range 280.5-298 K. The present study unequivocally establishes the carbohydrate-binding stoichiometry of the tetrameric agglutinin from castor bean as two, i.e. the (As-sB)2-type tetramer of the agglutinin has two equivalent sites that are non-interacting and independent. The site binding constants range from 2.2x10(3) M-1 at 282 K for galactose to 4.84x10(4) M-1 at 281 K for N-acetyl-lactosamine. The binding enthalpies range from -21.9 kJ. mol-1 at 293 K for 4-methylumbelliferyl-beta-galactoside to -50.2 kJ. mol-1 at 292.9 K for thiodigalactoside. The observation of limited entropy-enthalpy compensation for binding of the sugars to the lectin indicates that reorganization of water molecules plays an important role in binding. As the slope of the compensation plot is greater than unity, the reactions are largely enthalpically driven. These studies show that the stronger binding of N-acetyl-lactosamine than lactose is due to a favourable interaction between the acetamido group of the reducing-end N-acetylglucosamine of the former and the corresponding loci in the agglutinin molecule. Preferential binding of methyl-beta-galactoside over methyl-alpha-galactoside also indicates the apolar nature of the interaction with the methyl group of the former sugar.
Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
S
S
Sharma
S
S
Bharadwaj
A
A
Surolia
S K
SK
Podder
D002151:Calorimetry; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000737:chemistry*; D002367:Ricinus communis / Q000378:metabolism; D004187:Disaccharides / Q000737:chemistry / Q000378:metabolism; D037102:Lectins / Q000737:chemistry* / Q000378:metabolism; D009005:Monosaccharides / Q000737:chemistry / Q000378:metabolism; D037121:Plant Lectins; D010947:Plants, Toxic; D013816:Thermodynamics; D017186:Titrimetry
D002241:Carbohydrates; D004187:Disaccharides; D037102:Lectins; D009005:Monosaccharides; D037121:Plant Lectins; C017707:Ricinus communis agglutinin-1