Carbon-13 nuclear magnetic resonance studies on lectin-carbohydrate interactions: binding of specifically carbon-13-labeled methyl .beta.-D-lactoside to peanut agglutinin

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DOIResolve DOI: http://doi.org/10.1021/bi00532a013
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TypeArticle
Journal titleBiochemistry
ISSN0006-2960
1520-4995
Volume21
Issue3
Pages498503
AbstractThe binding of methyl β-D-lactoside, specifically ¹³C-labeled at C-1 of the D-galactose residue, to peanut agglutinin was studied by carbon-13 nuclear magnetic resonance. The high level of ¹³C enrichment and the high magnetic field employed allowed studies at low ligand concentrations and provided simple single line spectra of very good signal to noise ratios. The ¹³C-l resonance of the disaccharide shows line broadening upon binding to the lectin, which was inhibited by an excess of an unlabeled competitive binding sugar. Between 10 and 30ºC, the disaccharide was found to be in slow exchange with the protein. Residence times, rM, dissociation rate constants, kdiss, and association equilibrium constants were obtained from a study of the ¹³C-l line width of the disaccharide in the presence of the protein. Dissociation rate constants for the disaccharide were similar in magnitude to those found for the α and β anomers of methyl D-galactoside but showed a steeper temperature dependence. The activation enthalpy for dissociation, 13.4 kcal mol⁻¹, is larger than that for methyl β-D-galactopyranoside, while the activation entropy is less negative. The free energy of activation for dissociation is very close to that for both methyl α- and β-D-galactopyranoside. The association rate constants for the disaccharide at different temperatures, as calculated from the dissociation rate constants and from association constants determined independently, are also very similar to those found for the monosaccharide. The association activation parameters, ΔH⁺ = -3.5 kcal mol⁻¹ and TΔS⁺ = -14.6 kcal mol⁻¹, and the fact that the activation enthalpy for dissociation is smaller than the total enthalpy change are further evidence for the previously proposed two-step model for the binding of saccharides to peanut agglutinin. The larger dissociation activation enthalpy and total enthalpy change on binding of the disaccharide, as compared to the monosaccharide, indicate an extended binding site on peanut agglutinin.
Publication date
LanguageEnglish
AffiliationNRC Institute for Biological Sciences; National Research Council Canada
Peer reviewedYes
NRC numberNEUROHR1982
NPARC number9360712
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Record identifierc625a7db-fabc-48ec-900f-598a2dfa0f0a
Record created2009-07-10
Record modified2017-03-24
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