Thermodynamics of lectin-carbohydrate interactions. Titration microcalorimetry measurements of the binding of N-linked carbohydrates and ovalbumin to concanavalin A

Dipak K. Mandal, Nand Kishore, Curtis F. Brewer

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239 Citations (Scopus)

Abstract

The thermodynamics of binding of concanavalin A (Con A) with a series of linear and branched chain oligosaccharides including certain N-linked complex type and oligomannose type carbohydrates and a fraction of quail ovalbumin containing Man7 and Man8 oligomannose chains have been determined using titration microcalorimetry. Methyl 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranoside, a branch chain trisaccharide moiety found in all N-linked carbohydrates which possesses approximately 60-fold higher affinity than methyl α-D-mannopyranoside, exhibited a change in enthalpy of binding (ΔH) of -14.4 kcal mol-1 as compared to -8.2 kcal mol-1 for the monosaccharide. This demonstrates that Con A possesses an extended binding site for the trimannoside. However, a biantennary complex type carbohydrate with terminal β(1,2)-GlcNAc residues which binds with 3-fold higher affinity than the trimannoside possesses a ΔH of only -10.6 kcal mol-1. A plot of -ΔH versus -TΔS for the carbohydrates in the present study showed positive deviations in -TΔS for the complex type carbohydrate, as well as α(1,2)-di- and trimannosyl oligosaccharides which are part of the structures of oligomannose type carbohydrates. The relative favorable changes in binding entropies of these compounds are attributed to the presence of multiple internal and terminal residues in each molecule which can independently bind to the monosaccharide binding site of the lectin. The ΔH values for the complex type carbohydrate and the α(1,2) mannose oligosaccharides were also approximately -2.5 kcal mol-1 greater than that of methyl α-D-mannopyranoside, indicating some extended binding site interactions. The thermodynamics of binding of N-linked oligomannose type carbohydrates to dimeric Con A and its succinyl and acetyl derivatives were determined since these carbohydrates are bivalent and precipitate with the native tetrameric lectin but not with the dimeric protein and its two derivatives. Titration of succinyl-Con A with a Man5 oligomannose type oligosaccharide gave a ΔH of -14.5 kcal mol-1, which is similar to that of the branch chain trimannoside. This indicates that the α(1,6) core arm of Man5 which contains the trimannosyl moiety is the primary binding epitope for Con A. A fraction of quail ovalbumin containing a mixture of Man7 and Man8 chains at a single glycosylation site showed univalent binding to succinyl-Con A and a ΔH of -13.6 kcal mol-1. These results indicate that the trimannoside moiety on the α(1,6) arm(s) of the carbohydrate chains is the primary binding epitope and that its interactions with the lectin are relatively unaffected by the protein matrix of ovalbumin.

Original languageEnglish (US)
Pages (from-to)1149-1156
Number of pages8
JournalBiochemistry
Volume33
Issue number5
StatePublished - 1994

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Ovalbumin
Concanavalin A
Titration
Thermodynamics
Lectins
Carbohydrates
Oligosaccharides
Quail
Monosaccharides
Binding Sites
Mannose
Branched-Chain Oligosaccharides
Epitopes
Derivatives
Glycosylation
Trisaccharides
Entropy
Precipitates
Enthalpy
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

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title = "Thermodynamics of lectin-carbohydrate interactions. Titration microcalorimetry measurements of the binding of N-linked carbohydrates and ovalbumin to concanavalin A",
abstract = "The thermodynamics of binding of concanavalin A (Con A) with a series of linear and branched chain oligosaccharides including certain N-linked complex type and oligomannose type carbohydrates and a fraction of quail ovalbumin containing Man7 and Man8 oligomannose chains have been determined using titration microcalorimetry. Methyl 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranoside, a branch chain trisaccharide moiety found in all N-linked carbohydrates which possesses approximately 60-fold higher affinity than methyl α-D-mannopyranoside, exhibited a change in enthalpy of binding (ΔH) of -14.4 kcal mol-1 as compared to -8.2 kcal mol-1 for the monosaccharide. This demonstrates that Con A possesses an extended binding site for the trimannoside. However, a biantennary complex type carbohydrate with terminal β(1,2)-GlcNAc residues which binds with 3-fold higher affinity than the trimannoside possesses a ΔH of only -10.6 kcal mol-1. A plot of -ΔH versus -TΔS for the carbohydrates in the present study showed positive deviations in -TΔS for the complex type carbohydrate, as well as α(1,2)-di- and trimannosyl oligosaccharides which are part of the structures of oligomannose type carbohydrates. The relative favorable changes in binding entropies of these compounds are attributed to the presence of multiple internal and terminal residues in each molecule which can independently bind to the monosaccharide binding site of the lectin. The ΔH values for the complex type carbohydrate and the α(1,2) mannose oligosaccharides were also approximately -2.5 kcal mol-1 greater than that of methyl α-D-mannopyranoside, indicating some extended binding site interactions. The thermodynamics of binding of N-linked oligomannose type carbohydrates to dimeric Con A and its succinyl and acetyl derivatives were determined since these carbohydrates are bivalent and precipitate with the native tetrameric lectin but not with the dimeric protein and its two derivatives. Titration of succinyl-Con A with a Man5 oligomannose type oligosaccharide gave a ΔH of -14.5 kcal mol-1, which is similar to that of the branch chain trimannoside. This indicates that the α(1,6) core arm of Man5 which contains the trimannosyl moiety is the primary binding epitope for Con A. A fraction of quail ovalbumin containing a mixture of Man7 and Man8 chains at a single glycosylation site showed univalent binding to succinyl-Con A and a ΔH of -13.6 kcal mol-1. These results indicate that the trimannoside moiety on the α(1,6) arm(s) of the carbohydrate chains is the primary binding epitope and that its interactions with the lectin are relatively unaffected by the protein matrix of ovalbumin.",
author = "Mandal, {Dipak K.} and Nand Kishore and Brewer, {Curtis F.}",
year = "1994",
language = "English (US)",
volume = "33",
pages = "1149--1156",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "5",

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TY - JOUR

T1 - Thermodynamics of lectin-carbohydrate interactions. Titration microcalorimetry measurements of the binding of N-linked carbohydrates and ovalbumin to concanavalin A

AU - Mandal, Dipak K.

AU - Kishore, Nand

AU - Brewer, Curtis F.

PY - 1994

Y1 - 1994

N2 - The thermodynamics of binding of concanavalin A (Con A) with a series of linear and branched chain oligosaccharides including certain N-linked complex type and oligomannose type carbohydrates and a fraction of quail ovalbumin containing Man7 and Man8 oligomannose chains have been determined using titration microcalorimetry. Methyl 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranoside, a branch chain trisaccharide moiety found in all N-linked carbohydrates which possesses approximately 60-fold higher affinity than methyl α-D-mannopyranoside, exhibited a change in enthalpy of binding (ΔH) of -14.4 kcal mol-1 as compared to -8.2 kcal mol-1 for the monosaccharide. This demonstrates that Con A possesses an extended binding site for the trimannoside. However, a biantennary complex type carbohydrate with terminal β(1,2)-GlcNAc residues which binds with 3-fold higher affinity than the trimannoside possesses a ΔH of only -10.6 kcal mol-1. A plot of -ΔH versus -TΔS for the carbohydrates in the present study showed positive deviations in -TΔS for the complex type carbohydrate, as well as α(1,2)-di- and trimannosyl oligosaccharides which are part of the structures of oligomannose type carbohydrates. The relative favorable changes in binding entropies of these compounds are attributed to the presence of multiple internal and terminal residues in each molecule which can independently bind to the monosaccharide binding site of the lectin. The ΔH values for the complex type carbohydrate and the α(1,2) mannose oligosaccharides were also approximately -2.5 kcal mol-1 greater than that of methyl α-D-mannopyranoside, indicating some extended binding site interactions. The thermodynamics of binding of N-linked oligomannose type carbohydrates to dimeric Con A and its succinyl and acetyl derivatives were determined since these carbohydrates are bivalent and precipitate with the native tetrameric lectin but not with the dimeric protein and its two derivatives. Titration of succinyl-Con A with a Man5 oligomannose type oligosaccharide gave a ΔH of -14.5 kcal mol-1, which is similar to that of the branch chain trimannoside. This indicates that the α(1,6) core arm of Man5 which contains the trimannosyl moiety is the primary binding epitope for Con A. A fraction of quail ovalbumin containing a mixture of Man7 and Man8 chains at a single glycosylation site showed univalent binding to succinyl-Con A and a ΔH of -13.6 kcal mol-1. These results indicate that the trimannoside moiety on the α(1,6) arm(s) of the carbohydrate chains is the primary binding epitope and that its interactions with the lectin are relatively unaffected by the protein matrix of ovalbumin.

AB - The thermodynamics of binding of concanavalin A (Con A) with a series of linear and branched chain oligosaccharides including certain N-linked complex type and oligomannose type carbohydrates and a fraction of quail ovalbumin containing Man7 and Man8 oligomannose chains have been determined using titration microcalorimetry. Methyl 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranoside, a branch chain trisaccharide moiety found in all N-linked carbohydrates which possesses approximately 60-fold higher affinity than methyl α-D-mannopyranoside, exhibited a change in enthalpy of binding (ΔH) of -14.4 kcal mol-1 as compared to -8.2 kcal mol-1 for the monosaccharide. This demonstrates that Con A possesses an extended binding site for the trimannoside. However, a biantennary complex type carbohydrate with terminal β(1,2)-GlcNAc residues which binds with 3-fold higher affinity than the trimannoside possesses a ΔH of only -10.6 kcal mol-1. A plot of -ΔH versus -TΔS for the carbohydrates in the present study showed positive deviations in -TΔS for the complex type carbohydrate, as well as α(1,2)-di- and trimannosyl oligosaccharides which are part of the structures of oligomannose type carbohydrates. The relative favorable changes in binding entropies of these compounds are attributed to the presence of multiple internal and terminal residues in each molecule which can independently bind to the monosaccharide binding site of the lectin. The ΔH values for the complex type carbohydrate and the α(1,2) mannose oligosaccharides were also approximately -2.5 kcal mol-1 greater than that of methyl α-D-mannopyranoside, indicating some extended binding site interactions. The thermodynamics of binding of N-linked oligomannose type carbohydrates to dimeric Con A and its succinyl and acetyl derivatives were determined since these carbohydrates are bivalent and precipitate with the native tetrameric lectin but not with the dimeric protein and its two derivatives. Titration of succinyl-Con A with a Man5 oligomannose type oligosaccharide gave a ΔH of -14.5 kcal mol-1, which is similar to that of the branch chain trimannoside. This indicates that the α(1,6) core arm of Man5 which contains the trimannosyl moiety is the primary binding epitope for Con A. A fraction of quail ovalbumin containing a mixture of Man7 and Man8 chains at a single glycosylation site showed univalent binding to succinyl-Con A and a ΔH of -13.6 kcal mol-1. These results indicate that the trimannoside moiety on the α(1,6) arm(s) of the carbohydrate chains is the primary binding epitope and that its interactions with the lectin are relatively unaffected by the protein matrix of ovalbumin.

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