Quantitative analysis of the interaction between immune complex and Clq complement subcomponent. The role of interdomain interactions in rabbit IgG in binding of Clq to immune precipitates

F. Vonderviszt, J. Török, S. Lakatos, F. Kilár, P. Závodszky

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Abstract

A novel method was developed for the analysis of the interaction of large multivalent ligands with surfaces (matrices) to analyse the binding of complement subcomponent Clq to immune precipitates. Our new evaluation method provides quantitative data characteristic of the Clq-immune-complex interaction and of the structure of the immune complex as well. To reveal the functional role of domain-domain interactions in the Fc part of IgG the binding of Clq to different anti-ovalbumin IgG-ovalbumin immune complexes was studied. Immune-complex precipitates composed of rabbit IgG in which the non-covalent bonds were abolished by splitting off the CH3 domains, i.e. by using Facb fragments, and the covalent contact was broken by reduction and alkylation of the single inter-heavy-chain disulphide bond. The quantitative analysis of the binding curves provides a dissociation constant (K) of 200 nM for the interaction between Clq and immune precipitate formed from native IgG. Surprisingly, for immune precipitates composed of Facb fragments or IgG in which the inter-heavy-chain disulphide bond had been selectively reduced and alkylated, stronger binding (K = 30 nM) was observed. In this case, however, changes in the structure of the immune-complex matrix were also detected. These structural changes may account for the strengthening of the Clq-immune-complex interaction, which can be strongly influenced by the flexibility and the binding-site pattern of the immune-complex precipitates. These results suggest that domain-domain interactions in the Fc part of IgG affect the segmental mobility of IgG molecules and the spatial arrangement of the immune-complex matrix rather than the affinity of individual Clq-binding sites on IgG.

Original languageEnglish
Pages (from-to)449-456
Number of pages8
JournalBiochemical Journal
Volume243
Issue number2
Publication statusPublished - 1987

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Precipitins
Antigen-Antibody Complex
Immunoglobulin G
Rabbits
Chemical analysis
Ovalbumin
Disulfides
Precipitates
Binding Sites
Alkylation
Ligands
Molecules

ASJC Scopus subject areas

  • Biochemistry

Cite this

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title = "Quantitative analysis of the interaction between immune complex and Clq complement subcomponent. The role of interdomain interactions in rabbit IgG in binding of Clq to immune precipitates",
abstract = "A novel method was developed for the analysis of the interaction of large multivalent ligands with surfaces (matrices) to analyse the binding of complement subcomponent Clq to immune precipitates. Our new evaluation method provides quantitative data characteristic of the Clq-immune-complex interaction and of the structure of the immune complex as well. To reveal the functional role of domain-domain interactions in the Fc part of IgG the binding of Clq to different anti-ovalbumin IgG-ovalbumin immune complexes was studied. Immune-complex precipitates composed of rabbit IgG in which the non-covalent bonds were abolished by splitting off the CH3 domains, i.e. by using Facb fragments, and the covalent contact was broken by reduction and alkylation of the single inter-heavy-chain disulphide bond. The quantitative analysis of the binding curves provides a dissociation constant (K) of 200 nM for the interaction between Clq and immune precipitate formed from native IgG. Surprisingly, for immune precipitates composed of Facb fragments or IgG in which the inter-heavy-chain disulphide bond had been selectively reduced and alkylated, stronger binding (K = 30 nM) was observed. In this case, however, changes in the structure of the immune-complex matrix were also detected. These structural changes may account for the strengthening of the Clq-immune-complex interaction, which can be strongly influenced by the flexibility and the binding-site pattern of the immune-complex precipitates. These results suggest that domain-domain interactions in the Fc part of IgG affect the segmental mobility of IgG molecules and the spatial arrangement of the immune-complex matrix rather than the affinity of individual Clq-binding sites on IgG.",
author = "F. Vonderviszt and J. T{\"o}r{\"o}k and S. Lakatos and F. Kil{\'a}r and P. Z{\'a}vodszky",
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T1 - Quantitative analysis of the interaction between immune complex and Clq complement subcomponent. The role of interdomain interactions in rabbit IgG in binding of Clq to immune precipitates

AU - Vonderviszt, F.

AU - Török, J.

AU - Lakatos, S.

AU - Kilár, F.

AU - Závodszky, P.

PY - 1987

Y1 - 1987

N2 - A novel method was developed for the analysis of the interaction of large multivalent ligands with surfaces (matrices) to analyse the binding of complement subcomponent Clq to immune precipitates. Our new evaluation method provides quantitative data characteristic of the Clq-immune-complex interaction and of the structure of the immune complex as well. To reveal the functional role of domain-domain interactions in the Fc part of IgG the binding of Clq to different anti-ovalbumin IgG-ovalbumin immune complexes was studied. Immune-complex precipitates composed of rabbit IgG in which the non-covalent bonds were abolished by splitting off the CH3 domains, i.e. by using Facb fragments, and the covalent contact was broken by reduction and alkylation of the single inter-heavy-chain disulphide bond. The quantitative analysis of the binding curves provides a dissociation constant (K) of 200 nM for the interaction between Clq and immune precipitate formed from native IgG. Surprisingly, for immune precipitates composed of Facb fragments or IgG in which the inter-heavy-chain disulphide bond had been selectively reduced and alkylated, stronger binding (K = 30 nM) was observed. In this case, however, changes in the structure of the immune-complex matrix were also detected. These structural changes may account for the strengthening of the Clq-immune-complex interaction, which can be strongly influenced by the flexibility and the binding-site pattern of the immune-complex precipitates. These results suggest that domain-domain interactions in the Fc part of IgG affect the segmental mobility of IgG molecules and the spatial arrangement of the immune-complex matrix rather than the affinity of individual Clq-binding sites on IgG.

AB - A novel method was developed for the analysis of the interaction of large multivalent ligands with surfaces (matrices) to analyse the binding of complement subcomponent Clq to immune precipitates. Our new evaluation method provides quantitative data characteristic of the Clq-immune-complex interaction and of the structure of the immune complex as well. To reveal the functional role of domain-domain interactions in the Fc part of IgG the binding of Clq to different anti-ovalbumin IgG-ovalbumin immune complexes was studied. Immune-complex precipitates composed of rabbit IgG in which the non-covalent bonds were abolished by splitting off the CH3 domains, i.e. by using Facb fragments, and the covalent contact was broken by reduction and alkylation of the single inter-heavy-chain disulphide bond. The quantitative analysis of the binding curves provides a dissociation constant (K) of 200 nM for the interaction between Clq and immune precipitate formed from native IgG. Surprisingly, for immune precipitates composed of Facb fragments or IgG in which the inter-heavy-chain disulphide bond had been selectively reduced and alkylated, stronger binding (K = 30 nM) was observed. In this case, however, changes in the structure of the immune-complex matrix were also detected. These structural changes may account for the strengthening of the Clq-immune-complex interaction, which can be strongly influenced by the flexibility and the binding-site pattern of the immune-complex precipitates. These results suggest that domain-domain interactions in the Fc part of IgG affect the segmental mobility of IgG molecules and the spatial arrangement of the immune-complex matrix rather than the affinity of individual Clq-binding sites on IgG.

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