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STRUCTURAL BASES OF SPECIFICITY AND AFFINITY IN ANTIGEN-ANTIBODY REACTIONS

Objective

IMPROVED UNDERSTANDING OF THE MOLECULAR BASIS OF ANTIGEN-ANTIBODY INTERACTIONS LEADING TO BASIC INFORMATION ON SIGNAL GENERATION BY ANTIBODY BINDING AND SIGNAL TRANSDUCTION.

LONG TERM ECONOMIC BENEFITS IN HEALTH CARE AND INDUSTRIAL BIOTECHNOLOGY ARE TO BE EXPECTED.
Research was carried out to determine, by X-ray crystallography, the 3-dimensional structures of:
antigen antibody complexes for which hen eggwhite lysozyme (HEL) was used as the model antigen;
heterologous antigens (other avian lysozymes);
FabNQ10 (from a monoclonal antiphenyloxazolone antibody) and its complexes with specific haptens;
an idiotope antiidiotope complex.

Results obtained in the study of the 3-dimensional structures were:
the determination of those of FabNQ10 free and bound to the phenyloxazolone hapten;
the refinement of the atomic coordinates of the antigen antibody complex FabD1.3-HEL;
the determination and refinement of the structures of the heterologous antigens, turkey and Japanese quail lysozymes;
the determination of the structure of an idiotope antiidiotope complex.

Another major result obtained was that of reshaping human antibodies: the antilysozyme activity of the mouse monoclonal antibody D1.3 was grafted by recombinant deoxyribonucleic acid (DNA) techniques onto a human immunoglobulin, a technique which should facilitate the production of therapeutic human (chimaeric) antibodies. The variable domains of the heavy chains (VH) and light chains (VL) of D1.3 were cloned and expressed in Esherichia coli. It was found that the VH domain binds lysozyme with an affinity constant (Kd = 19 nM) which is only 1 order of magnitude lower than that of the parent antibody (Kd = 2 nM).
THIS IS A JOINT PROJECT TO STUDY THE MOLECULAR BASIS OF ANTIGEN-ANTIBODY INTERACTIONS.

THE SPECIFIC AIMS OF THE RESEARCH WORK TO BE CARRIED OUT AT PASTEUR, ARE :

1) THE DETERMINATION OF THE THREE-DIMENSIONAL STRUCTURE OF A FAB-LYSOZYME ANTIGEN-ANTIBODY COMPLEX, INCLUDING THE CRYSTALLOGRAPHIC REFINEMENT OF THE STRUCTURE;
2) CRYSTALLIZATION TRIALS OF ADDITIONAL FAB-LYSOZYME COMPLEXES FROM MURINE MONOCLONAL ANTI-LYSOZYME ANTIBODIES AND OTHER ANTIGEN-ANTIBODY COMPLEXES;
3) DETERMINATION OF THE THREE-DIMENSIONAL STRUCTURE OF MUTATED-ANTIGENS ANTIBODY COMPLEXES;
4) DETERMINATION OF THE THREE-DIMENSIONAL STRUCTURE OF FAB NQ10/12.5 DERIVED FROM A MURINE MONOCLONAL ANTI-PHENYL OXAZOLONE ANTIBODY.;
5) THE DETERMINATION OF THE THREE-DIMENSIONAL STRUCTURE OF THE COMPLEX ANTI-LYSOZYME FAB D1.3/ANTI-IDIOTYPIC FAB 225 WHICHRECOGNIZES THE IDIOTYPE OF D1.3;
6) DETERMINATION OF THE THREE-DIMENSIONAL STRUCTURE OF A COMPLEX BETWEEN A HETEROCLITIC ANTI-LYSOZYME FAB AND A HETEROLOGOUS ANTIGEN, PHEASANT LYSOZYME AND OF THE ANTIGEN-ANTIBODY INTERACTIONS OF THE COMPLEX.

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

INSTITUT PASTEUR
Address
Rue Du Docteur Roux 25
75724 Paris
France

Participants (3)

Laboratoire pour l'Utilisation du Rayonnement Électromagnetique
France
Address
Université De Paris Xi (Université Paris-sud)
91405 Orsay
MRC MOLECULAR BIOLOGY LABORATORY
United Kingdom
Address

Cambridge
UNIVERSITY OF LEEDS
United Kingdom
Address
Woodhouse Lane
LS2 9JT Leeds