title: Functional mapping of conserved, surface-exposed charges of antibody variable domains.
abstract: Surface-exposed charges can affect protein structure, stability and solubility as well as the kinetics of both the folding process and interaction with binding partners. We have investigated the influence on kinetic interaction parameters of 14 conserved, surface-exposed charges located away from the paratope in the variable domains of two antibodies of different specificity. We found that conserved, surface-exposed, charged framework residues are asymmetrically distributed on opposite faces of both VH and VL domains. Some of the charges play a critical role in protein folding and stability. While electrostatic forces within or close to the binding interface can be used to optimize the association rate, we confirmed the predicted minor effects of charge modifications remote from the binding site. They had no effect on the dissociation rate parameter. Our study demonstrates the role of residues remote from the interaction site in the recognition function as well as the limited effect of surface charge modifications in antibody fragments on kinetic interaction parameters.
authors.affiliation: Institut de Biologie Structurale Jean-Pierre Ebel, Laboratoire d'Ingénierie des Macromolécules, CEA/CNRS/UJF, 41 rue Jules Horowitz,-F-38027 Grenoble cedex 1, France.
authors.forename: Marianne
authors.initials: M
authors.lastname: Weidenhaupt
authors.forename: Myriam Ben
authors.initials: MB
authors.lastname: Khalifa
authors.forename: Nicolas
authors.initials: N
authors.lastname: Hugo
authors.forename: Laurence
authors.initials: L
authors.lastname: Choulier
authors.forename: Danièle
authors.initials: D
authors.lastname: Altschuh
authors.forename: Thierry
authors.initials: T
authors.lastname: Vernet
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody; D017124:Conserved Sequence; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007147:Immunoglobulin Light Chains / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000276:immunology; D010449:Peptide Mapping; D017433:Protein Structure, Secondary; D014027:Tobacco Mosaic Virus / Q000276:immunology
chemical_list: D000914:Antibodies, Viral; D007140:Immunoglobulin Fab Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D010446:Peptide Fragments