title: Three-dimensional structure of murine anti-p-azophenylarsonate Fab 36-71. 1. X-ray crystallography, site-directed mutagenesis, and modeling of the complex with hapten.
abstract: The structure of the antigen-binding fragment (Fab) of an anti-p-azophenylarsonate monoclonal antibody, 36-71, bearing a major cross-reactive idiotype of A/J mice has been refined to an R factor of 24.8% at a resolution of 1.85 A. The previously solved partial structure of this Fab at a resolution of 2.9 A (Rose et al., 1990) was used as an initial model for refinement against the high-resolution data. The complex with hapten has been modeled by docking the small-molecule crystal structure of phenylarsonic acid into the structure of the native Fab on the basis of a low-resolution electron density map of the complex. In this model, residue Arg-96 in the light chain and residues Asn-35, Trp-47, and Ser-99 in the heavy chain contact the arsonate moiety of the hapten; an additional bond is found between the arsonate group and a tightly bound water molecule. The phenyl moiety of the hapten packs against two tyrosine side chains at positions 50 and 106 in the heavy chain. Residue Arg-96 in the light chain had been implicated as involved in hapten binding on the basis of previous experiments, and indeed, this residue appears to play a crucial role in this model. Experiments employing site-directed mutagenesis directly support this conclusion. The heavy-chain complementarity-determining regions have novel conformations not previously observed in immunoglobulins except for the recently solved anti-p-azophenylarsonate Fab R 19.9 (Lascombe et al., 1989).
authors.affiliation: California Institute of Technology, Pasadena 91125.
authors.forename: R K
authors.initials: RK
authors.lastname: Strong
authors.forename: R
authors.initials: R
authors.lastname: Campbell
authors.forename: D R
authors.initials: DR
authors.lastname: Rose
authors.forename: G A
authors.initials: GA
authors.lastname: Petsko
authors.forename: J
authors.initials: J
authors.lastname: Sharon
authors.forename: M N
authors.initials: MN
authors.lastname: Margolies
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D001483:Base Sequence; D006241:Haptens / Q000737:chemistry* / Q000235:genetics; D007127:Immunoglobulin Constant Regions; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D013237:Stereoisomerism; D014961:X-Ray Diffraction; D010132:p-Azobenzenearsonate / Q000737:chemistry* / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D006241:Haptens; D007127:Immunoglobulin Constant Regions; D007140:Immunoglobulin Fab Fragments; D007135:Immunoglobulin Variable Region; D010132:p-Azobenzenearsonate