title: In vitro mutagenesis of HLA-B27: single and multiple amino acid substitutions at consensus B27 sites identify distinct monoclonal antibody-defined epitopes.
abstract: The consensus HLA-B27 sequence includes a unique constellation of amino acid residues along the peptide-binding cleft. To investigate the potential role of this region in the antigenic structure of HLA-B27, a panel of transfected cell lines was produced expressing 24 mutant B27 molecules with single or multiple substitutions within this constellation of residues. The cells were analyzed by flow cytometry with a panel of four anti-B27 mAb: ME1, GSP5.3, GS145.2, and B27M2. Previous studies have suggested that position 67 exerts a conformational effect on the ME1, GSP5.3, and GS145.2 epitopes. This was further supported in these studies by the observation that additional substitutions at the flanking residues 63 and 70 could reverse the disruption of these mAb epitopes by large residues at 67. Substitutions at positions 69-71 disrupted the binding of ME1 and GSP5.3, apparently by a direct effect. Individual substitutions at either of the two positions bearing residues unique to B27, 70 and 97, had no significant influence on the binding of any of the four mAb. The region of amino acid positions 63-71 in HLA-B27 thus appears to participate in the formation of at least three distinct epitopes shared by B27 and B7, identified by ME1, GSP5.3, and GS145.2.
authors.affiliation: Harold C. Simmons Arthritis Research Center, University of Texas Southwestern Medical Center, Dallas 75235-8884.
authors.forename: F A
authors.initials: FA
authors.lastname: el-Zaatari
authors.forename: J D
authors.initials: JD
authors.lastname: Taurog
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000235:genetics*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D001216:Asparagine / Q000235:genetics; D016384:Consensus Sequence; D000939:Epitopes / Q000032:analysis / Q000235:genetics* / Q000276:immunology; D015796:HLA-B27 Antigen / Q000032:analysis / Q000235:genetics* / Q000276:immunology; D015793:HLA-B7 Antigen / Q000032:analysis / Q000235:genetics / Q000276:immunology; D051379:Mice; D016297:Mutagenesis, Site-Directed; D013912:Threonine / Q000235:genetics
chemical_list: D000596:Amino Acids; D000911:Antibodies, Monoclonal; D000939:Epitopes; D015796:HLA-B27 Antigen; D015793:HLA-B7 Antigen; D013912:Threonine; D001216:Asparagine