Mapping the binding of monoclonal antibodies to histone H5.
The binding sites of nine monoclonal antibodies along the polypeptide chain of histone H5 were mapped. Immunoblotting experiments with peptides generated from H5 by trypsin digestion, N-bromosuccinimide cleavage, and cyanogen bromide cleavage revealed that all of the monoclonal antibodies reacted with the globular region of H5 which is encompassed by amino acid residues 22-98. Within this globular segment, the epitopes could be subdivided into three regions. Monoclonals 1G11, 2E5, and 2H5 bind to residues 28-31. The close proximity of the epitopes was verified by a competitive enzyme-linked immunosorbent assay and by their binding pattern to a tryptic digest of H5. Monoclonals 4C6, 6E12, and 2E12 bind to a region encompassed by amino acids 28-53 while monoclonals 4H7, 1C3, and 3H9 bind to a region encompassed by residues 53-98. Precise localization of the epitopes in the primary sequence of H5 will allow detailed studies on the mode of binding of H5 to core particles in chromatin.
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D003488:Cyanogen Bromide; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis*; D005260:Female; D006657:Histones / Q000032:analysis* / Q000276:immunology; D051379:Mice; D010434:Pepsin A; D010446:Peptide Fragments / Q000032:analysis; D014357:Trypsin
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000939:Epitopes; D006657:Histones; D010446:Peptide Fragments; D014357:Trypsin; D010434:Pepsin A; D003488:Cyanogen Bromide