Mapping the binding of monoclonal antibodies to histone H5

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Abstract
The binding sites of 9 monoclonal antibodies along the polypeptide chain of [chicken] 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 3 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.

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