Epitope mapping of monoclonal antibodies to keratin 19 using keratin fragments, synthetic peptides and phage peptide libraries.
To generate tools for monitoring processing and folding in keratin intermediate filaments, a group of monoclonal antibodies reacting with the intermediate filament protein keratin 19 were studied using different approaches to define the structure and localization of their epitopes. The binding pattern to bacterially expressed human keratin 19 fragments allowed the definition of minimal amino acid sequences required for antibody binding. The screening of overlapping 15-residue peptides confirmed and further specified the epitope locations for a subset of the tested antibodies. In addition, the epitope of an antibody with apparent species-restricted specificity (LE64) was revealed by isolating and characterizing a full-length keratin 19 clone from a PtK2 cDNA library. Taken together with species cross-reactivity of individual antibodies and sequence information obtained by probing a phage display library, specific amino acid residues could be highlighted as likely to be involved in the antibody binding.
Böttger
V
V
Department of Anatomy and Physiology, University of Dundee, Scotland.
Stasiak
P C
PC
Harrison
D L
DL
Mellerick
D M
DM
Lane
E B
EB
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001435:Bacteriophages / Q000235:genetics; D001483:Base Sequence; D001666:Binding Sites, Antibody; D018604:Epitope Mapping; D000939:Epitopes / Q000276:immunology; D015723:Gene Library; D006801:Humans; D015151:Immunoblotting; D007633:Keratins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010446:Peptide Fragments; D010455:Peptides; D011993:Recombinant Fusion Proteins; D007633:Keratins