An immobilised peptide array identifies antibodies to a discontinuous epitope in the extracellular domain of the bovine growth hormone receptor.
Using an array of overlapping decapeptides representing the extracellular domain of the bovine (b) growth-hormone receptor (GHR) we have mapped the continuous, dominant epitopes defined by five rabbit and one guinea pig polyclonal antisera to recombinant bovine growth-hormone-binding protein (rbGHBP). We report that six major epitopes are identified by these antisera and that these largely occur in areas of non-ordered secondary structure, although there is some contribution from the extensive beta-sheet structure of GHBP. Similar to our previously described studies for growth hormone (GH), we have again found slight differences between animals in the exact location of these epitopes. Using peptide-affinity chromatography we have isolated a population of antibodies reactive with epitope 1 (the N-terminal epitope:GHBP residues 21-38). Analysis of these antibodies by further peptide affinity chromatography and competitive radioimmunoassay experiments indicated cross-reactivity of epitope-1-specific antibodies with epitope 4 (in the interdomain hinge region of the GHBP:residues 111-126). We suggest that, although separate in the primary structure of the molecule, the tertiary fold exhibited by GHBP may bring into close proximity areas of sequence representing epitope 1 and epitope 4 such that they represent a conformational epitope. Under these conditions our experiments indicate that peptides 1 and 4 may represent partial functional epitopes for this antibody population and consequently demonstrate that this approach may be useful in describing discontinuous epitopes.
Beattie
J
J
Hannah Research Institute, Ayr, Scotland.
Shand
J H
JH
Flint
D J
DJ
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D000918:Antibody Specificity; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D002417:Cattle; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis*; D005260:Female; D006168:Guinea Pigs; D007700:Kinetics; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011817:Rabbits; D011986:Receptors, Somatotropin / Q000737:chemistry* / Q000276:immunology* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism
D000906:Antibodies; D002352:Carrier Proteins; D000939:Epitopes; D010455:Peptides; D011986:Receptors, Somatotropin; D011994:Recombinant Proteins; C051815:somatotropin-binding protein