Identification of a peptide which binds to the carbohydrate-specific monoclonal antibody B3.
The monoclonal antibody (mAb) B3 recognizes an antigen found on the surface of many adenocarcinoma cells. While the structure of the cellular antigen is unknown, epitope mapping using neoglycoproteins with known carbohydrate moieties indicates that the mAb B3 reacts with the LewisY (LeY) antigen [Pastan et al., Cancer Res. 51 (1991) 3781-3787]. We have used mAb B3 to select for peptides that mimic the carbohydrate structure using libraries of filamentous phage displaying random peptides on their surface. Phage that were selected coded for the sequence APWLYGPA. The corresponding peptide was synthesized and tested for its ability to bind to mAb B3. The peptide was found to inhibit specifically the binding of 111In-labeled mAb B3 to A431 adenocarcinoma cells, as well as to inhibit killing of these cells by a B3 immunotoxin. In addition, the LeY carbohydrate, lactodifucotetraose, was able to compete with the phage displaying this peptide for binding to mAb B3. Alanine-scanning mutagenesis of the sequence coding for this peptide indicates that four residues, PWLY, were critical for binding to the mAb. The sequence is similar to other sequences known to mimic carbohydrate structures.
Hoess
R
R
DuPont-Merck Pharmaceutical Company, Wilmington, DE 19980-0328.
Brinkmann
U
U
Handel
T
T
Pastan
I
I
D000230:Adenocarcinoma / Q000378:metabolism; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D000951:Antigens, Neoplasm / Q000378:metabolism; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D003001:Cloning, Molecular / Q000379:methods; D003090:Coliphages / Q000235:genetics*; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics; D006020:Glycopeptides / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D007983:Lewis Blood Group Antigens / Q000378:metabolism; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D000939:Epitopes; D006020:Glycopeptides; D006023:Glycoproteins; D007983:Lewis Blood Group Antigens; D009842:Oligopeptides