Immunological and structural features of the protein core of human polymorphic epithelial mucin.
The protein core of high mol. wt polymorphic epithelial mucin (PEM--approximately 400 kDa glycoprotein) which is associated with breast carcinomas, consists of a repeating 20 amino acid peptide motif [Gendler et al. (1988) J. biol. Chem. 263, 12,820-12,823]. Monoclonal antibodies C595 (anti-urinary mucin) and NCRC-11 (anti-breast carcinoma cells), and other antibodies against human milk fat globule membranes, were found to recognize determinants present within this 20 amino acid peptide. A model of the peptide was developed based on hydropathicity and structure prediction calculations and these indicated that the repeated structure is dominated by a hydrophilic domain of seven amino acids, extending into two flanking beta turns. NMR analysis of the 20 amino acid peptide was undertaken to probe the secondary structure. Epitope mapping experiments involving solid phase synthesis of overlapping heptapeptides in the repeat unit identified the minimum structures for antibody binding as Arg-Pro-Ala-Pro and Arg-Pro-Ala for the C595 and NCRC-11 antibodies, respectively. These determinants were found within the predicted hydrophilic turn region domain of the peptide. The epitopes for six other PEM-reactive monoclonal antibodies were also determined to reside within the predicted hydrophilic turn domain. This evidence is in accord with the disposition of this region of the PEM peptide core being at the exterior of the glycoprotein where it would be accessible to antibody recognition and binding events.
Cancer Research Campaign Laboratories, University of Nottingham, University Park, U.K.
M R
MR
Price
F
F
Hudecz
C
C
O'Sullivan
R W
RW
Baldwin
P M
PM
Edwards
S J
SJ
Tendler
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001943:Breast Neoplasms / Q000276:immunology; D003432:Cross-Linking Reagents; D000939:Epitopes / Q000276:immunology; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008562:Membrane Glycoproteins / Q000276:immunology* / Q000652:urine; D008969:Molecular Sequence Data; D018396:Mucin-1; D009077:Mucins / Q000276:immunology* / Q000652:urine; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D011487:Protein Conformation; D012710:Serum Albumin, Bovine
D000911:Antibodies, Monoclonal; D003432:Cross-Linking Reagents; D000939:Epitopes; D008562:Membrane Glycoproteins; D018396:Mucin-1; D009077:Mucins; D010455:Peptides; D012710:Serum Albumin, Bovine