Structural and dynamic consequences of increasing repeats in a MUC1 peptide tumor antigen.
MUC1 mucin is a large transmembrane glycoprotein whose extracelluler domain is composed of repeating units of a 20 amino acid sequence. In the cancer associated state, this protein expression becomes upregulated and underglycosylated. Previous studies, which show an enhanced binding of a 5-repeat over a 1-repeat MUC1 peptide to a panel of anti-MUC1 antibodies, have led us to investigate the structural and dynamic consequences of increasing repeat number. Two MUC1 peptides were studied: a 16mer corresponding to slightly less than one full repeat of the MUC1 tandem repeat sequence (GVTSAPDTRPAPGSTA) and a 40mer corresponding to two full repeats of the MUC1 sequence (VTSAPDTRPAPGSTAPPAHG)2. Isotopically labeled versions of these MUC1 peptides were cloned, expressed, purified, and evaluated structurally and dynamically using 15N- and 13C-edited NMR approaches. The data show that MUC1 structure, dynamics, and antibody binding affinity are invariant with increasing repeat number. In light of these results, we conclude that the enhanced antibody affinity of the 5-repeat over the 1-repeat MUC1 peptide is due to multivalency effects, and not due to the development of higher order structure in the longer length peptides. The implications of these results are discussed within the context of a multiple repeat MUC1 breast cancer vaccine design.
Schuman
Jason T
JT
Department of Chemistry, University of Washington, Seattle, WA 98195, USA.
Grinstead
Jeffrey S
JS
Apostolopoulos
Vasso
V
Campbell
A Patricia
AP
D000595:Amino Acid Sequence; D000918:Antibody Specificity / Q000276:immunology; D001666:Binding Sites, Antibody / Q000276:immunology; D001667:Binding, Competitive / Q000276:immunology; D019496:Cancer Vaccines / Q000276:immunology; D006801:Humans; D016056:Immunodominant Epitopes / Q000737:chemistry / Q000276:immunology; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000737:chemistry* / Q000276:immunology*; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry* / Q000276:immunology*; D011487:Protein Conformation; D020449:Repetitive Sequences, Amino Acid
D019496:Cancer Vaccines; D016056:Immunodominant Epitopes; C116055:MUC1 tandem repeat peptide; D018396:Mucin-1; D010446:Peptide Fragments