Continuous B-epitope maps of cytochrome P450cam (CYP101) obtained by peptide scanning: correlation to spatial structure.
Protein continuous B-epitopes can be revealed using short synthetic peptides that overlap a known protein sequence. Since the whole protein surface is considered to possess antigenic properties, a question that arises is whether a set of linear B-epitopes determined by peptide scanning correlates with a protein spatial structure. We have chosen cytochrome P450cam (CYP101) of Pseudomonas putida, with known 3D structure, as a template. Sera of two rabbits and antibody egg yolk preparations from three chickens were produced against the P450cam molecule. These polyclonals were analyzed separately in ELISA with 409 overlapping P450cam hexapeptides. The whole set of continuous antigenic sites of P450cam covered about 45% of the P450cam sequence. However, immunodominant sites (those revealed with more than 50% antibody preparations), the so-called "antigenic core," represent only 9% of the protein sequence. While the amount of water-accessible residues in the total antigenic map (42%) was close to that in the whole native P450cam molecule (39%), the amount of water-accessible residues in the antigenic core was significantly higher (64%). These results led to the conclusion that antigenic core epitopes can be associated to the molecular surface, whereas epitopes with low detection frequency may partly correspond to unfolded regions of the protein molecule.
Moshkovskii
Serguei A
SA
Institute of Biomedical Chemistry, 10, Pogodinskaya Street, Moscow, 119992, Russia.
Kolesanova
Ekatherina F
EF
Archakov
Alexander I
AI
D000818:Animals; D019475:Camphor 5-Monooxygenase / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D018604:Epitope Mapping; D018985:Epitopes, B-Lymphocyte / Q000276:immunology*; D016056:Immunodominant Epitopes / Q000235:genetics; D010449:Peptide Mapping; D017510:Protein Folding; D011817:Rabbits
D018985:Epitopes, B-Lymphocyte; D016056:Immunodominant Epitopes; D019475:Camphor 5-Monooxygenase