title: Characterization of IgE-binding epitopes on Candida albicans enolase.
abstract: Candida albicans enolase is one of the important allergens in Candida allergy. We isolated and purified 46kDa C. albicans enolase (CAE) from C. albicans and characterized epitopes for IgE antibody by lectin-blotting and enzymatic digestion followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. Lectin blotting and deglycozilation indicated that this protein did not contain polysaccharide side chains. The purified CAE and recombinant fusion protein produced from CAE gene possessed common epitopes for IgE antibody. We estimated IgE binding epitopes on the basis of reported amino acid sequences from the analysis of cDNA encoding CAE. V8 protease digestion of CAE gave six polypeptide fragments (A-F). The N-termini of each fragment were confirmed by amino acid sequence and the C-termini were estimated by molecular weights of each fragment and the specific cutting site of V8 protease. Fragment C (25.0 kDa; F-171-I-399) reacted to 90% IgE antibodies examined, whereas fragments D (21.0 kDa; F-171-I-360), E (16.2 kDa; F-171-D-317) and F (13.0 kDa; A-47-E-170) showed no IgE binding. Our results suggest that epitopes for IgE antibodies exist near the C-terminal of the protein.
authors.affiliation: Department of Paediatrics, Nagoya University School of Medicine, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Ito
authors.forename: A
authors.initials: A
authors.lastname: Ishiguro
authors.forename: T
authors.initials: T
authors.lastname: Kanbe
authors.forename: K
authors.initials: K
authors.lastname: Tanaka
authors.forename: S
authors.initials: S
authors.lastname: Torii
mesh_terms: D000595:Amino Acid Sequence; D002176:Candida albicans / Q000201:enzymology* / Q000276:immunology; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D015151:Immunoblotting; D007073:Immunoglobulin E / Q000276:immunology*; D008969:Molecular Sequence Data; D010751:Phosphopyruvate Hydratase / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D011485:Protein Binding / Q000276:immunology; D012697:Serine Endopeptidases
chemical_list: D000939:Epitopes; D007073:Immunoglobulin E; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D010751:Phosphopyruvate Hydratase