title: Comparative studies on tree pollen allergens. X. Further purification and N-terminal amino acid sequence analyses of the major allergen of birch pollen (Betula verrucosa).
abstract: The previously isolated major allergen of birch pollen (fraction BV45), Int. Archs Allergy appl. Immun. 68: 70-78 (1982), was further purified by recycling chromatography. The purified preparation was run on a high-performance liquid chromatography (HPLC) TSK-G-2000 gel filtration chromatography column and, finally, on paper high-volt electrophoresis. The protein recovered met the homogeneity criteria required for performing the N-terminal sequence analysis. The allergenic and antigenic reactivities of the HPLC-purified protein, designated BV45B, was examined. A single homogeneous precipitation line in crossed immunoelectrophoresis (CIE) was shown. Specific IgE-inhibition tests and immuno-autoradiographic prints indicated that this allergen could bind reaginic IgE specificially and with good affinity. The homogeneity of BV45B was examined by isoelectric focusing (IEF). Several minor bands of pI differences of less than 0.1 units were visible, demonstrating the existence of some molecular variants of this protein. The N-terminal sequence analysis of the molecule was performed, and the following four amino acids were tentatively shown by sequential cleavage: NH2-Ala-Gly-Ile-Val-. The demonstration of one dominant N-terminal 1-dimethyl-amino-5-naphthalene sulphonyl (DNS)-amino acid by polyamide thin-layer chromatography at each sequence step confirmed that the N-terminal residue of the protein was not blocked; the heterogeneity shown by the IEF system was merely due to the presence of several homologous polymorphic proteins with identical N-terminal amino acid, the adequacy of the purification repertoire used.
mesh_terms: D000042:Absorption; D000485:Allergens / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D001345:Autoradiography; D003429:Cross Reactions; D014644:Genetic Variation; D006023:Glycoproteins / Q000235:genetics / Q000276:immunology; D006801:Humans; D007122:Immunoelectrophoresis; D008970:Molecular Weight; D011058:Pollen / Q000276:immunology*; D011852:Radioallergosorbent Test; D012882:Skin Tests; D013057:Spectrum Analysis; D014197:Trees
chemical_list: D000485:Allergens; D006023:Glycoproteins