PURIFICATION AND N-TERMINAL AMINO-ACID-SEQUENCE OF 2 BIRCH POLLEN ISOALLERGENS (BET-VI AND BET-VII)

被引:11
作者
ELSAYED, S [1 ]
VIK, H [1 ]
机构
[1] UNIV BERGEN HOSP,CLIN BIOCHEM LAB,ALLERGY RES GRP,BERGEN,NORWAY
来源
INTERNATIONAL ARCHIVES OF ALLERGY AND APPLIED IMMUNOLOGY | 1990年 / 93卷 / 04期
关键词
D O I
10.1159/000235269
中图分类号
R392 [医学免疫学];
学科分类号
100102 ;
摘要
The major allergen of birch pollen BV45 (Bet v I) was previously isolated by molecular weight exclusion chromatography and eluted in the molecular weight region of 15-29 KD. Further purification of this fraction on an SP-Trisacryl M cation exchange matrix allowed 6 peaks of which the 4th (BV4A4) and 6th (BV4A6) included two dominant IgE-binding birch pollen isoallergens designated Bet v I and Bet v II. Final purification, using the 'Applied Biosystems' Peptide Micro Separation System, revealed two sharp peaks with a high degree of homogeneity. This was ascertained by automatic N-terminal amino acid (AA) sequence analyses which showed high average repetitive yields of the phenyl-thiohydantoin (PTH) AAs of the isoallergens sequenced. N-terminal AA analyses of the two fractions allowed 51 cleavages with correct identifications of PTH AAs for 3 replicates. The sequence data of the two isoallergens showed large homologies with the hazel pollen allergen, Cor a I, the birch pollen allergen, Ag 23, and the translated cDNA sequence derived from cloning birch pollen allergen genes. The sequence homologies support that Betula verrucosa allergens were derived from a gene family expressing several isologous allergens, 2 of which with 13 variable residues in a segment of 51 AAs. The antigenicity of the two fractions, Bet v I and Bet v II, was demonstrated by fused rocket immunoelectrophoresis (FRIE) and by crossed immunoelectrophoresis (CIE) giving single symmetrical antigenic precipitation. The specific IgE-binding capacity of the two isoallergens was supported by: (a) immunoautoradiography of the plates provided from FRIE; (b) specific IgE-binding inhibition in radioallergosorbent test (RAST) for fraction BV4A, and (c) similarly high RAST inhibition for the individual isoallergens. The in vivo allergenic activity was clearly shown by skin prick and nasal provocation tests. In conclusion, two isoallergens from birch pollen with proximate pI values (5.2 and 5.5) were isolated for the first time and the N-terminal sequence characterized.
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页码:378 / 384
页数:7
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