Helm R, Cockrell G, Herman E, Burks A, Sampson H, Bannon G
Department of Pediatrics, University of Arkansas for Medical Sciences, Arkansas Children's Hospital Research Institute, Little Rock, Ark., USA.
Int Arch Allergy Immunol. 1998 Sep;117(1):29-37. doi: 10.1159/000023987.
Soybean proteins share a large number of cross-reacting allergens with other members of the legume family; however, soy-allergic patients rarely react clinically to other members of the legume family. Gly m Bd 30K, an IgE-binding protein with a molecular weight of 30 kD, was identified in soybean extracts by Western IgE-immunoblot analysis. This monomeric allergen was shown to have an N-terminal amino acid sequence and amino acid composition identical to that of the seed 34-kD protein, P34, a thiol protease of the papain family. Electron-microscopic immunolocalization of P34 monoclonal antibodies and IgE binding to sections of soybean seeds showed dense staining throughout the vacuolar bodies, localizing the allergens in protein storage vacuoles of seed cotyledons. We used pooled serum from soybean-sensitive patients to determine the linear IgE-specific epitopes in the 34-kD allergen amino acid sequence. B-cell epitope mapping revealed 10 regions of IgE-binding activity using an overlapping peptide strategy of 15-mers offset by 8 amino acids throughout the P34 sequence. Smaller overlapping peptides, 10-mers offset by 2 amino acids, revealed 16 distinct linear epitopes, 9 of which were mapped to the mature protein. No obvious amino acid sequence motifs could be identified by the smaller IgE-binding epitopes. Using individual patient serum, 5 immunodominant epitopes were identified in this allergen.
大豆蛋白与豆科植物家族的其他成员有大量交叉反应性过敏原；然而，大豆过敏患者临床上很少对豆科植物家族的其他成员产生反应。通过Western IgE免疫印迹分析在大豆提取物中鉴定出一种分子量为30 kD的IgE结合蛋白Gly m Bd 30K。这种单体过敏原的N端氨基酸序列和氨基酸组成与种子34-kD蛋白P34相同，P34是木瓜蛋白酶家族的一种巯基蛋白酶。P34单克隆抗体和IgE与大豆种子切片结合的电子显微镜免疫定位显示，整个液泡体都有密集染色，将过敏原定位在种子子叶的蛋白质储存液泡中。我们使用大豆敏感患者的混合血清来确定34-kD过敏原氨基酸序列中的线性IgE特异性表位。B细胞表位图谱分析使用在整个P34序列中以8个氨基酸偏移的15肽重叠肽策略，揭示了10个IgE结合活性区域。以2个氨基酸偏移的10肽更小重叠肽揭示了16个不同的线性表位，其中9个定位到成熟蛋白。较小的IgE结合表位未发现明显的氨基酸序列基序。使用个体患者血清，在这种过敏原中鉴定出5个免疫显性表位。
