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    • 80. 发明公开
    • 단백질의 가용성2가 및 다가 헤테로 이량체 유사체
    • 蛋白质的可溶性多元和多重杂交模拟物
    • KR1020000005060A
    • 2000-01-25
    • KR1019980707687
    • 1997-03-28
    • 더 존스 홉킨스 유니버시티
    • 쉬넥조나단피오헤린신
    • C12N15/62C07K19/00C12N5/10
    • C07K14/7051A61K38/00A61K39/00A61K2039/605A61K2039/6056C07K14/70539C07K2319/00C07K2319/30C12N2799/026
    • PURPOSE: Soluble divalent and multivalent heterodimeric analogs of proteins and preparation method thereof are provided which are related to immuno-regulation are useful for studying TCR/MHC interactions, lymphocyte tracking, identifying new antigens. CONSTITUTION: Divalent peptide/MHC complexes or T cell receptors (superdimers) are constructed, to increase the avidity of soluble analogs for their cognates to biologically relevant levels. Using a recombinant DNA strategy, DNA encoding either the MHC class II/peptide or TCR heterodimers are ligated to DNA coding for murine Ig heavy and light chains. These constructs are subsequently expressed in a baculovirus expression system. Enzyme-linked immunosorbant assays (ELISA) specific for the Ig and polymorphic determinants of either the TCR or MHC fraction of the molecule indicates that infected insect cells secreted approximately 1 ug/ml of soluble, conformationally intact chimeric superdimers. SDS-PAGE analysis of purified protein shows that expected molecular weight species. The results of flow cytometry demonstrates that the TCR and class II chimeras bound specifically with high avidity to cells bearing their cognate receptors.
    • 目的:提供蛋白质的可溶性二价和多价异二聚体类似物及其制备方法,其与免疫调节有关,可用于研究TCR / MHC相互作用,淋巴细胞跟踪,鉴定新抗原。 构成:构建了二价肽/ MHC复合物或T细胞受体(superdimers),以增加其可溶性类似物对于相关生物学相关水平的亲合力。 使用重组DNA策略,将编码MHC II类/肽或TCR异源二聚体的DNA连接到编码鼠Ig重链和轻链的DNA。 这些构建体随后在杆状病毒表达系统中表达。 特异于分子的TCR或MHC级分的Ig和多态性决定簇的酶联免疫吸附测定(ELISA)表明感染的昆虫细胞分泌约1μg/ ml的可溶性,构象完整的嵌合超聚体。 纯化蛋白质的SDS-PAGE分析显示预期的分子量物种。 流式细胞术的结果表明,TCR和II类嵌合体具有高亲和力特异性结合携带其同源受体的细胞。