Heiss Kirsten, Junkes Christof, Guerreiro Nelson, Swamy Mahima, Camacho-Carvajal Margarita M, Schamel Wolfgang W A, Haidl Ian D, Wild Doris, Weltzien Hans Ulrich, Thierse Hermann-Josef
Max-Planck Institute for Immunobiology, Freiburg, Germany.
Proteomics. 2005 Sep;5(14):3614-22. doi: 10.1002/pmic.200401215.
Metal-protein interactions are vitally important in all living organisms. Metalloproteins, including structural proteins and metabolic enzymes, participate in energy transfer and redox reactions or act as metallochaperones in metal trafficking. Among metal-associated diseases, T cell mediated allergy to nickel (Ni) represents the most common form of human contact hypersensitivity. With the aim to elucidate disease-underlying mechanisms such as Ni-specific T cell activation, we initiated a proteomic approach to identify Ni-interacting proteins in human B cells. As antigen presenting cells, B cells are capable of presenting MHC-associated Ni-epitopes to T cells, a prerequisite for hapten-specific T cell activation. Using metal-affinity enrichment, 2-DE and MS, 22 Ni-interacting proteins were identified. In addition to known Ni-binding molecules such as tubulin, actin or cullin-2, we unexpectedly discovered that at least nine of these 22 proteins belong to stress-inducible heat shock proteins or chaperonins. Enrichment was particularly effective for the hetero-oligomeric TRiC/CCT complex, which is involved in MHC class I processing. Blue Native/SDS electrophoresis analysis revealed that Ni-NTA-beads specifically retained the complete protein machinery, including the associated chaperonin substrate tubulin. The apparent Ni-affinity of heat shock proteins suggests a new function of these molecules in human Ni allergy, by linking innate and adaptive immune responses.
金属 - 蛋白质相互作用在所有生物中都至关重要。金属蛋白,包括结构蛋白和代谢酶,参与能量转移和氧化还原反应,或在金属转运中充当金属伴侣。在与金属相关的疾病中,T细胞介导的对镍(Ni)的过敏是人类接触性超敏反应最常见的形式。为了阐明诸如镍特异性T细胞活化等疾病潜在机制,我们启动了一项蛋白质组学方法来鉴定人B细胞中与镍相互作用的蛋白质。作为抗原呈递细胞,B细胞能够将与MHC相关的镍表位呈递给T细胞,这是半抗原特异性T细胞活化的先决条件。使用金属亲和富集、二维电泳和质谱,鉴定出了22种与镍相互作用的蛋白质。除了已知的镍结合分子如微管蛋白、肌动蛋白或cullin - 2外,我们意外地发现这22种蛋白质中至少有9种属于应激诱导型热休克蛋白或伴侣蛋白。富集对参与MHC I类加工的异源寡聚体TRiC/CCT复合物特别有效。蓝色非变性/十二烷基硫酸钠电泳分析表明,镍 - 次氮基三乙酸珠粒特异性保留了完整的蛋白质机制,包括相关的伴侣蛋白底物微管蛋白。热休克蛋白明显的镍亲和力表明这些分子在人类镍过敏中具有新功能,通过连接先天免疫和适应性免疫反应。