Isolation of Native Soluble and Membrane-Bound Protein Complexes from Yeast Saccharomyces cerevisiae
Isolation of Native Soluble and Membrane-Bound Protein Complexes from Yeast Saccharomyces cerevisiae
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DOI：
10.1007/978-1-4939-6937-1_4
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发表时间：
2017-01-01
期刊：
影响因子：
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通讯作者：
Erdmann, Ralf
中科院分区：
文献类型：
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作者：
Hansen, Tobias;Chan, Anna;Erdmann, Ralf
免疫沉淀是从复杂的样品混合物中分离单一蛋白质或天然蛋白质复合物的传统方法。最初的方法利用针对内源性蛋白质或表位标签的特异性抗体，这些抗体首先与靶蛋白结合，然后用蛋白A珠分离。该方法的一个进步是融合到靶蛋白的蛋白A标签的应用和用化学交联到琼脂糖基质的人免疫球蛋白G亲和纯化标记的蛋白。该方法将通过从酵母酿酒酵母（Saccharomyces cerevisiae）中纯化过氧化物酶体膜的蛋白质复合物来举例说明。
Immunoprecipitation is a traditional approach to isolate single proteins or native protein complexes from a complex sample mixture. The original method makes use of specific antibodies against endogenous proteins or epitope tags, which are first bound to the target protein and then isolated with protein A beads. An advancement of this method is the application of a protein A tag fused to the target protein and the affinity-purification of the tagged protein with human Immunoglobulin G chemically cross-linked to a sepharose matrix. This method will be described exemplified by the purification of protein complexes of the peroxisomal membrane from yeast Saccharomyces cerevisiae.