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    • 5. 发明授权
    • Method for purification of a protein complex and identification of its components
    • 蛋白质复合物的纯化方法及其成分的鉴定
    • US07825227B2
    • 2010-11-02
    • US10984958
    • 2004-11-09
    • John J. BonifaceVladimir KeryJohn M. PeltierPaul B. RobbinsJustin SavageMoritz Von Rechenberg
    • John J. BonifaceVladimir KeryJohn M. PeltierPaul B. RobbinsJustin SavageMoritz Von Rechenberg
    • A23J1/00C12P21/04
    • C07K5/1021C07K1/22C07K5/101
    • The present invention provides a method for purifying a protein complex and the components comprising the complex from a cell, a cell or tissue lysate or a whole organism by employing a combined set of affinity tags of high affinity, specificity and ease of elution. The method involves using a protein or a peptide modified to contain one or more affinity tags separated by one or more specific protease cleavage sites to isolate any interacting proteins or fragments thereof. Specifically exemplified is a method employing a modified bait containing AviTag or GST and a removal tag of His6 hexapeptide positioned proximal to the bait such that any excess bait can be efficiently removed from the purified complex yielding enriched interacting proteins prior to subjecting the complex for further identification of individual components. The identification of the proteins or fragments thereof contained in the protein complex will provide new targets for the identification of new pharmaceuticals and diagnostic tools.
    • 本发明提供一种通过使用高亲和力,特异性和容易洗脱的组合的一组亲和标签来纯化蛋白质复合物和包含来自细胞,细胞或组织裂解物或整个生物体的复合物的组分的方法。 所述方法包括使用经修饰的蛋白质或肽含有由一个或多个特异性蛋白酶切割位点分离的一个或多个亲和标签,以分离任何相互作用的蛋白质或其片段。 具体举例说明了使用含有AviTag或GST的修饰诱饵和位于诱饵附近的His6六肽的去除标签的方法,使得任何过量的诱饵可以在将复合物进一步鉴定之前从纯化的复合物中有效地除去产生富集的相互作用蛋白质 的单个组件。 包含在蛋白质复合物中的蛋白质或其片段的鉴定将为鉴定新的药物和诊断工具提供新的目标。