Kim Heejae, Chen Wilfred
Chemical and Biomolecular Engineering Department, University of Delaware, 150 Academy St., Newark, DE 19716, United States.
Chemical and Biomolecular Engineering Department, University of Delaware, 150 Academy St., Newark, DE 19716, United States.
J Biotechnol. 2016 Sep 20;234:27-34. doi: 10.1016/j.jbiotec.2016.07.016. Epub 2016 Jul 25.
Protein purification using inverse phase transition of elastin-like polypeptide (ELP) domains is a useful alternative to chromatography. Genetic fusions of ELP domains to various proteins have the ability to reversibly transition between soluble monomers and micron-sized aggregates and this has been used to selectively purify many ELP fusions. Affinity domains can enhance this technology by using specific protein binding domains to enable ELP mediated affinity capture (EMAC) of proteins of interest (POI) that have been fused to corresponding affinity ligands. In this paper, we highlight the use of Src homology 3 (SH3) domains and corresponding peptide ligands in EMAC that have differential binding affinities towards SH3 for efficient capture and elution of proteins. Furthermore, differences between capture and elution of a monomeric and a multimeric protein were also studied.
利用类弹性蛋白多肽（ELP）结构域的逆相变进行蛋白质纯化是一种有用的色谱替代方法。ELP结构域与各种蛋白质的基因融合能够在可溶性单体和微米级聚集体之间可逆转变，这已被用于选择性纯化许多ELP融合蛋白。亲和结构域可以通过使用特定的蛋白质结合结构域来增强这项技术，从而实现对已融合相应亲和配体的目标蛋白（POI）进行ELP介导的亲和捕获（EMAC）。在本文中，我们重点介绍了在EMAC中使用Src同源3（SH3）结构域和相应的肽配体，它们对SH3具有不同的结合亲和力，可用于高效捕获和洗脱蛋白质。此外，还研究了单体蛋白和多聚体蛋白在捕获和洗脱方面的差异。
