Immunoaffinity purification of endogenous proteins from S. cerevisiae for post-translational modification and protein interaction analysis.
Immunoaffinity purification of endogenous proteins from S. cerevisiae for post-translational modification and protein interaction analysis.
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DOI：
10.1016/j.xpro.2021.100945
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发表时间：
2021-12-17
期刊：
影响因子：
--
通讯作者：
Green EM
中科院分区：
文献类型：
--
作者：
Jaiswal D;Turniansky R;Green EM
通过翻译后修饰和蛋白质-蛋白质相互作用的蛋白质调节对于控制分子途径至关重要。在这里，我们描述了一种免疫亲和纯化方法在酿酒酵母。该方案使用内源性表达的表位标记的蛋白质，并可应用于翻译后修饰或蛋白质结合伴侣的鉴定。这里使用赖氨酸甲基转移酶Set 5作为一个例子来纯化磷酸化Set 5并鉴定磷酸化位点;然而，这种方法可以应用于酵母中的一组不同的蛋白质。有关本方案使用和执行的完整详细信息，请参阅。使用FLAG标签制备用于蛋白质免疫亲和纯化的酵母免疫亲和纯化优化的详细建议用于鉴定翻译后修饰或蛋白质相互作用的通用方法通过翻译后修饰和蛋白质-蛋白质相互作用进行的蛋白质调节对于控制分子途径至关重要。在这里，我们描述了一种免疫亲和纯化方法在酿酒酵母。该方案使用内源性表达的表位标记的蛋白质，并可应用于翻译后修饰或蛋白质结合伴侣的鉴定。这里使用赖氨酸甲基转移酶Set 5作为一个例子来纯化磷酸化Set 5并鉴定磷酸化位点;然而，这种方法可以应用于酵母中的一组不同的蛋白质。
Protein regulation by post-translational modifications and protein-protein interactions is critical to controlling molecular pathways. Here, we describe an immunoaffinity purification approach in Saccharomyces cerevisiae. The protocol uses an endogenously-expressed epitope-tagged protein and can be applied to the identification of post-translational modifications or protein binding partners. The lysine methyltransferase Set5 is used as an example here to purify phosphorylated Set5 and identify phosphosites; however, this approach can be applied to a diverse set of proteins in yeast. For complete details on the use and execution of this protocol, please refer to. Preparation of yeast for immunoaffinity purification of proteins using the FLAG tag Detailed recommendations on optimization of immunoaffinity purifications Versatile method for identifying post-translational modifications or protein interactors Protein regulation by post-translational modifications and protein-protein interactions is critical to controlling molecular pathways. Here, we describe an immunoaffinity purification approach in Saccharomyces cerevisiae. The protocol uses an endogenously-expressed epitope-tagged protein and can be applied to the identification of post-translational modifications or protein binding partners. The lysine methyltransferase Set5 is used as an example here to purify phosphorylated Set5 and identify phosphosites; however, this approach can be applied to a diverse set of proteins in yeast.
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影响因子：
2.6
作者：
Moqtaderi, Zarrnk;Struhl, Kevin
通讯作者：
Struhl, Kevin
DOI：
10.1016/j.jbc.2021.100939
发表时间：
2021-08
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Separovich RJ;Wilkins MR
通讯作者：
Wilkins MR
DOI：
10.1074/jbc.ra120.015995
发表时间：
2021-01
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Separovich RJ;Wong MWM;Chapman TR;Slavich E;Hamey JJ;Wilkins MR
通讯作者：
Wilkins MR
影响因子：
3.3
作者：
Shang, C;Hazbun, TR;Barnes, G
通讯作者：
Barnes, G
影响因子：
5.3
作者：
Jaiswal, Deepika;Turniansky, Rashi;Green, Erin M.
通讯作者：
Green, Erin M.