Sodium fluoride

氟化钠

Company: Sigma-Aldrich

Catalog#: 201154

Detection and Quantification of Calcium Ions in the Endoplasmic Reticulum and Cytoplasm of Cultured Cells Using Fluorescent Reporter Proteins and ImageJ Software

Shunsuke Saito and Kazutoshi Mori

Published online: 2023-08-20

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MPM-2 Mediated Immunoprecipitation of Proteins Undergoing Proline-directed Phosphorylation

Author:

Roberta Antonelli and Paola Zacchi,

Date:

2016-12-05

[Abstract] Immunoprecipitation (IP) represents a widely utilized biochemical method to isolate a specific protein from a complex mixture taking advantage of an antibody that specifically recognizes that particular target molecule. This procedure is extremely versatile and can be applied to concentrate a specific protein, to identify interacting partners in complex with it or to detect post-translational modifications. The mitotic protein monoclonal 2 (MPM-2) is an antibody originally raised against extracts of synchronized mitotic HeLa cells to identify proteins selectively present in mitotic, and not in interphase-cells (Davis et al., 1983). MPM-2 recognizes phosphorylated serine or threonine residues followed by proline (pS/T-P), consensus epitopes generated by the concerted action of ... [摘要] 免疫沉淀（IP）代表广泛使用的生物化学方法，以利用特异性识别特定靶分子的抗体从复杂混合物中分离特异性蛋白质。该程序是非常通用的，可以应用于集中特定蛋白质，识别与其复合的相互作用的配偶体或检测翻译后修饰。有丝分裂蛋白单克隆2（MPM-2）是最初针对同步有丝分裂HeLa细胞的提取物产生的抗体，以鉴定选择性存在于有丝分裂中的蛋白质，而不是在间期细胞中（Davis等，1983）。 MPM-2识别磷酸化的丝氨酸或苏氨酸残基，随后是脯氨酸（pS / T-P），由脯氨酸指导的蛋白激酶和磷酸酶的共同作用产生的共有表位（Lu等，2002）。这些可逆磷酸化事件已经出现以通过促进目标上的构象变化来控制各种细胞过程，这不仅仅是由于磷酸化事件本身。这些基序一旦被磷酸化，就能够招募Pin1（肽基 - 脯氨酰异构酶NIMA相互作用蛋白1）（Lu等人，1996; Lu和Zhou，2007），这是一个促进肽键上的顺式/反式异构化反应的伴侣，在基础功能不同的构象之间切换底层（Lu，2004; Wulf等人，2005）。该方案描述了使用支架分子突触后密度蛋白-95（PSD-95）（Chen等人，2005），神经元Pin1靶（Antonelli等，2016）作为示例的基于MPM-2的免疫沉淀策略以说明详细的程序。
【背景】由MPM-2抗体识别的抗原的鉴定代表了发现经过磷酸化脯氨酰异构化调控机制的靶分子的有用起点。 ...

Detection of HBV C Protein Phosphorylation in the Cell

Author:

Jaesung Jung and Kyongmin Kim,

Date:

2015-08-05

[Abstract] Among the seven serines and one threonine in the carboxyl-terminus of HBV C protein, all but one (serine 183) appear in the context of RxxS/T consensus phosphoacceptor motifs and also overlap with other consensus motifs, such as S/TP, RS, SPRRR, RRRS/T, or RRxS/T, suggesting that various cellular kinases phosphorylate these residues. To determine whether threonine and/or serine (serines 157, 164, 170, 172, 178, and 180, and threonine 162, adw subtype) of HBV C protein are indeed phosphoacceptor residues in cells, Huh7 were transfected with a series of C-protein-expressing mutants, labeled with ³²P-orthophosphate for 14 h, and then lysed. The ³²Pi-labeled lysates were immunoprecipitated with anti-HBc antibody, and the ³²Pi-labeled immunoprecipitated C ... [摘要] 在HBV C蛋白的羧基末端中的七个丝氨酸和一个苏氨酸中，除了一个（丝氨酸183）外，所有其它丝氨酸出现在RxxS/T共有磷酸受体基序的上下文中，并且还与其他共有基序重叠，例如S/TP，RS ，SPRRR，RRRS/T或RRxS/T，表明各种细胞激酶磷酸化这些残基。为了确定HBV C蛋白的苏氨酸和/或丝氨酸（丝氨酸157,164,170,172,178和180以及苏氨酸162，adw亚型）是否确实是细胞中的磷酸受体残基，Huh7用一系列C- 蛋白表达突变体，用正磷酸32P标记14小时，然后裂解。用抗HBc抗体免疫沉淀 32 P标记的裂解物，并通过放射自显影检测32 P标记的免疫沉淀的C蛋白。

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Sodium fluoride

氟化钠

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