Assays of protein palmitoylation
Assays of protein palmitoylation
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DOI：
10.1016/j.ymeth.2006.04.015
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发表时间：
2006-10-01
期刊：
影响因子：
4.8
通讯作者：
Green, William N.
中科院分区：
文献类型：
--
作者：
Drisdel, Renaldo C.;Alexander, John K.;Green, William N.
蛋白质棕榈酰化在许多蛋白质的结构和功能中起着重要的作用。与其他脂质修饰不同，蛋白质棕榈酰化是高度动态的，棕榈酰化和脱棕榈酰化的循环可以调节蛋白质的功能和定位。由于测定方法的局限性，棕榈酰化的动态性质难以解决。在这里，我们讨论了各种方法，可用于测量蛋白质棕榈酰化，并确定棕榈酰化的网站。我们描述了新的方法的基础上“脂肪酰基交换标记”，其中棕榈酸酯通过羟胺介导的切割的棕榈酰-硫酯键，然后交换与巯基特异性标记化合物。该技术是高度敏感的，并允许棕榈酰化的定量估计。与其他用于检测翻译后修饰的技术不同，我们开发的技术可以用各种放射性或非放射性探针标记所有修饰位点，并可用于检测组织样品中蛋白质的棕榈酰化。(c)2006年爱思唯尔公司All rights reserved.
Protein palmitoylation plays an important role in the structure and function of a wide array of proteins. Unlike other lipid modifications, protein palmitoylation is highly dynamic and cycles of palmitoylation and depalmitoylation can regulate protein function and localization. The dynamic nature of palmitoylation is poorly resolved because of limitations in assay methods. Here, we discuss various methods that can be used to measure protein palmitoylation and identify sites of palmitoylation. We describe new methodology based on "fatty acyl exchange labeling" in which palmitate is removed via hydroxylamine-mediated cleavage of the palmitoyl-thioester bond and then exchanged with a sulfhydryl-specific labeling compound. The techniques are highly sensitive and allow for quantitative estimates of palmitoylation. Unlike other techniques used to assay posttranslational modifications, the techniques we have developed can label all sites of modification with a variety of probes, radiolabeled or non-radioactive, and can be used to assay the palmitoylation of proteins from tissue samples. (c) 2006 Elsevier Inc. All rights reserved.