Peptide microarrays to interrogate the "histone code".
Peptide microarrays to interrogate the "histone code".
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DOI：
10.1016/b978-0-12-391940-3.00006-8
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发表时间：
2012
影响因子：
--
通讯作者：
Fuchs, Stephen M.
中科院分区：
文献类型：
--
作者：
Rothbart, Scott B.;Krajewski, Krzysztof;Strahl, Brian D.;Fuchs, Stephen M.
组蛋白翻译后修饰在调控染色质的动力学和功能中起着关键作用。在越来越多的文献的支持下，组蛋白PTM如甲基化和乙酰化在“组蛋白密码”的背景下一起起作用，其由效应蛋白读取或解释，然后驱动染色质中的功能输出（例如，基因转录）。越来越多的与组蛋白和/或其PTM相互作用的结构域已被鉴定。虽然我们对这些结构域如何与组蛋白相互作用的理解已经取得了重大进展，但大量假定的组蛋白结合基序尚未被表征，毫无疑问，新的结构域将继续被发现。在这一章中，我们提供了一个详细的组合修饰的组蛋白肽，使用这些肽的微阵列制造的建设方法，和方法来表征效应蛋白，抗体和组蛋白修饰酶的底物特异性的相互作用。我们讨论这些方法的背景下，其他可用的技术，并提供一个用户友好的方法，使探索组蛋白-蛋白质-酶的相互作用和功能。
Histone posttranslational modifications (PTMs) play a pivotal role in regulating the dynamics and function of chromatin. Supported by an increasing body of literature, histone PTMs such as methylation and acetylation function together in the context of a “histone code,” which is read, or interpreted, by effector proteins that then drive a functional output in chromatin (e.g., gene transcription). A growing number of domains that interact with histones and/or their PTMs have been identified. While significant advances have been made in our understanding of how these domains interact with histones, a wide number of putative histone-binding motifs have yet to be characterized, and undoubtedly, novel domains will continue to be discovered. In this chapter, we provide a detailed method for the construction of combinatorially modified histone peptides, microarray fabrication using these peptides, and methods to characterize the interaction of effector proteins, antibodies, and the substrate specificity of histone-modifying enzymes. We discuss these methods in the context of other available technologies and provide a user-friendly approach to enable the exploration of histone–protein–enzyme interactions and function.