Wellcome Centre for Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh, UK.
Epigenetics Programme, Babraham Institute, Cambridge, UK.
Methods Mol Biol. 2022;2529:43-61. doi: 10.1007/978-1-0716-2481-4_2.
Histone methyltransferases (HMTs) catalyze the methylation of lysine and arginine residues in histone as well as nonhistone substrates. In vitro histone methyltransferase assays have been instrumental in identifying HMTs, and they continue to be invaluable tools for the study of these important enzymes, revealing novel substrates and modes of regulation.Here we describe a universal protocol to examine HMT activity in vitro that can be adapted to a range of HMTs, substrates, and experimental objectives. We provide protocols for the detection of activity based on incorporation of H-labeled methyl groups from S-adenosylmethionine (SAM), methylation-specific antibodies, and quantification of the reaction product S-adenosylhomocysteine (SAH).
组蛋白甲基转移酶（HMTs）催化赖氨酸和精氨酸残基在组蛋白以及非组蛋白底物上的甲基化。体外组蛋白甲基转移酶测定法在鉴定 HMTs 方面发挥了重要作用，并且仍然是这些重要酶的研究的宝贵工具，揭示了新的底物和调节方式。在这里，我们描述了一种通用的体外 HMT 活性检测方案，该方案可以适应多种 HMT、底物和实验目标。我们提供了基于 S-腺苷甲硫氨酸（SAM）中 H 标记的甲基掺入、甲基化特异性抗体以及反应产物 S-腺苷同型半胱氨酸（SAH）定量检测活性的方案。
