Pelletier Nadine, Champagne Nathalie, Lim Henry, Yang Xiang-Jiao
Molecular Oncology Group, Department of Medicine, McGill University Health Center, 687 Pine Avenue West, Montreal, Quebec, H3A 1A1, Canada.
Methods. 2003 Sep;31(1):24-32. doi: 10.1016/s1046-2023(03)00084-7.
Histone acetylation is one major mechanism by which chromatin structure and function are regulated. Besides histones, many nonhistone proteins are also acetylated in vivo. Aberrant acetylation has been linked to the development of various human diseases. Through acetylating histone and nonhistone proteins, histone acetyltransferases (HATs) play fundamental roles in regulating chromatin remodeling, transcription, and other nuclear processes. Known HATs belong to several groups, including the GCN5/PCAF, p300/CBP, and MYST families. ESA1, SAS3, MOF, TIP60, HBO1, MOZ, and MORF are the MYST family members with demonstrated HAT activity. The MOZ and MORF genes are rearranged by chromosome abnormalities associated with several types of leukemia, so these two HATs have been implicated in leukemogenesis. Compared with p300, CBP, and PCAF, much less is known about MOZ and MORF. To elucidate the function and regulation of these two interesting HATs, we have conducted their initial characterization. Here we describe the expression, purification, and activity analysis of MOZ and MORF. For comparison, we also include the procedure for expression and purification of PCAF. These methods are useful not only for functional characterization of MOZ, MORF, PCAF, and other HATs, but also for preparation of HAT proteins to screen compound libraries and obtain inhibitors with potential therapeutic value.
组蛋白乙酰化是调节染色质结构和功能的一种主要机制。除组蛋白外，许多非组蛋白在体内也会发生乙酰化。异常乙酰化与多种人类疾病的发生有关。通过乙酰化组蛋白和非组蛋白，组蛋白乙酰转移酶（HATs）在调节染色质重塑、转录及其他核过程中发挥着重要作用。已知的HATs可分为几个组，包括GCN5/PCAF、p300/CBP和MYST家族。ESA1、SAS3、MOF、TIP60、HBO1、MOZ和MORF是已证实具有HAT活性的MYST家族成员。MOZ和MORF基因因与几种类型白血病相关的染色体异常而发生重排，因此这两种HATs被认为与白血病发生有关。与p300、CBP和PCAF相比，人们对MOZ和MORF的了解要少得多。为阐明这两种有趣的HATs的功能和调控机制，我们对它们进行了初步表征。在此，我们描述了MOZ和MORF的表达、纯化及活性分析。为作比较，我们还纳入了PCAF的表达和纯化步骤。这些方法不仅对MOZ、MORF、PCAF及其他HATs的功能表征有用，而且对制备HAT蛋白以筛选化合物文库并获得具有潜在治疗价值的抑制剂也有用。
