Molecular enzymology of the catalytic domains of the Dnmt3a and Dnmt3b DNA methyltransferases
Molecular enzymology of the catalytic domains of the Dnmt3a and Dnmt3b DNA methyltransferases
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DOI：
10.1074/jbc.m202148200
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发表时间：
2002-06-07
影响因子：
4.8
通讯作者：
Jeltsch, A
中科院分区：
文献类型：
--
作者：
Gowher, H;Jeltsch, A
哺乳动物DNA甲基转移酶Dnmt 1、Dnmt 3a和Dnmt 3b的C-末端结构域具有胞嘧啶-C5甲基转移酶特有的所有保守基序。而孤立的催化结构域的Dnmt 1是无活性的，我们在这里显示，C-末端结构域的Dnmt 3a和Dnmt 3b的催化活性。Dnmt 3a和Dnmt 3b都没有显示出对卫星2序列的显著偏好，尽管Dnmt 3b是这些区域在体内甲基化所必需的。然而，Dnmt 3a的催化结构域在分布反应中甲基化DNA，而Dnmt 3b是进行性的，其在体外加速大分子DNA的甲基化。这种性质可以使Dnmt 3b成为卫星2重复甲基化的优选酶，因为它们高度富含CG。我们还分析了在Dnmt 3b催化结构域中发现的ICF（免疫缺陷、着丝粒不稳定和面部异常）患者中的六种不同突变的催化活性。其中5个突变体的催化活性降低了10-50倍; 1个突变体在我们的测定中无活性（残留活性
The C-terminal domains of the mammalian DNA methyltransferases Dnmt1, Dnmt3a, and Dnmt3b harbor all the conserved motifs characteristic for cytosine-C5 methyltransferases. Whereas the isolated catalytic domain of Dnmt1 is inactive, we show here that the C-terminal domains of Dnmt3a and Dnmt3b are catalytically active. Neither Dnmt3a nor Dnmt3b shows a significant preference for the satellite 2 sequence, although Dnmt3b is required for methylation of these regions in vivo. However, the catalytic domain of Dnmt3a methylates DNA in a distributive reaction, whereas Dnmt3b is processive, which accelerates methylation of macromolecular DNA in vitro. This property could make Dnmt3b a preferred enzyme for methylation at satellite 2 repeats, since they are highly CG-rich. We have also analyzed the catalytic activities of six different mutations found in ICF (immunodeficiency, centromeric instability, and facial abnormalities) patients in the catalytic domain of Dnmt3b. Five of them display catalytic activities reduced by 10-50-fold; one mutant was inactive in our assay (residual activity