Mechanism of stimulation of catalytic activity of Dnmt3A and Dnmt3B DNA-(cytosine-C5)-methyltransferases by Dnmt3L
Mechanism of stimulation of catalytic activity of Dnmt3A and Dnmt3B DNA-(cytosine-C5)-methyltransferases by Dnmt3L
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DOI：
10.1074/jbc.m413412200
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发表时间：
2005-04-08
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;Liebert, K;Jeltsch, A
Dnmt3L 已被鉴定为 DNA 从头甲基转移酶催化活性的刺激剂。它对于哺乳动物生殖细胞的发育至关重要。我们在此表明，Dnmt3L 通过其自身的 C 端结构域直接与 Dnmt3A 和 Dnmt3B 酶各自的催化结构域结合，从而刺激 Dnmt3A 和 Dnmt3B 酶的催化活性。 Dnmt3A和-3B的催化活性被刺激了15倍，并且Dnmt3L直接与DNA结合，但不与S-腺苷-L-甲硫氨酸(AdoMet)结合。 Dnmt3A 和 Dnmt3L 之间的复合物形成可将 Dnmt3A 的 DNA 结合加速 20 倍，并降低其与 DNA 的 Km。 Dnmt3L 与 Dnmt3A 的相互作用增加了辅酶 AdoMet 与 Dnmt3A 的结合，并降低了 Dnmt3A 对 AdoMet 的 Km。根据我们的数据，我们提出了一个模型，其中 Dnmt3A 与 Dnmt3L 的相互作用诱导 Dnmt3A 的构象变化，从而打开酶的活性位点并促进 DNA 和 AdoMet 的结合。我们证明 Dnmt3A 和 Dnmt3L 的相互作用是短暂的，DNA 与 Dnmt3A 结合后，Dnmt3L 从复合物中解离。 Dnmt3L 解离后，Dnmt3A 采用封闭构象，导致 DNA 释放速率减慢。因此，Dnmt3L 充当底物交换因子，加速 DNA 和 AdoMet 与 DNA 从头甲基转移酶的结合。
Dnmt3L has been identified as a stimulator of the catalytic activity of de novo DNA methyltransferases. It is essential in the development of germ cells in mammals. We show here that Dnmt3L stimulates the catalytic activity of the Dnmt3A and Dnmt3B enzymes by directly binding to their respective catalytic domains via its own C-terminal domain. The catalytic activity of Dnmt3A and -3B was stimulated similar to 15-fold, and Dnmt3L directly binds to DNA but not to S-adenosyl-L-methionine (AdoMet). Complex formation between Dnmt3A and Dnmt3L accelerates DNA binding by Dnmt3A 20-fold and lowers its Km for DNA. Interaction of Dnmt3L with Dnmt3A increases the binding of the coenzyme AdoMet to Dnmt3A, and it lowers the Km of Dnmt3A for AdoMet. On the basis of our data we propose a model in which the interaction of Dnmt3A with Dnmt3L induces a conformational change of Dnmt3A that opens the active site of the enzyme and promotes binding of DNA and the AdoMet. We demonstrate that the interaction of Dnmt3A and Dnmt3L is transient, and after DNA binding to Dnmt3A, Dnmt3L dissociates from the complex. Following dissociation of Dnmt3L, Dnmt3A adopts a closed conformation leading to slow rates of DNA release. Therefore, Dnmt3L acts as a substrate exchange factor that accelerates DNA and AdoMet binding to de novo DNA methyltransferases.