Dnmt3b Methylates DNA by a Noncooperative Mechanism, and Its Activity Is Unaffected by Manipulations at the Predicted Dimer Interface.
Dnmt3b Methylates DNA by a Noncooperative Mechanism, and Its Activity Is Unaffected by Manipulations at the Predicted Dimer Interface.
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DOI：
10.1021/acs.biochem.6b00964
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发表时间：
2018-07-24
期刊：
影响因子：
2.9
通讯作者：
Gowher H
中科院分区：
文献类型：
--
作者：
Norvil AB;Petell CJ;Alabdi L;Wu L;Rossie S;Gowher H
从头 DNA 甲基转移酶 Dnmt3a-C 和 Dnmt3b-C 的催化结构域高度同源。然而，它们独特的生化特性可能会导致这些酶的底物偏好或生物功能的差异。 Dnmt3a-C 通过二聚体界面的相互作用形成四聚体，这也促进 DNA 的多聚化和协同性。与持续酶的情况类似，协同作用允许 Dnmt3a-C 甲基化同一 DNA 分子上的多个位点；然而，尚不清楚 Dnmt3b-C 是否通过协作或持续机制甲基化 DNA。 DNMT3A 二聚体界面中的 R882H 突变在急性髓系白血病中非常普遍，并导致其活性大幅丧失，这一观察结果强调了四聚体结构和协作机制的重要性。在区分协同性和持续合成性的条件下，我们发现与 Dnmt3a-C 相比，Dnmt3b-C 的活性不是协同性的，并证实了 Dnmt3b-C 和全长 Dnmt3b 酶的持续合成性。尽管 R878H 突变（R882H 的小鼠同源物）导致 Dnmt3a-C 协同性丧失，但类似的 Dnmt3b-C R829H 变体的活性和持续合成能力与野生型酶相当。此外，减弱 Dnmt3a-C 二聚体界面相互作用的缓冲液酸化对 Dnmt3b-C 活性没有影响。综上所述，这些结果证明了 Dnmt3b 和 Dnmt3a 之间的重要机制差异，并表明二聚体界面处的相互作用可能在调节 Dnmt3b-C 活性方面发挥有限的作用。这些新见解对 Dnmt3a 和 Dnmt3b 的独特生物学作用具有潜在影响。
The catalytic domains of the de novo DNA methyltransferases Dnmt3a-C and Dnmt3b-C are highly homologous. However, their unique biochemical properties could potentially contribute to differences in the substrate preferences or biological functions of these enzymes. Dnmt3a-C forms tetramers through interactions at the dimer interface, which also promote multimerization on DNA and cooperativity. Similar to the case for processive enzymes, cooperativity allows Dnmt3a-C to methylate multiple sites on the same DNA molecule; however, it is unclear whether Dnmt3b-C methylates DNA by a cooperative or processive mechanism. The importance of the tetramer structure and cooperative mechanism is emphasized by the observation that the R882H mutation in the dimer interface of DNMT3A is highly prevalent in acute myeloid leukemia and leads to a substantial loss of its activity. Under conditions that distinguish between cooperativity and processivity, we show that in contrast to that of Dnmt3a-C, the activity of Dnmt3b-C is not cooperative and confirm the processivity of Dnmt3b-C and the full length Dnmt3b enzyme. Whereas the R878H mutation (mouse homologue of R882H) led to the loss of cooperativity of Dnmt3a-C, the activity and processivity of the analogous Dnmt3b-C R829H variant were comparable to those of the wild-type enzyme. Additionally, buffer acidification that attenuates the dimer interface interactions of Dnmt3a-C had no effect on Dnmt3b-C activity. Taken together, these results demonstrate an important mechanistic difference between Dnmt3b and Dnmt3a and suggest that interactions at the dimer interface may play a limited role in regulating Dnmt3b-C activity. These new insights have potential implications for the distinct biological roles of Dnmt3a and Dnmt3b.
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发表时间：
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期刊：
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影响因子：
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作者：
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