Excision of thymine and 5-hydroxymethyluracil by the MBD4 DNA glycosylase domain: structural basis and implications for active DNA demethylation.
Excision of thymine and 5-hydroxymethyluracil by the MBD4 DNA glycosylase domain: structural basis and implications for active DNA demethylation.
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DOI：
10.1093/nar/gks628
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发表时间：
2012-09-01
影响因子：
14.9
通讯作者：
Cheng X
中科院分区：
文献类型：
--
作者：
Hashimoto H;Zhang X;Cheng X
哺乳动物DNA糖基酶-甲基-CpG结合域蛋白4(MBD4)-通过碱基切除修复途径参与活性DNA去甲基化。MBD4含有一个N-端的MBD和一个C-端的DNA糖基酶结构域。MBD4可以切除与鸟嘌呤(G：X)配对的错配碱基，其中X是尿嘧啶、胸腺嘧啶或5-羟甲基尿嘧啶(5hmU)。它们分别是胞嘧啶、5-甲基胞嘧啶(5mC)和5-羟甲基胞嘧啶(5hmC)的脱氨产物。在这里，我们提出了三种结构的MBD4 C末端糖基酶结构域(野生型及其催化突变体D534N)，在含有G：T或G：5hmU错配的DNA的复合体中。MBD4从双链DNA中翻转目标核苷酸。催化突变体D534N在活性部位结合口袋中捕获完整的靶核苷酸。MBD4通过O2、N3和O4原子特异性识别胸腺嘧啶或5hmU的Watson-Crick极边，从而将其活性限制在基于胸腺嘧啶/尿嘧啶的修饰，同时排除胞嘧啶及其衍生物。野生型酶裂解N-糖苷键，使核糖环处于翻转状态，同时裂解的碱基被释放。出乎意料的是，糖的C1‘还没有被水解，似乎通过静电或共价作用与D534的一个侧链羧基氧原子形成了一个稳定的中间体，这表明了与其他DNA糖基酶不同的催化机制。
The mammalian DNA glycosylase—methyl-CpG binding domain protein 4 (MBD4)—is involved in active DNA demethylation via the base excision repair pathway. MBD4 contains an N-terminal MBD and a C-terminal DNA glycosylase domain. MBD4 can excise the mismatched base paired with a guanine (G:X), where X is uracil, thymine or 5-hydroxymethyluracil (5hmU). These are, respectively, the deamination products of cytosine, 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC). Here, we present three structures of the MBD4 C-terminal glycosylase domain (wild-type and its catalytic mutant D534N), in complex with DNA containing a G:T or G:5hmU mismatch. MBD4 flips the target nucleotide from the double-stranded DNA. The catalytic mutant D534N captures the intact target nucleotide in the active site binding pocket. MBD4 specifically recognizes the Watson–Crick polar edge of thymine or 5hmU via the O2, N3 and O4 atoms, thus restricting its activity to thymine/uracil-based modifications while excluding cytosine and its derivatives. The wild-type enzyme cleaves the N-glycosidic bond, leaving the ribose ring in the flipped state, while the cleaved base is released. Unexpectedly, the C1′ of the sugar has yet to be hydrolyzed and appears to form a stable intermediate with one of the side chain carboxyl oxygen atoms of D534, via either electrostatic or covalent interaction, suggesting a different catalytic mechanism from those of other DNA glycosylases.
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影响因子：
64.5
作者：
Bhutani N;Burns DM;Blau HM
通讯作者：
Blau HM
影响因子：
3.7
作者：
Globisch D;Münzel M;Müller M;Michalakis S;Wagner M;Koch S;Brückl T;Biel M;Carell T
通讯作者：
Carell T
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
DOI：
10.1107/s0907444909052925
发表时间：
2010-02
期刊：
Acta crystallographica. Section D, Biological crystallography
影响因子：
--
作者：
Adams PD;Afonine PV;Bunkóczi G;Chen VB;Davis IW;Echols N;Headd JJ;Hung LW;Kapral GJ;Grosse-Kunstleve RW;McCoy AJ;Moriarty NW;Oeffner R;Read RJ;Richardson DC;Richardson JS;Terwilliger TC;Zwart PH
通讯作者：
Zwart PH
影响因子：
64.5
作者：
Guo JU;Su Y;Zhong C;Ming GL;Song H
通讯作者：
Song H