Non-productive DNA damage binding by DNA glycosylase-like protein Mag2 from Schizosaccharomyces pombe.
Non-productive DNA damage binding by DNA glycosylase-like protein Mag2 from Schizosaccharomyces pombe.
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DOI：
10.1016/j.dnarep.2012.12.001
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发表时间：
2013-03-01
期刊：
影响因子：
3.8
通讯作者：
Eichman, Brandt F.
中科院分区：
文献类型：
--
作者：
Adhikary, Suraj;Cato, Marilyn C.;McGary, Kriston L.;Rokas, Antonis;Eichman, Brandt F.
裂殖酵母含有两个与酵母和细菌中的烷基嘌呤DNA糖基酶的螺旋-发夹-螺旋(HHH)超家族相关的类似蛋白Mag1和Mag2。对四个裂殖酵母和其他真菌物种相关蛋白质的系统发育分析表明，Mag1/Mag2重复序列是裂殖酵母属所特有的，很可能存在于其祖先中。Mag1从DNA中剔除N3-和N7-烷基鸟嘌呤以及1，N6-乙基腺嘌呤，而Mag2被报道没有检测到的烷基嘌呤碱基切除活性，尽管与Mag1有很高的序列和活性部位相似。为了理解这种差异，我们确定了与基本DNA结合的Mag2的晶体结构，并将其与我们之前确定的Mag1-DNA结构进行了比较。与Mag1相比，Mag2不能将基本部分翻转到活性部位，也不能将DNA链5‘稳定在病变上，表明它不能形成具有催化活性的蛋白质-DNA复合体。Mag1和Mag2与DNA双链相互作用的细微差异说明了Mag2如何在没有完全接触损伤的情况下在损伤部位失速。我们通过碱基切除的突变分析来检验我们的结构预测，发现一个单一的氨基酸至少在一定程度上与Mag2的S缺乏活性有关。用MAG1中相应的丝氨酸残基取代MAG2Asp56，使MAG2具有与MAG1相当的εA切割活性。这项工作为HHH糖基酶以催化产生DNA的方式与DNA作用的化学和物理决定因素提供了新的见解。
Schizosaccharomyces pombe contains two paralogous proteins, Mag1 and Mag2, related to the helix-hairpin-helix (HhH) superfamily of alkylpurine DNA glycosylases from yeast and bacteria. Phylogenetic analysis of related proteins from four Schizosaccharomyces and other fungal species shows that the Mag1/Mag2 duplication is unique to the genus Schizosaccharomyces and most likely occurred in its ancestor. Mag1 excises N3- and N7-alkylguanines and 1,N6-ethenoadenine from DNA, whereas Mag2 has been reported to have no detectible alkylpurine base excision activity despite high sequence and active site similarity to Mag1. To understand this discrepancy we determined the crystal structure of Mag2 bound to abasic DNA and compared it to our previously determined Mag1-DNA structure. In contrast to Mag1, Mag2 does not flip the abasic moiety into the active site or stabilize the DNA strand 5′ to the lesion, suggesting that it is incapable of forming a catalytically competent protein-DNA complex. Subtle differences in Mag1 and Mag2 interactions with the DNA duplex illustrate how Mag2 can stall at damage sites without fully engaging the lesion. We tested our structural predictions by mutational analysis of base excision and found a single amino acid responsible at least in part for Mag2’s lack of activity. Substitution of Mag2 Asp56, which caps the helix at the base of the DNA intercalation loop, with the corresponding serine residue in Mag1 endows Mag2 with εA excision activity comparable to Mag1. This work provides novel insight into the chemical and physical determinants by which the HhH glycosylases engage DNA in a catalytically productive manner.
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影响因子：
14.9
作者：
Lavery R;Moakher M;Maddocks JH;Petkeviciute D;Zakrzewska K
通讯作者：
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DOI：
10.1107/s0907444904019158
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DOI：
10.1016/j.bbapap.2012.10.005
发表时间：
2013-01
影响因子：
3.2
作者：
Brooks, Sonja C.;Adhikary, Suraj;Rubinson, Emily H.;Eichman, Brandt F.
通讯作者：
Eichman, Brandt F.