Characterization of human AlkB homolog 1 produced in mammalian cells and demonstration of mitochondrial dysfunction in ALKBH1-deficient cells.
Characterization of human AlkB homolog 1 produced in mammalian cells and demonstration of mitochondrial dysfunction in ALKBH1-deficient cells.
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DOI：
10.1016/j.bbrc.2017.10.158
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发表时间：
2018-01-01
影响因子：
3.1
通讯作者：
Hausinger RP
中科院分区：
文献类型：
--
作者：
Müller TA;Struble SL;Meek K;Hausinger RP
Alkbh 1是大肠杆菌DNA修复酶AlkB的哺乳动物同源物，AlkB是一种Fe（II）和2-酮戊二酸依赖性双加氧酶，可从DNA碱基上去除烷基损伤。ALKBH 1与六种不同的酶活性相关，包括DNA或tRNA羟基化、DNA中脱碱基（AP）位点的切割以及组蛋白的去甲基化。报道的这种蛋白质的细胞作用反映了不同的酶活性，包括直接DNA修复，tRNA修饰和组蛋白修饰。我们证明，在哺乳动物细胞（ALKBH 1293）中产生的ALKBH 1是类似的蛋白质产生的细菌（ALKBH 1 Ec）方面的m6 A脱甲基酶和AP裂解酶的活动。此外，我们发现ALKBH 1293以类似于ALKBH 1 Ec的方式与裂解酶产物的5′产物形成共价加合物。在两种人细胞株中内源性蛋白的定位和亚细胞分级分离研究证实ALKBH 1主要存在于线粒体中。两种CRISPR/Cas9产生的ALKBH 1缺陷型HEK 293细胞株显示出mtDNA拷贝数增加和线粒体功能障碍，如生长测量和柠檬酸合酶活性测定所揭示的。
Alkbh1 is a mammalian homolog of the Escherichia coli DNA repair enzyme AlkB, an Fe(II) and 2-oxoglutarate dependent dioxygenase that removes alkyl lesions from DNA bases. The human homolog ALKBH1 has been associated with six different enzymatic activities including DNA or tRNA hydroxylation, cleavage at abasic (AP) sites in DNA, as well as demethylation of histones. The reported cellular roles of this protein reflect the diverse enzymatic activities and include direct DNA repair, tRNA modification, and histone modification. We demonstrate that ALKBH1 produced in mammalian cells (ALKBH1293) is similar to the protein produced in bacteria (ALKBH1Ec) with regard to its m6A demethylase and AP lyase activities. In addition, we find that ALKBH1293 forms a covalent adduct with the 5′ product of the lyase product in a manner analogous to ALKBH1Ec. Localization and subcellular fractionation studies with the endogenous protein in two human cell strains confirm that ALKBH1 is primarily in the mitochondria. Two strains of CRISPR/Cas9-created ALKBH1-deficient HEK293 cells showed increases in mtDNA copy number and mitochondrial dysfunction as revealed by growth measurements and citrate synthase activity assays.
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影响因子：
14.9
作者：
Kawarada L;Suzuki T;Ohira T;Hirata S;Miyauchi K;Suzuki T
通讯作者：
Suzuki T
DOI：
10.1042/bj20121908
发表时间：
2013-06-15
期刊：
The Biochemical journal
影响因子：
--
作者：
Müller TA;Andrzejak MM;Hausinger RP
通讯作者：
Hausinger RP
影响因子：
3.8
作者：
Müller TA;Meek K;Hausinger RP
通讯作者：
Hausinger RP
DOI：
10.15252/embj.201694885
发表时间：
2016-10-04
期刊：
The EMBO journal
影响因子：
--
作者：
Haag S;Sloan KE;Ranjan N;Warda AS;Kretschmer J;Blessing C;Hübner B;Seikowski J;Dennerlein S;Rehling P;Rodnina MV;Höbartner C;Bohnsack MT
通讯作者：
Bohnsack MT
影响因子：
2.5
作者：
Pan, Zishu;Sikandar, Shaheen;Lipkin, Steven M.
通讯作者：
Lipkin, Steven M.