Biochemical properties of naturally occurring human bloom helicase variants.
Biochemical properties of naturally occurring human bloom helicase variants.
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DOI：
10.1371/journal.pone.0281524
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发表时间：
2023
期刊：
影响因子：
3.7
通讯作者：
中科院分区：
文献类型：
--
作者：
Bloom综合征解旋酶(BLM)是一种RecQ家族解旋酶，参与多种细胞过程，包括DNA复制、DNA修复和端粒维持。人类BLM的突变会导致Bloom综合征(BS)，这是一种常染色体隐性遗传病，会导致无数负面的健康影响，包括癌症的易感性。引起BLm的BS突变通常会对BLm的ATPase和解旋酶活性产生负面影响。虽然引起BS的BLM突变在体外和体内都有很好的特征，但还有其他较少研究的BLM突变存在于人类群体中，不会导致BS。其中两个非BS突变，编码BLMP868L和BLMG1120R，当纯合时，会增加人类细胞中的姐妹染色单体交换。为了在体外鉴定这些自然产生的BLM突变蛋白，我们用P868L或G1120R替换纯化了BLM催化核心(BLMcore，残基636-1298)。我们还纯化了一个BLMcore K869A K870A突变蛋白，它改变了P868残基附近的一个富含赖氨酸的环。我们发现BLMcore P868L和G1120R蛋白都能降解ATP，结合不同的DNA底物，解开G-四链和双链DNA结构。分子动力学模拟表明，P868L取代减弱了DNA与BLM的翼状螺旋结构域的相互作用，并改变了ATPase结构域的一个叶的方向。由于BLMcore P868L和G1120R在体外保留了解旋酶功能，因此基因组不稳定性的增加可能是由于体内突变蛋白的特定影响所致。有趣的是，我们发现BLMcore K869A K870A与野生型BLMcore相比，ATPase活性降低，与双链DNA结构的结合减弱，解旋酶活性降低。因此，富含赖氨酸的环可能在BLM的ATPase活性、特异结合和DNA解离功能中起重要作用。
Bloom syndrome helicase (BLM) is a RecQ-family helicase implicated in a variety of cellular processes, including DNA replication, DNA repair, and telomere maintenance. Mutations in human BLM cause Bloom syndrome (BS), an autosomal recessive disorder that leads to myriad negative health impacts including a predisposition to cancer. BS-causing mutations in BLM often negatively impact BLM ATPase and helicase activity. While BLM mutations that cause BS have been well characterized both in vitro and in vivo, there are other less studied BLM mutations that exist in the human population that do not lead to BS. Two of these non-BS mutations, encoding BLM P868L and BLM G1120R, when homozygous, increase sister chromatid exchanges in human cells. To characterize these naturally occurring BLM mutant proteins in vitro, we purified the BLM catalytic core (BLMcore, residues 636–1298) with either the P868L or G1120R substitution. We also purified a BLMcore K869A K870A mutant protein, which alters a lysine-rich loop proximal to the P868 residue. We found that BLMcore P868L and G1120R proteins were both able to hydrolyze ATP, bind diverse DNA substrates, and unwind G-quadruplex and duplex DNA structures. Molecular dynamics simulations suggest that the P868L substitution weakens the DNA interaction with the winged-helix domain of BLM and alters the orientation of one lobe of the ATPase domain. Because BLMcore P868L and G1120R retain helicase function in vitro, it is likely that the increased genome instability is caused by specific impacts of the mutant proteins in vivo. Interestingly, we found that BLMcore K869A K870A has diminished ATPase activity, weakened binding to duplex DNA structures, and less robust helicase activity compared to wild-type BLMcore. Thus, the lysine-rich loop may have an important role in ATPase activity and specific binding and DNA unwinding functions in BLM.
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影响因子：
15
作者：
Dudev, T;Lim, C
通讯作者：
Lim, C
影响因子：
14.9
作者：
Zittel MC;Keck JL
通讯作者：
Keck JL
影响因子：
5.5
作者：
Leighty, Ralph E.;Varma, Sameer
通讯作者：
Varma, Sameer
影响因子：
5.5
作者：
Hess, Berk
通讯作者：
Hess, Berk
DOI：
10.1172/jci147301
发表时间：
2022-03-01
期刊：
The Journal of clinical investigation
影响因子：
--
作者：
Abu-Libdeh B;Jhujh SS;Dhar S;Sommers JA;Datta A;Longo GM;Grange LJ;Reynolds JJ;Cooke SL;McNee GS;Hollingworth R;Woodward BL;Ganesh AN;Smerdon SJ;Nicolae CM;Durlacher-Betzer K;Molho-Pessach V;Abu-Libdeh A;Meiner V;Moldovan GL;Roukos V;Harel T;Brosh RM Jr;Stewart GS
通讯作者：
Stewart GS