在DNA聚合酶β E295K突变体中不利的静电和固体相互作用干扰了酶的通路
Yunlang Li1, Chelsea L Gridley, Joachim Jaeger
1Department of Chemistry and Courant Institute of Mathematical Sciences, New York University, 251 Mercer Street, New York, New York 10012, USA.
Journal of the American Chemical Society
|June 2, 2012
概括
在DNA聚合酶β (pol β) 中的突变可以导致癌症. E295K多β突变导致扭曲的活性部位和更高的能量屏障,阻碍DNA修复,并可能导致不活性.
科学领域:
- 生物化学和分子生物学
- 结构生物学 结构生物学
- 酶学 是一种酶学.
背景情况:
- 在DNA聚合酶β (pol β) 中的突变涉及~30%的人类癌症.
- E295K多β突变通过基切除修复干扰与胃癌有关.
研究的目的:
- 阐明野生型聚β和E295K突变体之间的原子和能量差异.
- 了解E295K突变的酶途径的结构变化和能量格局.
主要方法:
- 在2.5 Å时,E295K突变二进制复合物的晶体结构分辨率为2.5 Å.
- 应用过渡路径采样 (TPS) 来绘制E295K多β突变的关闭路径.
- 对形状变化,过渡状态和能量障碍的计算分析.
主要成果:
- 与野生类型的pol β相比,E295K突变体表现出明显的关闭途径,具有改变的过渡状态和能量.
- 封闭状态的E295K具有更扭曲的活性部位和明显更高的能量屏障 (65±11kJ/mol),而不是野生类型的pol β.
- 由于不良相互作用,Arg258的旋转被确定为E295K关闭通路中的速度限制步骤.
结论:
- 在E295K突变体中,扭曲的活性部位和高能量屏障可能导致其观察到的不活性.
- E295K可能与类似的亲和力结合DNA,但在随后的化学步骤中受损,可能超越野生类型的pol β.
- 这些发现凸显了结构完整性和能量学在聚β功能的关键作用,并建议进一步研究突变效应.
相关概念视频
01:31Proofreading
Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore, it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
Errors During Replication are Corrected by the DNA Polymerase Enzyme
Errors During Replication are Corrected by the DNA Polymerase Enzyme
01:43Proofreading
Overview
01:20Mismatch Repair
Organisms are capable of detecting and fixing nucleotide mismatches that occur during DNA replication. This sophisticated process requires identifying the new strand and replacing the erroneous bases with correct nucleotides. Mismatch repair is coordinated by many proteins in both prokaryotes and eukaryotes.
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...
01:36Mismatch Repair
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02:37Restarting Stalled Replication Forks
DNA replication is initiated at sites containing predefined DNA sequences known as origins of replication. DNA is unwound at these sites by the minichromosome maintenance (MCM) helicase and other factors such as Cdc45 and the associated GINS complex.The unwound single strands are protected by replication protein A (RPA) until DNA polymerase starts synthesizing DNA at the 5’ end of the strand in the same direction as the replication fork. To prevent the replication fork from falling apart, a...
02:10Translesion DNA Polymerases
Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...