四种Y家族损伤旁路DNA聚合酶的同源性建模:大肠杆菌Pol IV与人Pol κ为直系同源物,大肠杆菌Pol V与人Pol η为直系同源物的情况。

Homology modeling of four Y-family, lesion-bypass DNA polymerases: the case that E. coli Pol IV and human Pol kappa are orthologs, and E. coli Pol V and human Pol eta are orthologs.

作者信息

Lee Chiu Hong, Chandani Sushil, Loechler Edward L

机构信息

Department of Physics, Harvard University, Cambridge, MA 02138, USA.

出版信息

J Mol Graph Model. 2006 Sep;25(1):87-102. doi: 10.1016/j.jmgm.2005.10.009. Epub 2006 Jan 4.

Abstract

Y-family DNA polymerases (DNAPs) are a superfamily of evolutionarily related proteins that exist in cells to bypass DNA damage caused by both radiation and chemicals. Cells have multiple Y-family DNAPs, presumably to conduct translesion synthesis (TLS) on DNA lesions of varying structure and conformation. The potent, ubiquitous environmental mutagen/carcinogen benzo[a]pyrene (B[a]P) induces all classes of mutations with G-->T base substitutions predominating. We recently showed that a G-->T mutagenesis pathway for the major adduct of B[a]P ([+ta]-B[a]P-N2-dG) in Escherichia coli depends on Y-family member DNAP V. Since no X-ray crystal study for DNAP V has been reported, no structure is available to help in understanding the structural basis for dATP insertion associated with G-->T mutations from [+ta]-B[a]P-N2-dG. Herein, we do homology modeling to construct a model for UmuC, which is the polymerase subunit of DNAP V. The sequences of eight Y-family DNAPs were aligned based on the positioning of conserved amino acids and an analysis of conserved predicted secondary structure, as well as insights gained from published X-ray structures of five Y-family members. Starting coordinates for UmuC were generated from the backbone coordinates for the Y-family polymerase Dpo4 for reasons discussed, and were refined using molecular dynamics with CHARMM 27. A survey of the literature revealed that E. coli DNAP V and human DNAP eta show a similar pattern of dNTP insertion opposite a variety of DNA lesions. Furthermore, E. coli DNAP IV and human DNAP kappa show a similar dNTP insertional pattern with these same DNA lesions, although the insertional pattern for DNAP IV/kappa differs from the pattern for DNAPs V/eta. These comparisons prompted us to construct and refine models for E. coli DNAP IV and human DNAPs eta and kappa as well. The dNTP/template binding pocket of all four DNAPs was inspected, focusing on the array of seven amino acids that contact the base of the incoming dNTP, as well as the template base. DNAPs V and eta show similarities in this array, and DNAPs IV and kappa also show similarities, although the arrays are different for the two pairs of DNAPs. Thus, there is a correlation between structural similarities and insertional similarities for the pairs DNAPs V/eta and DNAPs IV/kappa. Although the significance of this correlation remains to be elucidated, these observations point the way for future experimental studies.

摘要

Y家族DNA聚合酶(DNAPs)是一组进化相关蛋白的超家族,存在于细胞中以绕过由辐射和化学物质引起的DNA损伤。细胞中有多种Y家族DNAPs,推测是为了对不同结构和构象的DNA损伤进行跨损伤合成(TLS)。强效且普遍存在的环境诱变剂/致癌物苯并[a]芘(B[a]P)可诱导所有类型的突变,其中G→T碱基替换占主导。我们最近发现,大肠杆菌中B[a]P主要加合物([+ta]-B[a]P-N2-dG)的G→T诱变途径依赖于Y家族成员DNAP V。由于尚未有关于DNAP V的X射线晶体学研究报道,所以没有结构信息可用于帮助理解与[+ta]-B[a]P-N2-dG导致的G→T突变相关的dATP插入的结构基础。在此,我们进行同源建模以构建DNAP V的聚合酶亚基UmuC的模型。基于保守氨基酸的定位、保守预测二级结构的分析以及从已发表的五个Y家族成员的X射线结构中获得的见解,对八个Y家族DNAPs的序列进行了比对。出于所讨论的原因,UmuC的起始坐标由Y家族聚合酶Dpo4的主链坐标生成,并使用CHARMM 27通过分子动力学进行了优化。文献调查显示,大肠杆菌DNAP V和人类DNAP eta在面对多种DNA损伤时显示出相似的dNTP插入模式。此外,大肠杆菌DNAP IV和人类DNAP kappa在面对这些相同的DNA损伤时也显示出相似的dNTP插入模式,尽管DNAP IV/kappa的插入模式与DNAPs V/eta的模式不同。这些比较促使我们也构建并优化了大肠杆菌DNAP IV以及人类DNAPs eta和kappa的模型。检查了所有四种DNAPs的dNTP/模板结合口袋,重点关注与进入的dNTP碱基以及模板碱基接触的七个氨基酸阵列。DNAPs V和eta在这个阵列中显示出相似性,DNAPs IV和kappa也显示出相似性,尽管这两对DNAPs的阵列不同。因此,DNAPs V/eta和DNAPs IV/kappa这两对在结构相似性和插入相似性之间存在相关性。尽管这种相关性的意义仍有待阐明,但这些观察结果为未来的实验研究指明了方向。

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