Role of M. tuberculosis error-prone DNA polymerase DnaE2 in mutagenesis and drug resistance

结核分枝杆菌易错 DNA 聚合酶 DnaE2 在诱变和耐药性中的作用

基本信息

批准号：
9262376
负责人：
Kathleen A McDonough
金额：
$ 24.94万
依托单位：
WADSWORTH CENTER
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-12-01 至 2018-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9262376
关键词：
Bacteria Bacterial DNA Biochemical Biological Biological Assay Cells Closure by clamp Complex DNA DNA Damage DNA Polymerase III DNA biosynthesis DNA replication fork DNA-Directed DNA Polymerase Disease Drug resistance Drug resistance in tuberculosis Environment Evolution Family Fluoroquinolones Foundations Frequencies Future Genes Genetic Genetic Predisposition to Disease Genome Genomics Goals In Vitro Incidence Individual Induced Mutation Infection Intervention Mediating Modeling Molecular Molecular Genetics Multienzyme Complexes Mus Mutagenesis Mutation Mycobacterium tuberculosis Nucleotides Pathogenicity Plasmids Polymerase Process Property Proteins Research Resistance Role Site Slide Stress Testing Therapeutic Tuberculosis Two-Hybrid System Techniques UV induced Work Yeasts base clinically relevant combat global health improved inhibitor/antagonist light effects novel prevent protein complex protein protein interaction reconstitution resistance gene resistance mechanism response stressor success therapeutic target tool tuberculosis drugs

项目摘要

An increasing challenge in the control of tuberculosis (TB) worldwide is the emergence and spread of drug resistance in Mycobacterium tuberculosis (Mtb), the causative agent of TB. Evolution of drug resistance in Mtb is associated with chromosomal mutations, not with the acquisition of resistance plasmids or transferred resistance genes. DnaE2 is required for induced mutagenesis in Mtb and belongs to the C-family of bacterial DNA polymerases that are responsible for genome replication. dnaE2 is a component of the imuA-imuB-dnaE2 cassette and all three genes are essential for DNA damage-induced mutagenesis. The DnaE2-ImuA-ImuB protein complex is proposed to associate with the sliding clamp processivity factor (the β-subunit of DNA pol III) through interactions with the ImuB subunit, allowing DnaE2 access to sites of replication where it can perform error-prone DNA synthesis. However, this enzyme complex has not been well characterized. We hypothesize that error prone DNA synthesis by the DnaE2-ImuA-ImuB polymerase complex contributes to drug resistance during TB infection. Here, we will (Aim 1) evaluate the importance of the complex for Mtb survival, mutagenesis and drug resistance in host-associated stress conditions, and (Aim 2) characterize the assembly, catalytic activity and mutational properties of this putative error-prone polymerase. These studies are essential steps towards understanding the mechanistic bases of drug resistance emergence in Mtb and the potential use of this error-prone DNA polymerase complex as a target for novel adjunctive therapies to combat drug- resistance in Mtb.

全球结核病（TB）的控制越来越大的挑战是药物的出现和传播结核分枝杆菌（MTB）的耐药性，TB的耐药性进化。与染色体突变有关，而与耐药质粒的获取或转移抗性基因。负责基因组复制的DNA聚合酶。盒子和所有三个基因对于DNA损伤诱变的诱变至关重要蛋白质复合物被支撑以与滑动夹具的加工性因子（DNA POL的β亚基）相关联 iii）通过与IMUB亚基的相互作用，允许DNAE2访问复制位置但是，易于使用误差的DNA合成假设该误差易受DNAE2-IMUA-IMUB聚合酶复合物的DNA合成，从而有助于药物在TB感染期间，我们将（AIM 1）评估该建筑群的重要性在与宿主相关的应力条件下的mutajenesis和耐药性，（AIM 2）将组件表征该假定误差的催化活性和突变特性了解MTB中耐药性出现的机械基础的步骤和潜在用途在这个容易出错的DNA聚合酶复合酶时，忘记了忘记忘记忘记忘记忘记忘记忘记忘记忘记忘记忘记忘记接受疗法对抗药物 - MTB的阻力。