Characterization of the Bacterial BREX Phage Restriction System

细菌 BREX 噬菌体限制系统的表征

• 批准号: 10291285
• 负责人: Brett Kian Kaiser
• 金额: $ 28.4万
• 依托单位: SEATTLE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-15 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10291285
• 关键词: ATPase Domain; Acinetobacter; Address; Adsorption; Affect; Affinity; Archaeal Genome; Architecture; Area; Bacillus cereus; Bacteria; Bacterial Antibiotic Resistance; Bacterial Genome; Bacteriophages; Biochemical; Bioinformatics; Biological; Biological Assay; Complement; Complex; Cryoelectron Microscopy; Crystallization; DNA; DNA Binding; DNA Modification Methylases; DNA Restriction-Modification Enzymes; DNA biosynthesis; Data; Discrimination; Disease; Electrophoretic Mobility Shift Assay; Enzymes; Escherichia coli; Exclusion; Family; Fibrinogen; Gel; Gene Expression; Genes; Genetic Transcription; Genome; Genomics; Goals; Health; Human; Individual; Infection; Lead; Location; Maps; Mass Spectrum Analysis; Mediating; Methylation; Methyltransferase; Mutation; Mutation Analysis; Open Reading Frames; Peptide Hydrolases; Phosphoric Monoester Hydrolases; Proteins; Race; Regulation; Research Design; Resolution; Role; Sequence Homology; Specificity; Structure; System; Testing; Virus Diseases; arm; base; disorder prevention; endopeptidase Clp; endopeptidase La; genomic locus; insight; member; microbiome; mutant; novel; overexpression; particle; prevent; protein degradation

SUMMARY The arms race between bacteria and phage has driven bacteria to evolve a wide range of defense systems, many of which remain to be characterized. A largely uncharacterized phage restriction system called ‘BREX’ (‘Bacteriophage Exclusion’), initially described 35 years ago, is widely distributed in bacterial and archaeal genomes. The mechanism of BREX-mediated phage restriction is likely complex given the large number of genes (4-8 genes, depending on the subfamily) and range of enzymatic activities encoded by BREX systems. Studies of type 1 and type 2 BREX systems (there are 6 total subfamilies) have demonstrated that the PglX DNA methyltransferase establishes self / non-self discrimination by modifying the bacterial genome. However, BREX-mediated restriction does not appear to function by degrading the phage genome, which is a surprise given the well-characterized role for similar methyltransferase enzymes in restriction-modification (R-M) systems. Type 1 BREX systems inhibit phage after adsorption and prior to DNA replication, but further details about the mechanism of phage restriction are not known. The long-term goal is to understand the mechanism by which BREX restricts phage, including elucidating the functional role of each BREX component. Our understanding of BREX-mediated phage restriction is limited because the function of most individual BREX proteins is not understood, nor is it understood how they are regulated. To address these questions, we are characterizing the Acinetobacter type 1 BREX system using biochemical and structural studies combined with biological activity assays using an E. coli-based phage infection system. The first aim will address the hypothesis that Brx0, an Acinetobacter-specific factor (i.e. not present in other BREX Type 1 systems), transcriptionally regulates other BREX ORFs. The second aim will address the hypothesis that BrxL is a AAA+ chambered protease that specifically targets substrate proteins for degradation. Uncovering the roles of both proteins will provide key insight into the mechanism of BREX-mediated phage restriction, not only in Acinetobacter but as a general strategy in other bacterial clades. Gaining a fundamental understanding of bacterial defense systems could impact human health in numerous ways, including contributing to strategies to treat antibiotic-resistant bacteria and understanding how the microbiome relates to health and disease.

总结 细菌和噬菌体之间的军备竞赛促使细菌进化出各种各样的防御系统， 其中许多仍有待鉴定。一个很大程度上未表征的噬菌体限制性系统，称为“BREX” （“噬菌体排除”），最初描述于35年前，广泛分布于细菌和古细菌中。 基因组BREX介导的噬菌体限制的机制可能是复杂的，因为存在大量的噬菌体。 基因（4-8个基因，取决于亚家族）和由BREX系统编码的酶活性范围。 对1型和2型BREX系统（总共有6个亚家族）的研究表明，PglX DNA甲基转移酶通过修饰细菌基因组建立自我/非自我区分。然而，在这方面， BREX介导的限制似乎并不通过降解噬菌体基因组来发挥作用，这是一个令人惊讶的结果。 考虑到类似的甲基转移酶在限制修饰（R-M）中的良好表征的作用， 系统. 1型BREX系统在吸附后和DNA复制前抑制噬菌体，但进一步的细节 关于噬菌体限制的机制尚不清楚。长期目标是了解机制 BREX限制噬菌体的机制，包括阐明每个BREX组分的功能作用。 我们对BREX介导的噬菌体限制的理解是有限的，因为大多数单个噬菌体的功能是有限的。 BREX蛋白不被理解，也不知道它们是如何被调节的。为了解决这些问题，我们 正在使用生物化学和结构研究相结合的方法来表征不动杆菌1型BREX系统 用E.大肠杆菌噬菌体感染系统。第一个目标是解决 假设Brx 0，一种不动杆菌特异性因子（即不存在于其他BREX 1型系统中）， 转录调控其他BREX ORF。第二个目标将解决BrxL是AAA+的假设 专门针对底物蛋白质进行降解的有室蛋白酶。揭示两者的作用 蛋白质将提供BREX介导的噬菌体限制机制的关键见解，不仅在 不动杆菌，但作为一般战略，在其他细菌的分支。 获得对细菌防御系统的基本了解可能会在许多方面影响人类健康 方法，包括有助于治疗耐药细菌的策略，并了解 微生物组与健康和疾病有关。