Phage Lysis

噬菌体裂解

• 批准号: 10631067
• 负责人: RYLAND F YOUNG
• 金额: $ 40.02万
• 依托单位: TEXAS A&M AGRILIFE RESEARCH
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10631067
• 关键词: Anabolism; Antibiotics; Antimicrobial Cationic Peptides; Area; Autolysis; Bacteria; Bacterial Infections; Bacteriophages; Binding; Bioinformatics; Biophysics; Cell Wall; Cells; Complex; Cytolysis; Enzymes; Event; Functional disorder; Genes; Genetic; Genomics; Homeostasis; In Vitro; Infection; Licensing; Lipids; Mammalian Cell; Membrane; Membrane Fusion; Membrane Proteins; Metagenomics; Modeling; Molecular; Multi-Drug Resistance; Nucleic Acids; Pathway interactions; Peptidoglycan; Phenotype; Process; Proteins; Single-Stranded DNA; System; Testing; Virus; clinically relevant; design; ds-DNA; endolysin; mutant; programs; tool

Lysis of the host cell by bacteriophage is, as the most frequent cytocidal event in the biosphere, a truly fundamental process. In addition, understanding the molecular basis of phage lysis is now clinically relevant because phage therapy is emerging as an important tool against multi-drug resistant bacterial infections. There are two general modes: Multi-Gene Lysis (MGL), used by dsDNA phages, and Single-Gene Lysis (SGL), used by small single-strand nucleic acid phages. At minimum, MGL systems require a muralytic enzyme, the endolysin, that degrades the cell wall or peptidoglycan (PG), and a small membrane protein, the holin, that actively programs the function of the endolysin. At least 10 more classes of phage lysis proteins have also been identified, including spanins functioning in destruction of the outer membrane in Gram-negative infections or acting as regulators of holin and endolysin function. The lysis pathways have steps that both respond to and cause biophysical changes in the host membrane, as well as featuring multiple examples of dynamic membrane topology and massive quaternary rearrangements, ultimately resulting in holes in the bacterial membrane of unprecedented micron-scale. Overall, these complex MGL systems make lysis a precisely-controlled, all-or- nothing phenomenon. In contrast, the small ssDNA and ssRNA phages have no genomic room for MGL systems. Instead a single Sgl (single gene lysis) protein acts to cause dysfunction in host PG biosynthesis or homeostasis, eventually leading to a host autolysis. One class of Sgl’s that block steps in cell wall biosynthesis has been established and designated as Protein Antibiotics, but the target of more than 20 other Sgl’s identified by bioinformatics and phage genetics is not known. In the next five years, the focus will not only be on the remarkable spanins, which fuse membranes during lysis, but also on two new classes of MGL proteins: releasins and disruptins. Releasins are unique in licensing dynamic membrane topology of endolysins. Disruptins are small, amphipathic proteins that are used to weaken the outer membrane; surprisingly, when purified and used in vitro, they function as phage-encoded versions of the cationic antimicrobial peptides (CAMPs) produced by mammalian cells. The unique power of phage genetics will be used to determine the mechanisms of both these new MGL proteins. Our biophysical and structural collaborators will be supp;oed with mutants, phenotypes and constructs to be used in characterizing lysis at both the atomic level and in the context of the infected single cell. In the SGL area, the recent hyper-expansion of the metagenomics of ssRNA phages will be exploited to solve the targets of many new Sgl proteins. The hypothesis is that ssRNA phage Sgl proteins have evolved to attack every step in host cell wall synthesis and homeostasis. Also, a new model that a major class of Sgl’s acts by binding the universal cell wall precursor, Lipid II, will be tested.

作为生物圈中最常见的杀细胞事件，噬菌体裂解宿主细胞是一种真正的杀细胞作用。 基本过程。此外，了解噬菌体裂解的分子基础现在具有临床意义 因为噬菌体疗法正在成为对抗多重耐药细菌感染的重要工具。 有两种通用模式：双链 DNA 噬菌体使用的多基因裂解 (MGL) 和单基因裂解 (SGL)， 由小型单链核酸噬菌体使用。至少，MGL 系统需要一种胞壁溶解酶，即 内溶素，可降解细胞壁或肽聚糖 (PG)，以及一种小膜蛋白，穴蛋白，可降解细胞壁或肽聚糖 (PG) 主动编程内溶素的功能。至少另外 10 类噬菌体裂解蛋白也已被 已鉴定，包括在革兰氏阴性感染中破坏外膜的西班牙蛋白或 作为穴蛋白和细胞内溶素功能的调节剂。裂解途径具有既响应又响应的步骤 引起宿主膜的生物物理变化，以及具有动态膜的多个例子 拓扑结构和大规模四元重排，最终导致细菌膜上出现孔洞 前所未有的微米级。总体而言，这些复杂的 MGL 系统使裂解成为一种精确控制的、全或- 没什么现象。 相比之下，小型 ssDNA 和 ssRNA 噬菌体没有 MGL 系统的基因组空间。而是单个 Sgl（单基因裂解）蛋白最终会导致宿主 PG 生物合成或体内平衡功能障碍 导致宿主自溶。已经建立了一类阻止细胞壁生物合成步骤的 Sgl 被指定为蛋白质抗生素，但生物信息学和其他 20 多个 Sgl 的目标 噬菌体遗传学尚不清楚。 在接下来的五年里，重点将不仅是非凡的跨膜蛋白，它在过程中融合膜 裂解，而且还作用于两类新的 MGL 蛋白：释放素和破坏素。发行版在许可方面是独一无二的 内溶素的动态膜拓扑结构。干扰素是一种小的两亲性蛋白质，用于 削弱外膜；令人惊讶的是，当在体外纯化和使用时，它们充当噬菌体编码的 由哺乳动物细胞产生的阳离子抗菌肽（CAMP）的版本。独特的力量 噬菌体遗传学将用于确定这两种新 MGL 蛋白的机制。我们的生物物理学 和结构合作者将获得突变体、表型和构建体用于 在原子水平和受感染的单细胞背景下表征裂解。在 SGL 区域， 最近 ssRNA 噬菌体宏基因组学的超扩展将被用来解决许多目标 新的Sgl蛋白。假设 ssRNA 噬菌体 Sgl 蛋白已经进化到可以攻击宿主细胞的每一步 壁合成和稳态。另外，Sgl的主要类通过绑定通用单元来起作用的新模型 将测试壁前体脂质 II。