Phage Lysis

噬菌体对宿主细胞的裂解是生物圈中最常见的细胞杀灭事件，是一种真正的 基本流程。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。 一般有两种模式：dsDNA噬菌体使用的多基因裂解(MGL)和单基因裂解(SGL)， 被小的单链核酸噬菌体使用。至少，MGL系统需要一种溶菌酶，即 内溶素，它降解细胞壁或肽聚糖(PG)，以及一种小的膜蛋白，Holin，它 积极规划内毒素的功能。至少另外10类噬菌体裂解蛋白也被 已确定，包括在革兰氏阴性感染中起破坏外膜功能的Spanins或 作为霍林和内溶素功能的调节者。裂解途径有两个步骤，既能对 引起宿主膜的生物物理变化，以及具有多个动态膜的例子 拓扑和大量的四元重排，最终导致细菌膜上的孔洞 前所未有的微米级。总体而言，这些复杂的MGL系统使Lysis成为一种精确控制的、全或- 没有任何现象。 相反，小的单链DNA和单链RNA噬菌体没有MGL系统的基因组空间。取而代之的是一张 SGL(单基因裂解)蛋白最终导致宿主PG生物合成或动态平衡的功能障碍 导致宿主自溶。已经建立了一类阻断细胞壁生物合成步骤的SGL，并 被指定为蛋白质抗生素，但生物信息学和生物信息学确定的20多种其他SGL的目标 噬菌体遗传学尚不清楚。 在接下来的五年里，重点将不仅放在引人注目的Spanins上，这些Spanins在 裂解，但也对两类新的MGL蛋白质：释放素和破碎素。Releasin在许可方面是独一无二的 内毒素的动态膜拓扑结构。Disruptins是一种小的两亲性蛋白质，用于 削弱外膜；令人惊讶的是，当它们被纯化并在体外使用时，它们的功能就像噬菌体编码的 哺乳动物细胞产生的阳离子抗菌肽(CAMP)的不同版本。独一无二的力量 噬菌体遗传学将被用来确定这两种新的MGL蛋白的机制。我们的生物物理 结构合作者将得到突变体、表型和结构的支持，以用于 在原子水平和受感染的单个细胞的背景下表征裂解。在SGL地区， 最近对单链RNA噬菌体元基因组的超扩增将被用来解决许多 新的SGL蛋白。假设单链RNA噬菌体SGL蛋白已经进化到攻击宿主细胞的每一步 壁合成和动态平衡。此外，一种新的模型，即SGL的主要类别通过绑定通用细胞来起作用 WALL前体，Lipid II，将进行测试。

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.