摘要

肽聚糖（PG）是围绕大多数细菌细胞以防止细胞质膜的渗透破裂的基本交联聚合物。其合成依赖于青霉素结合蛋白，即β-内酰胺抗生素的靶标。包括机会性病原体铜绿假单胞菌在内的许多革兰氏阴性细菌由于被称为AmpC的染色体编码的β-内酰胺酶而对β-内酰胺具有抗性。在铜绿假单胞菌中，ampC基因的表达受到严格调控，其诱导与细胞壁应激相关。我们推测报告基因融合到ampC启动子将允许我们鉴定在维持细胞壁内稳态方面有缺陷的突变体，从而揭示在该过程中涉及的新因素。因此筛选具有升高的ampC启动子活性的突变体的转座子诱变的铜绿假单胞菌文库。作为屏幕如预期工作的指示，分离了具有破坏dacB基因的转座子的突变体。 DacB的缺陷以前涉及ampC诱导和对β-内酰胺抗生素的临床抗性。该屏幕还揭露了murU和PA3172突变体，经进一步表征后，其显示出几乎相同的耐药性和灵敏度特征。我们提供的遗传证据表明，PA3172重新命名为mupP，编码预测在广泛分布于革兰氏阴性菌中的MurU PG回收途径中发挥功能的缺失磷酸酶。

Peptidoglycan (PG) is an essential cross-linked polymer that surrounds most bacterial cells to prevent osmotic rupture of the cytoplasmic membrane. Its synthesis relies on penicillin-binding proteins, the targets of beta-lactam antibiotics. Many Gram-negative bacteria, including the opportunistic pathogenPseudomonas aeruginosa, are resistant to beta-lactams because of a chromosomally encoded beta-lactamase called AmpC. In P. aeruginosa, expression of the ampC gene is tightly regulated and its induction is linked to cell wall stress. We reasoned that a reporter gene fusion to the ampC promoter would allow us to identify mutants defective in maintaining cell wall homeostasis and thereby uncover new factors involved in the process. A library of transposon-mutagenizedP. aeruginosa was therefore screened for mutants with elevated ampC promoter activity. As an indication that the screen was working as expected, mutants with transposons disrupting the dacB gene were isolated. Defects in DacB have previously been implicated in ampC induction and clinical resistance to beta-lactam antibiotics. The screen also uncovered murU and PA3172 mutants that, upon further characterization, displayed nearly identical drug resistance and sensitivity profiles. We present genetic evidence that PA3172, renamed mupP, encodes the missing phosphatase predicted to function in the MurU PG recycling pathway that is widely distributed among Gram-negative bacteria.

http://mbio.asm.org/content/8/2/e00102-17.short