A Novel in vivo Cell-Wall Labeling Approach Sheds New Light on Peptidoglycan Synthesis in Escherichia coli
A Novel in vivo Cell-Wall Labeling Approach Sheds New Light on Peptidoglycan Synthesis in Escherichia coli
复制标题
DOI：
10.1002/cbic.201000552
复制
发表时间：
2011-05-02
期刊：
影响因子：
3.2
通讯作者：
den Blaauwen, Tanneke
中科院分区：
文献类型：
--
作者：
Olrichs, Nick K.;Aarsman, Mirjam E. G.;den Blaauwen, Tanneke
用一种新的标记方法研究了肽聚糖的合成和周转与细胞生长和分裂的关系。该方法涉及通过细胞壁再循环途径将荧光标记的肽聚糖前体掺入细胞壁中。我们表明，大肠杆菌是能够进口外源性添加的murein三肽标记的N-7-硝基-2，1，3-苯并恶二唑-4-基（AeK-NBD）进入细胞质中，它进入肽聚糖的生物合成途径，导致荧光标记，特别是位于细胞壁。当野生型细胞在荧光肽存在下生长时，肽聚糖在经历延伸的细胞中被均匀地标记。处于分裂过程中的细胞在细胞中期缺乏标记的肽聚糖。细胞分裂突变体的标记模式的分析表明，未标记的肽聚糖的发生是依赖于FtsZ的存在，但独立的FtsQ和FtsI。在三酰胺酶突变体（Δ amiABC）的分裂位点处的荧光积累揭示了AeK-NBD被并入隔膜肽聚糖中。AmiC被证明参与在野生型细胞分裂位点标记的肽聚糖侧链的快速去除。由于AmiC的间隔定位依赖于FtsQ和FtsI，这表明存在另一种直接依赖于FtsZ的肽聚糖水解酶活性。
Peptidoglycan synthesis and turnover in relation to cell growth and division has been studied by using a new labeling method. This method involves the incorporation of fluorescently labeled peptidoglycan precursors into the cell wall by means of the cell-wall recycling pathway. We show that Escherichia coli is able to import exogenous added murein tripeptide labeled with N-7-nitro-2,1,3-benzoxadiazol-4-yl (AeK-NBD) into the cytoplasm where it enters the peptidoglycan biosynthesis route, resulting in fluorescent labels specifically located in the cell wall. When wild-type cells were grown in the presence of the fluorescent peptide, peptidoglycan was uniformly labeled in cells undergoing elongation. Cells in the process of division displayed a lack of labeled peptidoglycan at mid-cell. Analysis of labeling patterns in cell division mutants showed that the occurrence of unlabeled peptidoglycan is dependent on the presence of FtsZ, but independent of FtsQ and FtsI. Accumulation of fluorescence at the division sites of a triple amidase mutant (Delta amiABC) revealed that AeK-NBD is incorporated into septal peptidoglycan. AmiC was shown to be involved in the rapid removal of labeled peptidoglycan side chains at division sites in wild-type cells. Because septal localization of AmiC is dependent on FtsQ and FtsI, this points to the presence of another peptidoglycan hydrolase activity directly dependent on FtsZ.