Bernhardt Thomas G, de Boer Piet A J
Case Western Reserve University, School of Medicine, Department of Molecular Biology and Microbiology, W239, 10900 Euclid Ave., Cleveland, OH 44106, USA.
Mol Microbiol. 2003 Jun;48(5):1171-82. doi: 10.1046/j.1365-2958.2003.03511.x.
The N-acetylmuramoyl-l-alanine amidases of Escherichia coli (AmiA, B and C) are periplasmic enzymes that remove murein cross-links by cleaving the peptide moiety from N-acetylmuramic acid. Ami- cells form chains, indicating that the amidases help to split the septal murein. Interestingly, cells defective in the twin-arginine protein transport (Tat) pathway show a similar division defect. We find that both AmiA and AmiC are routed to the periplasm via Tat, providing an explanation for the Tat- division phenotype. Taking advantage of the ability of Tat to export prefolded (fluorescent) green fluorescent protein (GFP) to the periplasm, we sublocalized AmiA and AmiC in live cells using functional fusions to GFP. Interestingly, the periplasmic localization of the fusions differed markedly. AmiA-GFP appeared to be dispersed throughout the periplasm in all cells. AmiC-GFP similarly appeared throughout the periplasm in small cells, but was concentrated almost exclusively at the septal ring in constricting cells. Recruitment of AmiC to the ring was mediated by an N-terminal non-amidase targeting domain and required the septal ring component FtsN. AmiC therefore replaces FtsN as the latest known recruit to the septal ring and is the first entirely periplasmic component to be localized.
大肠杆菌的N - 乙酰胞壁酰 - L - 丙氨酸酰胺酶（AmiA、B和C）是周质酶，通过从N - 乙酰胞壁酸上切割肽部分来去除肽聚糖交联。Ami - 细胞形成链状，表明酰胺酶有助于分裂隔膜肽聚糖。有趣的是，双精氨酸蛋白转运（Tat）途径有缺陷的细胞表现出类似的分裂缺陷。我们发现AmiA和AmiC都通过Tat途径被转运到周质，这为Tat - 分裂表型提供了解释。利用Tat将预折叠的（荧光）绿色荧光蛋白（GFP）输出到周质的能力，我们通过与GFP的功能融合在活细胞中对AmiA和AmiC进行亚定位。有趣的是，融合蛋白的周质定位明显不同。AmiA - GFP在所有细胞中似乎分散在整个周质中。AmiC - GFP在小细胞中同样出现在整个周质中，但在正在缢缩的细胞中几乎完全集中在隔膜环处。AmiC募集到环上是由一个N端非酰胺酶靶向结构域介导的，并且需要隔膜环成分FtsN。因此，AmiC取代FtsN成为隔膜环上最新发现的募集蛋白，并且是第一个被定位的完全位于周质的成分。
