Tulum Isil, Tahara Yuhei O, Miyata Makoto
Graduate School of Science, Osaka City University, Osaka 558-8585, Japan.
The OCU Advanced Research Institute for Natural Science and Technology (OCARINA), Osaka City University, Osaka 558-8585, Japan.
Microscopy (Oxf). 2019 Dec 3;68(6):441-449. doi: 10.1093/jmicro/dfz033.
Peptidoglycan, which is the main component of the bacterial cell wall, is a heterogeneous polymer of glycan strands cross-linked with short peptides and is synthesized in cooperation with the cell division cycle. Although it plays a critical role in bacterial survival, its architecture is not well understood. Herein, we visualized the architecture of the peptidoglycan surface in Bacillus subtilis at the nanometer resolution, using quick-freeze, deep-etch electron microscopy (EM). Filamentous structures were observed on the entire surface of the cell, where filaments about 11 nm wide formed concentric circles on cell poles, filaments about 13 nm wide formed a circumferential mesh-like structure on the cylindrical part and a 'piecrust' structure was observed at the boundary. When growing cells were treated with lysozyme, the entire cell mass migrated to one side and came out from the cell envelope. Fluorescence labeling showed that lysozyme preferentially bound to a cell pole and cell division site, where the peptidoglycan synthesis was not complete. Ruffling of surface structures was observed during EM. When cells were treated with penicillin, the cell mass came out from a cleft around the cell division site. Outward curvature of the protoplast at the cleft seen using EM suggested that turgor pressure was applied as the peptidoglycan was not damaged at other positions. When muropeptides were depleted, surface filaments were lost while the rod shape of the cell was maintained. These changes can be explained on the basis of the working points of the chemical structure of peptidoglycan.
肽聚糖是细菌细胞壁的主要成分，是一种由聚糖链与短肽交联而成的异质聚合物，并与细胞分裂周期协同合成。尽管它在细菌存活中起着关键作用，但其结构尚未得到充分了解。在此，我们使用快速冷冻、深度蚀刻电子显微镜（EM），以纳米分辨率观察了枯草芽孢杆菌中肽聚糖表面的结构。在细胞的整个表面观察到丝状结构，其中约11nm宽的细丝在细胞极上形成同心圆，约13nm宽的细丝在圆柱部分形成圆周网状结构，并在边界处观察到“馅饼皮”结构。当生长中的细胞用溶菌酶处理时，整个细胞团迁移到一侧并从细胞膜中出来。荧光标记显示溶菌酶优先结合到肽聚糖合成未完成的细胞极和细胞分裂位点。在电子显微镜观察期间观察到表面结构的褶皱。当细胞用青霉素处理时，细胞团从细胞分裂位点周围的裂缝中出来。使用电子显微镜观察到的裂缝处原生质体的向外弯曲表明，由于肽聚糖在其他位置未受损，因此施加了膨压。当胞壁肽耗尽时，表面细丝消失，而细胞的杆状形状得以维持。这些变化可以根据肽聚糖化学结构的作用点来解释。
