Bacterial cell division requires septal peptidoglycan (sPG) synthesis by the divisome complex. Treadmilling of the essential tubulin homologue FtsZ has been implicated in septal constriction, though its precise role remains unclear. Here we used live-cell single-molecule imaging of the divisome transpeptidase PBP2B to investigate sPG synthesis dynamics in Bacillus subtilis. In contrast to previous models, we observed a single population of processively moving PBP2B molecules whose motion is driven by peptidoglycan synthesis and is not associated with FtsZ treadmilling. However, despite the asynchronous motions of PBP2B and FtsZ, a partial dependence of PBP2B processivity on FtsZ treadmilling was observed. Additionally, through single-molecule counting experiments we provide evidence that the divisome synthesis complex is multimeric. Our results support a model for B. subtilis division where a multimeric synthesis complex follows a single track dependent on sPG synthesis whose activity and dynamics are asynchronous with FtsZ treadmilling.

细菌细胞分裂需要分裂体复合物合成隔膜肽聚糖（sPG）。重要的微管蛋白同源物 FtsZ 的跑动与隔膜收缩有关，但其确切作用仍不清楚。在这里，我们使用分裂体转肽酶 PBP2B 的活细胞单分子成像来研究枯草芽孢杆菌中 sPG 的合成动力学。与之前的模型相比，我们观察到单个群体的持续运动的 PBP2B 分子，其运动由肽聚糖合成驱动，与 FtsZ 跑步无关。然而，尽管 PBP2B 和 FtsZ 进行异步运动，但仍观察到 PBP2B 持续性对 FtsZ 跑步的部分依赖性。此外，通过单分子计数实验，我们提供了分裂合成复合物是多聚体的证据。我们的结果支持枯草芽孢杆菌分裂的模型，其中多聚体合成复合物遵循依赖于 sPG 合成的单一轨道，其活性和动态与 FtsZ 跑步机异步。