Bacterial Cell Biology and Physiology, Swammerdam Institute for Life Science, University of Amsterdam, 1098 XH Amsterdam, The Netherlands.
Int J Mol Sci. 2023 Nov 15;24(22):16355. doi: 10.3390/ijms242216355.
Balancing peptidoglycan (PG) synthesis and degradation with precision is essential for bacterial growth, yet our comprehension of this intricate process remains limited. The NlpI-Prc proteolytic complex plays a crucial but poorly understood role in the regulation of multiple enzymes involved in PG metabolism. In this paper, through fluorescent D-amino acid 7-hydroxycoumarincarbonylamino-D-alanine (HADA) labeling and immunolabeling assays, we have demonstrated that the NlpI-Prc complex regulates the activity of PG transpeptidases and subcellular localization of PBP3 under certain growth conditions. PBP7 (a PG hydrolase) and MltD (a lytic transglycosylase) were confirmed to be negatively regulated by the NlpI-Prc complex by an in vivo degradation assay. The endopeptidases, MepS, MepM, and MepH, have consistently been demonstrated as redundantly essential "space makers" for nascent PG insertion. However, we observed that the absence of NlpI-Prc complex can alleviate the lethality of the mutant. A function of PG lytic transglycosylases MltA and MltD as "space makers" was proposed through multiple gene deletions. These findings unveil novel roles for NlpI-Prc in the regulation of both PG synthesis and degradation, shedding light on the previously undiscovered function of lytic transglycosylases as "space makers" in PG expansion.
精确平衡肽聚糖 (PG) 的合成与降解对于细菌生长至关重要，但我们对这一复杂过程的理解仍然有限。NlpI-Prc 蛋白水解复合物在调节参与 PG 代谢的多种酶的活性方面发挥着关键但尚未被充分理解的作用。在本文中，我们通过荧光 D-氨基酸 7-羟基香豆素羰基氨基-D-丙氨酸 (HADA) 标记和免疫标记测定，证明了 NlpI-Prc 复合物在某些生长条件下调节 PG 转肽酶的活性和 PBP3 的亚细胞定位。通过体内降解测定证实了 PBP7（一种 PG 水解酶）和 MltD（一种裂解转糖基酶）受 NlpI-Prc 复合物的负调控。内切酶 MepS、MepM 和 MepH 一直被证明是新生 PG 插入的冗余必需“空间制造者”。然而，我们观察到 NlpI-Prc 复合物的缺失可以减轻 突变体的致死性。通过多个基因缺失，提出了 PG 裂解转糖基酶 MltA 和 MltD 作为“空间制造者”的功能。这些发现揭示了 NlpI-Prc 在 PG 合成和降解调节中的新作用，揭示了裂解转糖基酶作为 PG 扩张“空间制造者”的以前未被发现的功能。
