Center for Health and Behavioral Sciences, Department of Science and Technology, Bryant University, 1150 Douglas Pike, Smithfield, RI 02917, USA.
Department of Basic Sciences, Touro University Nevada, College of Osteopathic Medicine, Henderson, NV 89014, USA.
Microbiology (Reading). 2022 Apr;168(4). doi: 10.1099/mic.0.001182.
Despite renewed interest, development of chemical biology methods to study peptidoglycan metabolism has lagged in comparison to the glycobiology field in general. To address this, a panel of diamides were screened against the Gram-positive bacterium to identify inhibitors of bacterial growth. The screen identified the diamide masarimycin as a bacteriostatic inhibitor of growth with an MIC of 8 µM. The diamide inhibited detergent-induced autolysis in a concentration-dependent manner, indicating perturbation of peptidoglycan degradation as the mode-of-action. Cell based screening of masarimycin against a panel of autolysin mutants, identified a higher MIC against a Δ strain lacking an endo-N-acetylglucosaminidase involved in cell division. Subsequent biochemical and phenotypic analyses suggested that the higher MIC was due to an indirect interaction with LytB. Further analysis of changes to the cell surface in masarimycin treated cells identified the overexpression of several moonlighting proteins, including elongation factor Tu which is implicated in regulating cell shape. Checkerboard assays using masarimycin in concert with additional antibiotics identified an antagonistic relationship with the cell wall targeting antibiotic fosfomycin, which further supports a cell wall mode-of-action.
尽管人们重新产生了兴趣，但与一般的糖生物学领域相比，用于研究肽聚糖代谢的化学生物学方法的发展一直较为滞后。为了解决这个问题，一组二酰胺被筛选针对革兰氏阳性菌，以鉴定抑制细菌生长的抑制剂。该筛选发现二酰胺马沙霉素是一种具有 8 μM MIC 的抑菌性生长抑制剂。该二酰胺以浓度依赖的方式抑制去污剂诱导的自溶，表明肽聚糖降解受到干扰是其作用模式。针对一系列自溶酶突变体的基于细胞的马沙霉素筛选发现，一种缺乏参与细胞分裂的内-N-乙酰氨基葡萄糖苷酶的Δ菌株对马沙霉素的 MIC 更高。随后的生化和表型分析表明，较高的 MIC 是由于与 LytB 的间接相互作用。对马沙霉素处理细胞的表面变化的进一步分析确定了几种月光蛋白的过表达，包括伸长因子 Tu，其涉及调节细胞形状。使用马沙霉素与其他抗生素协同进行棋盘试验鉴定与细胞壁靶向抗生素磷霉素具有拮抗关系，这进一步支持了细胞壁作用模式。
