Bacterial peptidoglycan hydrolases play an essential role in cell wall metabolism during bacterial growth, division, and elongation (autolysins) or in the elimination of closely related species from the same ecological niche (bacteriocins). Most studies concerning the peptidoglycan hydrolases present in Gram-positive bacteria have focused on clinically relevant Staphylococcus aureus or the model organism Bacillus subtilis, while knowledge relating to other species remains limited. Here, we report two new peptidoglycan hydrolases from the M23 family of metallopeptidases derived from the same staphylococcal species, Staphylococcus pettenkoferi. They share modular architecture, significant sequence identity (60%), catalytic and binding residue conservation, and similar modes of activation, but differ in gene distribution, putative biological role, and, strikingly, in their isoelectric points (pIs). One of the peptides has a high pI, similar to that reported for all M23 peptidases evaluated to date, whereas the other displays a low pI, a unique feature among M23 peptidases. Consequently, we named them SpM23_B (Staphylococcus pettenkoferi M23 “Basic”) and SpM23_A (Staphylococcus pettenkoferi M23 “Acidic”). Using genetic and biochemical approaches, we have characterized these two novel lytic enzymes, both in vitro and in their physiological context. Our study presents a detailed characterization of two novel and clearly distinct peptidoglycan hydrolases to understand their role in bacterial physiology.

细菌肽聚糖水解酶在细菌生长、分裂和伸长（自溶素）或从同一生态位（细菌素）中消除密切相关物种的过程中，在细胞壁代谢中起重要作用。大多数关于革兰氏阳性菌中存在的肽聚糖水解酶的研究都集中在临床相关的金黄色葡萄球菌 或模式生物 枯草芽孢杆菌，而与其他物种有关的知识仍然有限。在这里，我们报告了来自相同葡萄球菌物种的 M23 金属肽酶家族的两种新肽聚糖水解酶，小金黄色葡萄球菌. 它们具有模块化结构、显着的序列同一性 (60%)、催化和结合残基保守性以及类似的激活模式，但在基因分布、推定的生物学作用以及引人注目的等电点 (pI) 方面有所不同。其中一种肽具有高 pI，类似于迄今为止评估的所有 M23 肽酶所报告的 pI，而另一种则显示出低 pI，这是 M23 肽酶中的独特特征。因此，我们将它们命名为 SpM23_B（Staphylococcus pettenkoferi M23“Basic”）和 SpM23_A（Staphylococcus pettenkoferi M23“Acidic”）。使用遗传和生化方法，我们对这两种新型裂解酶进行了表征，体外并在他们的生理环境中。我们的研究详细描述了两种新型且明显不同的肽聚糖水解酶，以了解它们在细菌生理学中的作用。