Bacteria resist to the turgor pressure of the cytoplasm through a net-like macromolecule, the peptidoglycan, made of glycan strands connected via peptides cross-linked by penicillin-binding proteins (PBPs). We recently reported the emergence of β-lactam resistance resulting from a bypass of PBPs by the YcbB L,D-transpeptidase (LdtD), which form chemically distinct 3→3 cross-links compared to 4→3 formed by PBPs. Here we show that peptidoglycan expansion requires controlled hydrolysis of cross-links and identify among eight endopeptidase paralogues the minimum enzyme complements essential for bacterial growth with 4→3 (MepM) and 3→3 (MepM and MepK) cross-links. Purified Mep endopeptidases unexpectedly displayed a 4→3 and 3→3 dual specificity implying recognition of a common motif in the two cross-link types. Uncoupling of the polymerization of glycan chains from the 4→3 cross-linking reaction was found to facilitate the bypass of PBPs by YcbB. These results illustrate the plasticity of the peptidoglycan polymerization machinery in response to the selective pressure of β-lactams.

中文翻译：

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内肽酶在大肠杆菌替代交联酶介导的肽聚糖合成中的作用

细菌通过网状大分子肽聚糖抵抗细胞质的膨胀压力，肽聚糖由通过青霉素结合蛋白 (PBP) 交联的肽连接的聚糖链构成。我们最近报道了由 YcbB L,D-转肽酶 (LdtD) 绕过 PBP 引起的 β-内酰胺耐药性的出现，与 PBP 形成的 4→3 相比，它形成化学上不同的 3→3 交联。在这里，我们表明肽聚糖扩增需要交联的受控水解，并在八种内肽酶旁系同源物中鉴定出具有 4→3（MepM）和 3→3（MepM 和 MepK）交联的细菌生长所必需的最小酶补体。纯化的 Mep 内肽酶出人意料地表现出 4→3 和 3→3 双重特异性，这意味着识别两种交联类型中的共同基序。发现从 4→3 交联反应中解偶联聚糖链的聚合有助于 YcbB 绕过 PBP。这些结果说明了肽聚糖聚合机制响应 β-内酰胺的选择压力的可塑性。