Univ. Grenoble Alpes, CNRS, CERMAV, 38000, Grenoble, France.
Chemistry. 2023 Jan 27;29(6):e202202991. doi: 10.1002/chem.202202991. Epub 2022 Dec 5.
Soluble fragments of peptidoglycan called muropeptides are released from the cell wall of bacteria as part of their metabolism or as a result of biological stresses. These compounds trigger immune responses in mammals and plants. In bacteria, they play a major role in the induction of antibiotic resistance. The development of efficient methods to produce muropeptides is, therefore, desirable both to address their mechanism of action and to design new antibacterial and immunostimulant agents. Herein, we engineered the peptidoglycan recycling pathway of Escherichia coli to produce N-acetyl-β-D-glucosaminyl-(1→4)-1,6-anhydro-N-acetyl-β-D-muramic acid (GlcNAc-anhMurNAc), a common precursor of Gram-negative and Gram-positive muropeptides. Inactivation of the hexosaminidase nagZ gene allowed the efficient production of this key disaccharide, providing access to Gram-positive muropeptides through subsequent chemical peptide conjugation. E. coli strains deficient in both NagZ hexosaminidase and amidase activities further enabled the in vivo production of Gram-negative muropeptides containing meso-diaminopimelic acid, a rarely available amino acid.
可溶的肽聚糖片段，称为肽聚糖，作为其代谢的一部分或由于生物压力从细菌细胞壁中释放出来。这些化合物在哺乳动物和植物中引发免疫反应。在细菌中，它们在诱导抗生素耐药性方面起着重要作用。因此，开发有效的方法来生产肽聚糖是可取的，既可以解决它们的作用机制，又可以设计新的抗菌和免疫刺激剂。在这里，我们对大肠杆菌的肽聚糖回收途径进行了工程改造，以生产 N-乙酰-β-D-葡糖胺基-(1→4)-1,6-脱水-N-乙酰-β-D-乳酰氨酸（GlcNAc-anhMurNAc），这是革兰氏阴性和革兰氏阳性肽聚糖的常见前体。nagZ 基因的失活允许这种关键二糖的有效生产，通过随后的化学肽键合获得革兰氏阳性肽聚糖。同时缺乏 NagZ 六糖胺酶和酰胺酶活性的大肠杆菌菌株进一步使含有内消旋二氨基庚二酸的革兰氏阴性肽聚糖的体内生产成为可能，内消旋二氨基庚二酸是一种很少见的氨基酸。
