Nonribosomal peptide antibiotics, including polymyxin, vancomycin, and teixobactin, most of which contain d-amino acids, are highly effective against multidrug-resistant bacteria. However, overusing antibiotics while ignoring the risk of resistance arising has inexorably led to widespread emergence of resistant bacteria. Therefore, elucidation of the emerging mechanisms of resistance to nonribosomal peptide antibiotics is critical to their implementation. Here we describe a networking-associated genome-mining platform for linking biosynthetic building blocks to resistance components associated with biosynthetic gene clusters. By applying this approach to 5,585 complete bacterial genomes spanning the entire domain of bacteria, with subsequent chemical and enzymatic analyses, we demonstrate a mechanism of resistance toward nonribosomal peptide antibiotics that is based on hydrolytic cleavage by d-stereospecific peptidases. Our finding reveals both the widespread distribution and broad-spectrum resistance potential of d-stereospecific peptidases, providing a potential early indicator of antibiotic resistance to nonribosomal peptide antibiotics.

非核糖体肽抗生素，包括多粘菌素，万古霉素，和teixobactin，其中大部分含有d-氨基酸对多药耐药细菌非常有效。然而，过度使用抗生素而忽视产生耐药性的风险不可避免地导致了耐药菌的广泛出现。因此，阐明对非核糖体肽抗生素耐药的新兴机制对其实施至关重要。在这里，我们描述了一个与网络相关的基因组挖掘平台，用于将生物合成构件与生物合成基因簇相关的抗性成分联系起来。通过将这种方法应用于横跨细菌整个域的5,585个完整细菌基因组，并进行后续的化学和酶促分析，我们证明了对非核糖体肽抗生素具有抗性的机制是基于d的水解切割-立体特异性肽酶。我们的发现揭示了d-立体特异性肽酶的广泛分布和广谱耐药性，为非核糖体肽抗生素的耐药性提供了潜在的早期指标。