Non-ribosomal peptide synthetases (NRPSs) are multienzymes that produce complex natural metabolites with many applications in medicine and agriculture. They are composed of numerous catalytic domains that elongate and chemically modify amino acid substrates or derivatives and of non-catalytic carrier protein domains that can tether and shuttle the growing products to the different catalytic domains. The intrinsic flexibility of NRPSs permits conformational rearrangements that are required to allow interactions between catalytic and carrier protein domains. Their large size coupled to this flexibility renders these multi-domain proteins very challenging for structural characterization. Here, we summarize recent studies that offer structural views of multi-domain NRPSs in various catalytically relevant conformations, thus providing an increased comprehension of their catalytic cycle. A better structural understanding of these multienzymes provides novel perspectives for their re-engineering to synthesize new bioactive metabolites.

非核糖体肽合成酶 (NRPS) 是一种多酶，可产生复杂的天然代谢物，在医学和农业中有许多应用。它们由许多催化结构域组成，这些催化结构域可以延长和化学修饰氨基酸底物或衍生物，也由非催化载体蛋白结构域组成，这些非催化载体蛋白结构域可以将生长的产物束缚和穿梭到不同的催化结构域。NRPS 的内在灵活性允许催化和载体蛋白结构域之间相互作用所需的构象重排。它们的大尺寸加上这种灵活性使得这些多域蛋白质对结构表征非常具有挑战性。在这里，我们总结了最近的研究，这些研究提供了各种催化相关构象的多域 NRPS 的结构视图，从而提高对它们的催化循环的理解。更好地理解这些多酶的结构为它们重新设计以合成新的生物活性代谢物提供了新的视角。