Department of Chemistry , University of Warwick , Coventry CV4 7AL , United Kingdom.
ACS Chem Biol. 2019 May 17;14(5):845-849. doi: 10.1021/acschembio.9b00095. Epub 2019 Apr 8.
The terminal step in the biosynthesis of nonribosomal peptides is the hydrolytic release and, frequently, macrocyclization of an aminoacyl-S-thioester by an embedded thioesterase. The surugamide biosynthetic pathway is composed of two nonribosomal peptide synthetase (NRPS) assembly lines in which one produces surugamide A, which is a cyclic octapeptide, and the other produces surugamide F, a linear decapeptide. The terminal module of each system lacks an embedded thioesterase, which led us to question how the peptides are released from the assembly line (and cyclized in the case of surugamide A). We characterized a cyclase belonging to the β-lactamase superfamily in vivo, established that it is a trans-acting release factor for both compounds, and verified this functionality in vitro with a thioester mimic of linear surugamide A. Using bioinformatics, we estimate that ∼11% of filamentous Actinobacteria harbor an NRPS system lacking an embedded thioesterase and instead employ a trans-acting cyclase. This study improves the paradigmatic understanding of how nonribosomal peptides are released from the terminal peptidyl carrier protein and adds a new dimension to the synthetic biology toolkit.
非核糖体肽的生物合成的末端步骤是通过嵌入的硫酯酶将氨酰-S-硫酯的水解释放，并且通常是大环化。苏拉菌生物合成途径由两条非核糖体肽合酶（NRPS）组装线组成，其中一条产生苏拉菌 A，这是一种环状八肽，另一条产生苏拉菌 F，这是一种线性十肽。每个系统的末端模块都缺乏嵌入的硫酯酶，这导致我们质疑肽是如何从装配线上释放出来的（在苏拉菌 A 的情况下被环化）。我们在体内鉴定了属于β-内酰胺酶超家族的环化酶，确定它是两种化合物的反式作用释放因子，并通过线性苏拉菌 A 的硫酯模拟物在体外验证了这种功能。通过生物信息学，我们估计大约 11%的丝状放线菌含有缺乏嵌入硫酯酶的 NRPS 系统，而是采用反式作用的环化酶。这项研究提高了对非核糖体肽如何从末端肽酰载体蛋白释放的范例理解，并为合成生物学工具包增添了新的维度。
