Discovery and Biosynthesis of Pepticinnamins G–M Featuring Three Enzymes-Catalyzed Nonproteinogenic Amino Acid Formation
Yuanjie Ge, Guiyang Wang, Jing Jin, Tan Liu, Xueyang Ma, Zhongyi Zhang, Tongtong Geng, Juan Song, Xiaojie Ma, Yingtao Zhang, Donghui Yang, Ming Ma
Abstract
Since pepticinnamin E was discovered almost 30 years ago, no other pepticinnamin family of natural products has been reported to date. Here, we report the discovery of pepticinnamins G–I (1–3) from a marine Streptomyces sp. PKU-MA01144 and pepticinnamins J–M (4–7) from several mutants, and these new compounds contain different N-methyl-l-alanine and l-tyrosine residues compared to pepticinnamin E. Genome sequencing, heterologous expression, gene deletion, and reconstitution of enzymatic reaction in vitro identified the biosynthetic gene cluster of 1–7 and first experimentally established the biosynthesis of the nonproteinogenic 2-chloro-3-hydroxy-4-methoxy-l-phenylalanine residue by a biopterin-dependent hydroxylase Pep10, an O-methyltransferase Pep9, and a flavin-dependent halogenase Pep1. The biosynthetic research and heterologous expression system in this study set the stage for pathway engineering for more pepticinnamins generation in the future.