A Plug-and-Play T7 Expression System for Heterologous Production of Lanthipeptides in <i>Bacillus subtilis</i>

Ribosomally synthesized lanthionine-containing peptides (lanthipeptides) have emerged as a promising source of antimicrobials against multidrug resistance pathogens. An effective way to discover and engineer lanthipeptides is through heterologous expression of their biosynthetic gene clusters (BGCs) in a host of choice. Here we report a plug-and-play pathway refactoring strategy for rapid evaluation of lanthipeptide BGCs in <i>Bacillus subtilis</i> based on the T7 expression system. As a proof of concept, we used this strategy to not only observe the successful production of a known lanthipeptide haloduracin β but also discover two new human-microbiota-derived lanthipeptides that previously failed to be produced in <i>Escherichia coli</i>. The resulting <i>B. subtilis</i> plug-and-play T7 expression system should enable the genome mining of new lanthipeptides in a high-throughput manner.