Genome-Mining Based Discovery of Pyrrolomycin K and L from the Termite-Associated <i>Micromonospora</i> sp. RB23

Natural products derived from symbiotic microbes remain a rich source of structurally diverse and bioactive molecules. In this study, we report <i>de novo</i> genome sequencing of the termite-associated isolate <i>Micromonospora</i> sp. RB23. Genome mining uncovered a type I polyketide synthase (T1PKS) biosynthetic gene cluster encoding five halogenases, predicted to produce pyrrolomycin-like antimicrobial compounds. Mass-spectrometry-based molecular networking facilitated the identification and isolation of <i>N</i>-methylated pyrrolomycin K and mycothiol-adduct, pyrrolomycin L. Structure elucidation was accomplished based on liquid chromatography high-resolution tandem mass spectrometry (LC-HRMS/MS) alongside 1D and 2D nuclear magnetic resonance (NMR) spectroscopy. Based on the evaluated of antimicrobial activity, we propose that <i>N</i>-methylation and mycothiol-based conjugation in pyrrolomycins are possible detoxification mechanisms that play a role in enhancing self-tolerance.