Characterization of a Cryptic NRPS Gene Cluster in Bacillus velezensis FZB42 Reveals a Discrete Oxidase Involved in Multithiazole Biosynthesis

Polythiazole-containing
nonribosomal peptides (NRPs) represent
a rare and important group of natural products with intriguing structures
and promising bioactivities. However, the biosynthetic mechanism of
polythiazole in NRPS pathways remains poorly understood. Genome mining
of the model biocontrol bacterium Bacillus velezensis FZB42 revealed that a cryptic NRPS gene cluster nrs might be involved in the synthesis of polyheterocycle family products.
Heterologous expression of this gene cluster led to the identification
of rare trithiazole-containing products, bacillothiazols A–N
(1–14), with potent inhibitory activity
against protein tyrosine phosphatase PTP1B. Unlike the general incis manner in which the oxidation of polythiazole in NRPs
is canonically catalyzed by integrated oxidase (Ox) domains, our gene
deletion and biochemical characterization revealed that a stand-alone
oxidase NrsB in trans catalyzed the iterative oxidation
of multithiazoline to multithiazole in bacillothiazol biosynthesis,
which implied an unprecedented pathway of polythiazole formation in
microbial NRPs. Notably, rather than the integrated NRPS Ox domains,
the catalytic residues in NrsB were seemingly the same as ribosomally
synthesized and post-translationally modified peptide (RiPP) dehydrogenases,
which can catalyze iterative oxidation reactions, suggesting that
NrsB might represent an interesting case in the evolution of integrated
NRPS Ox domains and RiPP dehydrogenases. These findings not only resolve
the long-standing puzzle of the natural products from the model strain B. velezensis FZB42 but also provide insight into
the versatility of polythiazole biosynthesis.