Mining the Biosynthetic Landscape of Lactic Acid Bacteria Unearths a New Family of RiPPs Assembled by a Novel Type of ThiF-like Adenylyltransferases

Ribosomally synthesized
and post-translationally modified peptides
(RiPPs) are chemically diverse natural products of ribosomal origin.
These peptides, which frequently act as signals or antimicrobials,
are biosynthesized by conserved enzymatic machinery, making genome
mining a powerful strategy for unearthing previously uncharacterized
members of their class. Herein, we investigate the untapped biosynthetic
potential of Lactobacillales (i.e., lactic acid bacteria), an order
of Gram-positive bacteria closely associated with human life, including
pathogenic species and industrially relevant fermenters of dairy products.
Through genome mining methods, we systematically explored the distribution
and diversity of ThiF-like adenylyltransferase-utilizing RiPP systems
in lactic acid bacteria and identified a number of unprecedented biosynthetic
gene clusters. In one of these clusters, we found a previously undescribed
group of macrocyclic imide biosynthetic pathways containing multiple
transporters that may be involved in a potential quorum sensing (QS)
system. Through in vitro assays, we determined that
one such adenylyltransferase specifically catalyzes the intracyclization
of its precursor peptide through macrocyclic imide formation. Incubating
the enzyme with various primary amines revealed that it could effectively
amidate the C-terminus of the precursor peptide. This new transformation
adds to the growing list of Nature’s peptide macrocyclization
strategies and expands the impressive catalytic repertoire of the
adenylyltransferase family. The diverse RiPP systems identified herein
represent a vast, unexploited landscape for the discovery of a novel
class of natural products and QS systems.