An Obligate Peptidyl Brominase Underlies the Discovery of Highly Distributed Biosynthetic Gene Clusters in Marine Sponge Microbiomes

Abstract

Marine sponges are prolific sources of bioactive natural products, several of which are produced by bacteria symbiotically associated with the sponge host. Bacteria-derived natural products, and the specialized bacterial symbionts that synthesize them, are not shared among phylogenetically distant sponge hosts. This is in contrast to nonsymbiotic culturable bacteria in which the conservation of natural products and natural product biosynthetic gene clusters (BGCs) is well established. Here, we demonstrate the widespread conservation of a BGC encoding a cryptic ribosomally synthesized and post-translationally modified peptide (RiPP) in microbiomes of phylogenetically and geographically dispersed sponges from the Pacific and Atlantic oceans. Detection of this BGC was enabled by mining for halogenating enzymes in sponge metagenomes, which, in turn, allowed for the description of a broad-spectrum regiospecific peptidyl tryptophan-6-brominase which possessed no chlorination activity. In addition, we demonstrate the cyclodehydrative installation of azoline heterocycles in proteusin RiPPs. This is the first demonstration of halogenation and cyclodehydration for proteusin RiPPs and the enzymes catalyzing these transformations were found to competently interact with other previously described proteusin substrate peptides. Within a sponge microbiome, many different generalized bacterial taxa harbored this BGC with often more than 50 copies of the BGC detected in individual sponge metagenomes. Moreover, the BGC was found in all sponges queried that possess high diversity microbiomes but it was not detected in other marine invertebrate microbiomes. These data shed light on conservation of cryptic natural product biosynthetic potential in marine sponges that was not detected by traditional natural product-to-BGC (meta)genome mining.

Comparative metagenomic analysis of biosynthetic diversity across sponge microbiomes highlights metabolic novelty, conservation and diversification

“…The potential for azol(in)e-containing RiPPs has been found widespread in bacterial genomes (92), but has only recently been reported in marine sponge microbiomes with the identification of the srp clusters by Nguyen et . al (15). We predict an additional structurally diversified set of azol(in)e-containing RiPPs to this repertoire, expanding the combinatorial space of modifying enzymes and hence the potential for production of bioactive NPs.…”

“…Even though sponges are recognized as extensive sources of diverse NPs, their inventory of ribosomally synthesized and post-translationally modified peptides (RiPP) has remained largely undescribed, with one exception being the proteusin polytheonamides (15,23,80). Here we expand the known repertoire in sponges and showcase sponges’ RiPP diversity by identifying 17 uncharacterized RiPP families which seem to be widespread in sponge holobionts.…”

“…Natural Products (NPs) are ubiquitous small molecules that play a key role in symbiosis, as mediators of interactions within the holobiont (15,16). In this generally mutualistic relationship, the microbes provide their host with chemical compounds to prevent predation, fouling and infection, while receiving primary metabolic nutrition and a hospitable habitat (17)(18)(19)(20).…”

Trikoramides B–D, Bioactive Cyanobactins from the Marine Cyanobacterium Symploca hydnoides

“…Incidentally, a recent paper by Nguyen and co-workers reported the presence of broad-spectrum regiospecific peptidyl tryptophan-6-brominase on RiPP substrate. This brominase enzyme was detected through the mining for halogenating enzymes in sponge metagenomes [16]. To our knowledge, trikoramides C (2) and D (3) are the first hydroxylamine-and brominated cyclotryptophan-containing cyanobactins, respectively, from S. hydnoides.…”

Beyond Soil-Dwelling Actinobacteria: Fantastic Antibiotics and Where to Find Them

“…For example, endosymbiotic cyanobacteria have been identified as the producers of the highly toxic polybrominated diphenyl ethers found in sponges of the family Dysideidae [ 40 ]. The enormous amount of information generated by metagenomic sequencing of these complex microbial communities constitutes excellent database material for genome mining, as recently demonstrated by the discovery of a widespread family of brominated RiPPs [ 41 ]. This implies that the metabolic potential of these symbiotic communities could also include numerous cryptic compounds.…”