Cadasides, Calcium-Dependent Acidic Lipopeptides from the Soil Metagenome That Are Active against Multidrug-Resistant Bacteria

Abstract

The growing threat of antibiotic resistance necessitates the discovery of antibiotics that are active against resistant pathogens. Calcium-dependent antibiotics are a small family of structurally diverse acidic lipopeptides assembled by non-ribosomal peptide synthetases (NRPSs) that are known to display various modes of action against antibiotic-resistant pathogens. Here we use NRPS adenylation (AD) domain sequencing to guide the identification, recovery and cloning of the cde biosynthetic gene cluster from a soil metagenome. Heterologous expression of the cde biosynthetic gene cluster led to the production of cadasides A (1) and B (2), a sub-family of acidic lipopeptides that is distinct from previously characterized calcium-dependent antibiotics in terms of both overall structure and acidic residue rich peptide core. The cadasides inhibit the growth of multidrug-resistant Gram-positive pathogens by disrupting cell wall biosynthesis in the presence of high concentrations of calcium. Interestingly, sequencing of AD domains from diverse soils found that sequences predicted to arise from cadaside-like gene clusters are predominantly found in soils containing high levels of calcium carbonate.

The Desotamide Family of Antibiotics

“…The biosynthetic pathway leading to the production of the phytotoxin coronatine in the phytopathogenic bacteria Pseudomonas syringae , although not solely NRPS dependent, utilises NRPS biosynthetic logic and l - allo -Ile as a precursor to synthesise coronamic acid, an integral component of the phytotoxin [ 33 ]. Recently, cadasides A and B, calcium-dependent acidic lipopeptides whose BGC was characterised in a functional metagenomics study, were discovered [ 34 ]. The BGC encodes 13 NRPS modules as well as six other operons specifying the regulation, biosynthesis and transport of chemical precursors or the final compound.…”

Viennamycins: Lipopeptides Produced by a Streptomyces sp.

“…Finally, the lipophilic moiety of the molecule was identified as (2 Z ,4 E )-8-methylnona-2,4-dienoic acid (MNDA) using COSY and HMBC correlations and comparison of the NMR data to those of cadaside B and ramoplanin A3. 3 , 9 The Z and E configurations of the Δ 2 and Δ 4 double bonds present in this fragment were assigned based on the measured J 2,3 and J 4,5 coupling constant values of 11.3 and 15.0 Hz, respectively. 3 , 9 The attachment of this residue to the α-NH of the peptide was confirmed by HMBC correlations from the NH (δ H 8.19 ppm) and Hα (δ H 4.63 ppm) protons of the Asn residue to carbon C-1 of MNDA (δ C 166.1 ppm).…”

“… 3 , 9 The Z and E configurations of the Δ 2 and Δ 4 double bonds present in this fragment were assigned based on the measured J 2,3 and J 4,5 coupling constant values of 11.3 and 15.0 Hz, respectively. 3 , 9 The attachment of this residue to the α-NH of the peptide was confirmed by HMBC correlations from the NH (δ H 8.19 ppm) and Hα (δ H 4.63 ppm) protons of the Asn residue to carbon C-1 of MNDA (δ C 166.1 ppm). De novo sequencing of the compound using HRMS/MS led to results that were not in full agreement with the amino acid sequence deduced from BGC analysis (see below).…”

“…Recently, a family of novel lipopeptides named cadasides has been discovered via heterologous expression of a biosynthetic gene cluster identified in a soil metagenome library. 3 The cyclic peptide core of cadasides lacks the canonical Ca 2+ -binding motif, and the complete peptide moiety features five aspartate residues, making these compounds more acidic compared to other reported lipopeptide antibiotics. Maximum antibacterial activity of cadasides required a significantly higher concentration of Ca 2+ ions (100 mM), which is unlikely to be relevant for antibacterial therapy, but might reflect the environmental conditions at the spot where the metagenomics DNA was isolated from (high calcium carbonate concentration).…”

Natural Products and Synthetic Biology: Where We Are and Where We Need To Go

“…1b). Circumventing native producers is relevant because it allows us, for example, to obtain compounds from organisms that have yet to be cultured or to obtain compounds from metagenomic data sets (9). In addition, for cultured organisms, it provides a more streamlined way to increase yields and engineer biosynthesis because native producers may not be genetically tractable.…”