Activating and Attenuating the Amicoumacin Antibiotics.

Hyun Park,Jason Crawford,Corey Perez,Elena Perry

doi:10.3390/molecules21070824

Abstract

The amicoumacins belong to a class of dihydroisocoumarin natural products and display antibacterial, antifungal, anticancer, and anti-inflammatory activities. Amicoumacins are the pro-drug activation products of a bacterial nonribosomal peptide-polyketide hybrid biosynthetic pathway and have been isolated from Gram-positive Bacillus and Nocardia species. Here, we report the stimulation of a “cryptic” amicoumacin pathway in the entomopathogenic Gram-negative bacterium Xenorhabdus bovienii, a strain not previously known to produce amicoumacins. X. bovienii participates in a multi-lateral symbiosis where it is pathogenic to insects and mutualistic to its Steinernema nematode host. Waxmoth larvae are common prey of the X. bovienii-Steinernema pair. Employing a medium designed to mimic the amino acid content of the waxmoth circulatory fluid led to the detection and characterization of amicoumacins in X. bovienii. The chemical structures of the amicoumacins were supported by 2D-NMR, HR-ESI-QTOF-MS, tandem MS, and polarimeter spectral data. A comparative gene cluster analysis of the identified X. bovienii amicoumacin pathway to that of the Bacillus subtilis amicoumacin pathway and the structurally-related Xenorhabdus nematophila xenocoumacin pathway is presented. The X. bovienii pathway encodes an acetyltransferase not found in the other reported pathways, which leads to a series of N-acetyl-amicoumacins that lack antibacterial activity. N-acetylation of amicoumacin was validated through in vitro protein biochemical studies, and the impact of N-acylation on amicoumacin’s mode of action was examined through ribosomal structural analyses.

Highlights

The amicoumacins have been identified in select Gram-positive Bacillus and Nocardia species and belong to a larger group of bacterial dihydroisocoumarin natural products [1,2]
Organic extractable metabolites from X. bovienii Moldova were compared from cultures grown in Hemolymph-mimetic medium (HMM), Lysogeny Broth (LB), (LB), and and LB
LB supplemented supplemented with high high concentrations concentrations of LL-proline (72.6 mM), a known free amino amino acid nutrient signal that enhances the production of some secondary metabolites in Xenorhabdus

Summary

Introduction

The amicoumacins have been identified in select Gram-positive Bacillus and Nocardia species and belong to a larger group of bacterial dihydroisocoumarin natural products [1,2]. A number of amicoumacin analogs have since been structurally and functionally characterized [5,6,7,8,9] and synthetic routes have been developed to access their core scaffolds [10,11]. These secondary metabolites harbor potent antibacterial activities against clinically-relevant bacterial pathogens, such as Helicobacter pylori and methicillin-resistant. Staphylococcus aureus [12,13] Their antibacterial activity can be attributed to inhibiting the bacterial ribosome, which has been supported by extensive biochemical and X-ray crystallographic studies [14].

Methods

Results

Conclusion