Isolation, Structure Elucidation, and Biosynthesis of an Unusual Hydroxamic Acid Ester-Containing Siderophore from Actinosynnema mirum

Abstract

In this study we report the isolation, structure elucidation, and biosynthesis of mirubactin (1), a siderophore containing an unprecedented chemical functionality in natural products, namely, an O-acyl hydroxamic acid ester. Mirubactin represents the first siderophore isolated from the genus Actinosynnema and the first natural product produced by Actinosynnema mirum whose biosynthetic gene cluster could be identified. Structure elucidation was accomplished through a combination of spectroscopic (NMR, IR, and UV/vis) and mass spectrometric methods and revealed the presence of an unusual ester bond between the δ-N-hydroxyl group of δ-N-formyl-δ-N-hydroxyornithine and a 2,3-dihydroxybenzoate moiety. Bioinformatic analysis of the A. mirum genome and subsequent biochemical characterization of the putative biosynthetic machinery identified the gene cluster responsible for mirubactin assembly. The proposed biosynthesis of mirubactin comprises the iterative use of a stand-alone carrier-protein-bound substrate, as well as an ester-bond-forming step catalyzed by a C-terminal condensation domain, thus revealing an interesting system for further biochemical studies to gain a deeper understanding of nonribosomal peptide synthetase-catalyzed siderophore biosynthesis.

Chlorocatechelins A and B from Streptomyces sp.: New Siderophores Containing Chlorinated Catecholate Groups and an Acylguanidine Structure

“…The molecular formula of chlorocatechelin B (2) was established as C 18 H and 13 C NMR spectra that were simpler than those of 1. The planar structure of 2 was determined by 2D NMR analysis (Figure 2b) to be composed of one unit each of CDB, Orn, and hfOrn.…”

Identification of a New Siderophore Acinetoamonabactin Produced by a Salt‐Tolerant Bacterium Acinetobacter Soli

“…1 H NMR spectrum of the siderophore (Figure S2 and Table S2) is quite clean and simple, showing three signals of equal intensities in the downfield region ( δ 7.05‐7.25 ppm). The positions and splitting patterns, observed in the aromatic region, are indicative of the presence of 2,3‐dihydroxybenzoyl moiety, a common building block found frequently in catechol type siderophores ,. A singlet at δ 3.16 ppm has also appeared in the 1 H NMR of it (Figure S2).…”

“…Fragment ion peak, at m/z 134.90, is probably appearing due to the formation of dehydrated 2,3‐DHB. Literature has the precedence that ortho‐hydroxylated benzoic acid derivative transforms to benzodioxetane during gas phase fragmentation . Another fragment ion peak at m/z 117.08, corresponds to the existence of tris(aminomethyl)methylamine residue (Figure ).…”

Comparative Metabolomic Analysis of an Alternative Biosynthetic Pathway to Pseudosugars in Actinosynnema mirum DSM 43827

“…None of these bacteria, however, has been reported to produce pseudosugar‐containing natural products. Among them is Actinosynnema mirum DSM 43827, a soil bacterium that is known to produce the β‐lactam antibiotic nocardicins2 and the unique siderophore antibiotic mirubactin 3. Whole‐genome analysis of A. mirum by the antiSMASH (antibiotics and Secondary Metabolite Analysis Shell) program4 resulted in the prediction of 25 cryptic biosynthetic gene clusters for secondary metabolites; interestingly, the PsGT cluster was not one of them.…”