Structural expansion and assembly line promiscuity in the biosynthesis of actinochelins, a widespread family of siderophores from Actinobacteria Javier Santos-Aberturas, Rodney Lacret, Alicia H. Russell, Daniel Heine, Eleni Vikeli, Barrie Wilkinson and Andrew W. Truman John Innes Centre, UK We report the discovery of actinochelins, a new group of non-ribosomal peptide siderophores produced under specific stress culture conditions by Actinomadura atramentaria DSM43919 and Saccharopolyspora sp. KY21. Actinochelins feature an acylated pseudotripeptide backbone (2,3-dihydroxybenzoic acid/threonine/N- hydroxyornithine) akin to the rhodochelins 1 (previously described in Rhodococcus jostii HA1), but they differ in the rich structural diversification of the N-hydroxyornithine moiety and in their alternative tailoring. Genome analysis led to the identification of putative actinochelin biosynthetic gene clusters (BGCs) both in A. atramentaria and Sacchopolyspora sp. KY21, showing a progressive clustering compared to the three subclusters reported as required for the biosynthesis of rhodochelins. CRISPR/Cas9 genomic editing in Sacchopolyspora sp. KY21 and heterologous gene complementation experiments were performed to confirm the function of the BGC. Supragenomic analysis indicated the presence of similar BGCs in multiple species belonging to Actinomadura and Saccharopolyspora . Actinochelins are essential for the growth of Sacchopolyspora sp. KY21 under iron depletion conditions, but strikingly the gene deletion of the standalone adenylation domain responsible for the incorporation of 2,3-dihydroxybenzoic acid to the assembly line resulted in a mutant strain still able to grow under iron depletion. Untargeted metabolomics revealed that this mutant produces low iron-affinity actinochelin congeners to support its growth, thus suggesting an unusual assembly line promiscuity phenomenon combined with pathway crosstalk that ensures the redundancy of an essential ecological function. References 1. Bosello M, Robbel L, Linne U, Xie X, Marahiel MA. Biosynthesis of the siderophore rhodochelin requires the coordinated expression of three independent gene clusters in Rhodococcus jostii RHA1. J Am Chem Soc. 2011 Mar 30;133(12):4587-95. doi: 10.1021/ja1109453.
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