Alternative Benzoxazole Assembly Discovered in Anaerobic Bacteria Provides Access to Privileged Heterocyclic Scaffold

Abstract

Benzoxazole scaffolds feature prominently in diverse synthetic and natural product-derived pharmaceuticals. Our understanding of their bacterial biosynthesis is, however, limited to ortho-substituted heterocycles from actinomycetes. We report an overlooked biosynthetic pathway in anaerobic bacteria (typified in Clostridium cavendishii) that expands the benzoxazole chemical space to meta-substituted heterocycles and heralds a distribution beyond Actinobacteria. The first benzoxazoles from the anaerobic realm (closoxazole A and B) were elucidated by NMR and chemical synthesis. By genome editing in the native producer, heterologous expression in Escherichia coli, and systematic pathway dissection we show that closoxazole biosynthesis invokes an unprecedented precursor usage (3-amino-4-hydroxybenzoate) and manner of assembly. Synthetic utility was demonstrated by the precursor-directed biosynthesis of a tafamidis analogue. A bioinformatic survey reveals the pervasiveness of related gene clusters in diverse bacterial phyla.

Genomics-Driven Discovery of Benzoxazole Alkaloids from the Marine-Derived Micromonospora sp. SCSIO 07395

“…Benzoxazoles represent a family of heterocyclic compounds containing a benzene-fused oxazole ring core structure, which are frequently found in pharmaceutically active compounds [ 2 ], and natural products, for example caboxamycin [ 3 ], calcimycin [ 4 ], A33853 [ 5 ], nataxazole [ 6 ], and nocarbenzoxazoles [ 7 ] from actinomycetes, pseudopteroxazole [ 8 ] and nakijinol [ 9 ] from invertebrate, and salvianans [ 10 , 11 ] from plant. The unique structure and remarkable pharmaceutical potential of benzoxazoles have attracted significant attention for biosynthesis, including the construction of building blocks, the manner of backbone formation and subsequent heterocyclization and the mechanism of tailoring modifications ( Figure S1 ) [ 12 , 13 , 14 , 15 , 16 , 17 , 18 ].…”

Myxococcus xanthus as Host for the Production of Benzoxazoles

“…Benzoxazole‐containing compounds are used in different therapeutic areas, as exemplified by the approved drugs chlorzoxazone (muscle relaxant), tafamidis (transthyretin stabilizer) or suvorexant (treatment of insomnia) [9] . Their potent biological activities combined with the understanding of benzoxazole biosynthesis have stimulated the biotechnological production of benzoxazole analogs [6–8,10–12] . Recently, the genes involved in nataxazole biosynthesis were expressed in the bacterium Escherichia coli and, upon feeding of 3‐HAA together with other aryl carboxylic acids, various benzoxazoles were generated, albeit in low titers [12] …”

“…[9] Their potent biological activities combined with the understanding of benzoxazole biosynthesis have stimulated the biotechnological production of benzoxazole analogs. [6][7][8][10][11][12] Recently, the genes involved in nataxazole biosynthesis were expressed in the bacterium Escherichia coli and, upon feeding of 3-HAA together with other aryl carboxylic acids, various benzoxazoles were generated, albeit in low titers. [12] In this study, we describe an alternative heterologous production system for benzoxazoles.…”

“… [6] Natural products such as the antibiotics caboxamycin and A‐33853 or the anticancer agent nataxazole are synthesized in this way (Figure 1B). [6,7] A distinct assembly strategy is pursued in the biosynthesis of the closoxazoles, which were recently identified from the anaerobic bacterium Clostridium cavendishii DSM21758 [8] . The closoxazole pathway represents the first example of a benzoxazole biosynthetic pathway that utilizes a 3,4‐disubstituted aryl carboxylic acid as a building block and, hence, generates meta ‐substituted benzoxazoles.…”

Hot off the Press

“…A novel family of benzoxazoles with a meta -substituted scaffold, including closoxazole A 31 have been isolated from the anaerobic bacterium Clostridium cavendishii . 26 Genome editing and heterologous expression in Escherichia coli revealed the biosynthetic pathway and the unusual precursor, 3-amino-4-hydroxybenzoate.…”