Integration of Genomic Data with NMR Analysis Enables Assignment of the Full Stereostructure of Neaumycin B, a Potent Inhibitor of Glioblastoma from a Marine-Derived Micromonospora

Abstract

The microbial metabolites known as the macrolides are some of the most successful natural products used to treat infectious and immune diseases. Describing the structures of these complex metabolites, however, is often extremely difficult due to the presence of multiple stereogenic centers inherent in this class of polyketide-derived metabolites. With the availability of genome sequence data and a better understanding of the molecular genetics of natural product biosynthesis, it is now possible to use bioinformatic approaches in tandem with spectroscopic tools to assign the full stereostructures of these complex metabolites. In our quest to discover and develop new agents for the treatment of cancer, we observed the production of a highly cytotoxic macrolide, neaumycin B, by a marine-derived actinomycete bacterium of the genus Micromonospora. Neaumycin B is a complex polycyclic macrolide possessing 19 asymmetric centers, usually requiring selective degradation, crystallization, derivatization, X-ray diffraction analysis, synthesis, or other time-consuming approaches to assign the complete stereostructure. As an alternative approach, we sequenced the genome of the producing strain and identified the neaumycin gene cluster ( neu). By integrating the known stereospecificities of biosynthetic enzymes with comprehensive NMR analysis, the full stereostructure of neaumycin B was confidently assigned. This approach exemplifies how mining gene cluster information while integrating NMR-based structure data can achieve rapid, efficient, and accurate stereostructural assignments for complex macrolides.

Secondary Metabolites from Marine Micromonospora: Chemistry and Bioactivities

“…The biological assay uncovered that 10 showed significant potency and selectivity toward human glioblastoma (U87) cell lines with LD 50 of 6.4×10 –6 μ m , which indicated 10 to be a lead structure for glioblastoma treatment. The results also illustrated the practicability of applying bioinformatic approaches to assign the full stereostructure of complex macrolides …”

The genus Micromonospora as a model microorganism for bioactive natural product discovery

“…This unique metabolite revealed potent and selective in vitro inhibitory activity against glioblastoma cell line indicating it as a promising drug candidate. 50 All anticancer Micromonospora-derived macrolides are depicted in Fig. 6.…”

“…The very rst of this group was discovered in 1995 when the lipopeptides rakicidins A and B (139,140) were discovered from a soil-derived Micromonospora strain. Both lipo-tripeptides are cyclized through a b-hydroxy fatty acid and exhibited potent in vitro antiproliferative activity against M109 cell line (IC 50 55, 230 ng mL À1 ). Rakicidin B (140) differs from A (139) by one methylene group in the lipid side chain, indicating that the length of the lipid side chain may signicantly affect the cytotoxic activity of this class of compounds.…”

“…Macrolides derived from Micromonospora with anticancer activity(46)(47)(48)(49)(50)(51)(52)(53), and other miscellaneous macrolides (compounds 54-62).20946 | RSC Adv., 2020, 10, 20939-20959…”

Stereocontrolled Synthesis as an Enabling Tool for the Configurational Assignment of Marine Polyketide Natural Products

“…These aforementioned contemporary structural elucidation tools do not provide all the solutions and ab initio computational methods, including DP4, become prohibitively costly to run on large and especially flexible molecules; the nascent cryo‐EM technology for small molecules relies on the sample being a solid and available in reasonable quantities often not feasible after an isolation campaign; and the use of bioinformatics as a prospective tool, requires the characterisation of the gene sequence encoding the biosynthetic machinery, itself a formidable task, especially when the actual producing organism may be highly elusive . Even then, these techniques are seldom able to fully rationalise the complete stereochemistry of a natural product.…”