Structural Elucidation of Cryptic Algaecides in Marine Algal‐Bacterial Symbioses by NMR Spectroscopy and MicroED

Abstract

Microbial secondary metabolite discovery is often conducted in pure monocultures. In a natural setting, however, where metabolites are constantly exchanged, biosynthetic precursors are likely provided by symbionts or hosts. In the current work, we report eight novel and architecturally unusual secondary metabolites synthesized by the bacterial symbiont Phaeobacter inhibens from precursors that, in a native context, would be provided by their algal hosts. Three of these were produced at low titres and their structures were determined de novo using the emerging microcrystal electron diffraction method. Some of the new metabolites exhibited potent algaecidal activity suggesting that the bacterial symbiont can convert algal precursors, tryptophan and sinapic acid, into complex cytotoxins. Our results have important implications for the parasitic phase of algal‐bacterial symbiotic interactions.

Hakuhybotrol, a polyketide produced byHypomyces pseudocorticiicola, characterized with the assistance of 3D ED/MicroED

“…2,3 3D ED/MicroED is currently attracting attention as a method for structure determination of small molecules. 1,22–26 To the best of our knowledge, this is the first time that the absolute configuration of natural compounds is directly elucidated by 3D ED/MicroED and confirmed by derivatization and SCXRD. Our data suggest that 3D ED/MicroED could be applicable to determine the absolute configuration of natural products by the previously reported method.…”

Electron Diffraction of 3D Molecular Crystals

“…Given these limitations, if attempting to solve a challenging novel structure such as a complex natural product (generally replete with heteroatoms such as O and N) via 3D ED, rigorous corroboration with external data from NMR spectroscopy and mass spectrometry remains essential. 207,212 Second, 3D ED always requires well-formed single crystals. Serendipitously, many compounds may inhabit a specific "Goldilocks zone" where they refuse to form X-ray-scale crystals yet grudgingly aggregate into crystalline assemblies at the nanoscale.…”

“…In a field continually grappling with the myriad pitfalls associated with analysis of complex 2D NMR spectra, the clarity provided by a corroborating crystal structure seems almost cathartic. Furthermore, when applied in tandem with comparative genomics or metabolomics (to mine relevant biosynthetic gene clusters) and synthetic biology (to express those genes in model organisms), 3D ED could also significantly accelerate the rate of NP discovery. − More broadly, 3D ED is rapidly finding a complementary niche within the wider context of synthetic chemistry; a growing number of reports now feature 3D ED structures of relevant synthetic targets or intermediates which proved unsuitable for single-crystal XRD. − These structures include two noncanonical amino acids bearing all-carbon quaternary stereocenters, a trio of organic semiconductors solved at ultrahigh resolution (one of which is depicted in Figure E), a family of electron-deficient expanded helicenes, a pentacyclic indole-derived ester, and a synthetic mimic of the cuboidal subunit in the oxygen-evolving complex of photosystem II …”

“…In these cases, even at the refinement stage, elemental identity can be arduous or impossible to deduce based solely on experimental difference Fourier maps. Given these limitations, if attempting to solve a challenging novel structure such as a complex natural product (generally replete with heteroatoms such as O and N) via 3D ED, rigorous corroboration with external data from NMR spectroscopy and mass spectrometry remains essential. , …”

Ecological Niche-Inspired Genome Mining Leads to the Discovery of Crop-Protecting Nonribosomal Lipopeptides Featuring a Transient Amino Acid Building Block

“…Instead of brute force screenings of microbial monocultures, tapping into the metabolome associated with mutualistic or antagonistic ecological interactions between prokaryotes and eukaryotes is an effective approach to unlock the antibiotic treasure trove of nature. − Especially when in combination with genome mining, structure prediction of natural products from genetic information using bioinformatics tools is greatly facilitated . As a consequence, the biosynthetic potential of symbiotic bacteria can be uncovered in a timely fashion .…”