An official website of the United States government
Summary Plant‐specialized metabolism is complex, with frequent examples of highly branched biosynthetic pathways, and shared chemical intermediates. As such, many plant‐specialized metabolic networks are poorly characterized.TheN‐methyl Δ1‐pyrrolinium cation is a simple pyrrolidine alkaloid and precursor of pharmacologically important tropane alkaloids. Silencing of pyrrolidine ketide synthase (AbPyKS) in the roots ofAtropa belladonna(Deadly Nightshade) reduces tropane alkaloid abundance and causes highN‐methyl Δ1‐pyrrolinium cation accumulation. The consequences of this metabolic shift on alkaloid metabolism are unknown. In this study, we utilized discovery metabolomics coupled withAbPyKSsilencing to reveal major changes in the root alkaloid metabolome ofA. belladonna.We discovered and annotated almost 40 pyrrolidine alkaloids that increase whenAbPyKSactivity is reduced. Suppression of phenyllactate biosynthesis, combined with metabolic engineeringin planta, and chemical synthesis indicates several of these pyrrolidines share a core structure formed through the nonenzymatic Mannich‐like decarboxylative condensation of theN‐methyl Δ1‐pyrrolinium cation with 2‐O‐malonylphenyllactate. Decoration of this core scaffold through hydroxylation and glycosylation leads to mono‐ and dipyrrolidine alkaloid diversity.This study reveals the previously unknown complexity of theA. belladonnaroot metabolome and creates a foundation for future investigation into the biosynthesis, function, and potential utility of these novel alkaloids.
more »
« less
Synthesis of (±)‐Setigerumine I: Biosynthetic Origins of the Elusive Racemic Papaveraceae Isoxazolidine Alkaloids**
Abstract The biosynthetic origins of the structurally related racemic isoxazolidinePapaveraceaealkaloids Setigerumine I, Dactylicapnosinine and Dactylicapnosine have remained elusive since their original isolation over two decades ago. Herein we report the first biosynthetic hypothesis for their formation and, inspired by it, the first synthesis of (±)‐Setigerumine I with accompanying computational rationale. Based on the results, these isoxazolidine alkaloids arise from racemizing oxidative rearrangements of prominent isoquinoline alkaloids Noscapine and Hydrastine. The key steps featured in this synthesis are a room temperature Cope elimination and a domino oxidation/inverse‐electron demand 1,3‐dipolar cycloaddition of an axially chiral, yet configurationally unstable, intermediate. The work opens this previously inaccessible family of natural products for biological studies.
more »
« less
- Award ID(s):
- 2102462
- PAR ID:
- 10302661
- Publisher / Repository:
- Wiley Blackwell (John Wiley & Sons)
- Date Published:
- Journal Name:
- Angewandte Chemie International Edition
- Volume:
- 60
- Issue:
- 52
- ISSN:
- 1433-7851
- Page Range / eLocation ID:
- p. 27236-27240
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
Abstract Monoterpene indole alkaloids (MIAs) are a class of natural products comprised of thousands of structurally unique bioactive compounds with significant therapeutic values. Due to difficulties associated with isolation from native plant species and organic synthesis of these structurally complex molecules, microbial production of MIAs using engineered hosts are highly desired. In this work, we report the engineering of fully integrated Saccharomyces cerevisiae strains that allow de novo access to strictosidine, the universal precursor to thousands of MIAs at 30–40 mg/L. The optimization efforts were based on a previously reported yeast strain that is engineered to produce high titers of the monoterpene precursor geraniol through compartmentalization of mevalonate pathway in the mitochondria. Our approaches here included the use of CRISPR-dCas9 interference to identify mitochondria diphosphate transporters that negatively impact the titer of the monoterpene, followed by genetic inactivation; the overexpression of transcriptional regulators that increase cellular respiration and mitochondria biogenesis. Strain construction included the strategic integration of genes encoding both MIA biosynthetic and accessory enzymes into the genome under a variety of constitutive and inducible promoters. Following successful de novo production of strictosidine, complex alkaloids belonging to heteroyohimbine and corynantheine families were reconstituted in the host with introduction of additional downstream enzymes. We demonstrate that the serpentine/alstonine pair can be produced at ∼5 mg/L titer, while corynantheidine, the precursor to mitragynine can be produced at ∼1 mg/L titer. Feeding of halogenated tryptamine led to the biosynthesis of analogs of alkaloids in both families. Collectively, our yeast strain represents an excellent starting point to further engineer biosynthetic bottlenecks in this pathway and to access additional MIAs and analogs through microbial fermentation. One Sentence SummaryAn Saccharomyces cerevisiae-based microbial platform was developed for the biosynthesis of monoterpene indole alkaloids, including the universal precursor strictosidine and further modified heteroyohimbine and corynantheidine alkaloids.more » « less
-
Traxler, Matthew F. (Ed.)ABSTRACT Marine sponge holobionts are prolific sources of natural products. One of the most geographically widespread classes of sponge-derived natural products is the bromotyrosine alkaloids. A distinguishing feature of bromotyrosine alkaloids is that they are present in phylogenetically disparate sponges. In this study, using sponge specimens collected from Guam, the Solomon Islands, the Florida Keys, and Puerto Rico, we queried whether the presence of bromotyrosine alkaloids potentiates metabolomic and microbiome conservation among geographically distant and phylogenetically different marine sponges. A multi-omic characterization of sponge holobionts revealed vastly different metabolomic and microbiome architectures among different bromotyrosine alkaloid-harboring sponges. However, we find statistically significant correlations between the microbiomes and metabolomes, signifying that the microbiome plays an important role in shaping the overall metabolome, even in low-microbial-abundance sponges. Molecules mined from the polar metabolomes of these sponges revealed conservation of biosynthetic logic between bromotyrosine alkaloids and brominated pyrrole-imidazole alkaloids, another class of marine sponge-derived natural products. In light of prior findings postulating the sponge host itself to be the biosynthetic source of bromotyrosine alkaloids, our data now set the stage for investigating the causal relationships that dictate the microbiome-metabolome interconnectedness for marine sponges in which the microbiome may not contribute to natural product biogenesis. IMPORTANCE Our work demonstrates that phylogenetically and geographically distant sponges with very different microbiomes can harbor natural product chemical classes that are united in their core chemical structures and biosynthetic logic. Furthermore, we show that independent of geographical dispersion, natural product chemistry, and microbial abundance, overall sponge metabolomes tightly correlate with their microbiomes.more » « less
-
Dynemicin is an enediyne natural product fromMicromonospora chersinaATCC53710. Access to the biosynthetic gene cluster of dynemicin has enabled thein vitrostudy of gene products within the cluster to decipher their roles in assembling this unique molecule. This paper reports the crystal structure of DynF, the gene product of one of the genes within the biosynthetic gene cluster of dynemicin. DynF is revealed to be a dimeric eight-stranded β-barrel structure with palmitic acid bound within a cavity. The presence of palmitic acid suggests that DynF may be involved in binding the precursor polyene heptaene, which is central to the synthesis of the ten-membered ring of the enediyne core.more » « less
