Family Of Natural Products Research Articles

Lipidation Engineering in Daptomycin Biosynthesis.

Lipopeptides are an important family of natural products, some of which are clinically used as antibiotics to treat multidrug-resistant pathogens. Although the lipid moieties play a crucial role in balancing antibacterial activity and hemolytic toxicity, modifying the lipid moieties has been challenging due to the complexity of the lipidation process in lipopeptide biosynthesis. Here, we show that the lipid profile can be altered by engineering both secondary and primary metabolisms, using daptomycin as an example. First, swapping the fatty acyl AMP ligase (FAAL) gene dptF with foreign FAAL homologs improved the fatty acyl specificity of the lipidation process for decanoic acid. Then, the introduction of Mycobacterium type I fatty acid synthase operon (MvFAS-Ib/MvAcpS) and Cryptosporidium thioesterase (CpTEII) enriched the fatty acid pool with decanoic acid in Streptomyces roseosporus. The engineered fatty acid metabolism eliminates the need for external decanoic acid supplementation by enabling S. roseosporus to biosynthesize decanoic acid. By complete engineering of the lipidation process, we achieved, for the first time, high-purity, natural production of daptomycin. The lipidation engineering approach we demonstrate here lays the foundation for the lipidation control in lipopeptide biosynthesis.

A chiral hydrogen atom abstraction catalyst for the enantioselective epimerization of meso-diols.

Hydrogen atom abstraction is an important elementary chemical process but is very difficult to carry out enantioselectively. We have developed catalysts, readily derived from the Cinchona alkaloid family of natural products, which can achieve this by virtue of their chiral amine structure. The catalyst, following single-electron oxidation, desymmetrizes meso-diols by selectively abstracting a hydrogen atom from one carbon center, which then regains a hydrogen atom by abstraction from a thiol. This results in an enantioselective epimerization process, forming the chiral diastereomer with high enantiomeric excess. Cyclic and acyclic 1,2-diols are compatible, as are acyclic 1,3-diols. Additionally, we demonstrate the viability of combining our approach with carbon-carbon bond formation in Giese addition. Given the increasing number of synthetic methods involving hydrogen atom transfer steps, we anticipate that this work will have a broad impact in the field of enantioselective radical chemistry.

A kalihinol analog disrupts apicoplast function and vesicular trafficking in P. falciparum malaria.

We report the discovery of MED6-189, an analog of the kalihinol family of isocyanoterpene natural products that is effective against drug-sensitive and drug-resistant Plasmodium falciparum strains, blocking both asexual replication and sexual differentiation. In vivo studies using a humanized mouse model of malaria confirm strong efficacy of the compound in animals with no apparent hemolytic activity or toxicity. Complementary chemical, molecular, and genomics analyses revealed that MED6-189 targets the parasite apicoplast and acts by inhibiting lipid biogenesis and cellular trafficking. Genetic analyses revealed that a mutation in PfSec13, which encodes a component of the parasite secretory machinery, reduced susceptibility to the drug. Its high potency, excellent therapeutic profile, and distinctive mode of action make MED6-189 an excellent addition to the antimalarial drug pipeline.

Maduraflavacins A-E, Unusual Phenyl Polyene Metabolites Produced by a Rare Marine-Derived Actinomadura sp.

Phenyl polyenes comprise a small family of bacterial natural products with broad and potent bioactivities, primarily found in actinobacteria. Here we report the discovery of five new phenyl polyene metabolites, maduraflavacins A-E (1-5), from a rare, marine-derived actinobacteria strain Actinomadura glauciflava NA03286. The structures of these natural products were determined by NMR spectroscopy, HRESIMS, LC-MS/MS, and chemical derivatization. All of these new maduraflavacins feature methyl substitutions at the polyene side chain, and maduraflavacins A-C (1-3) possessed a 1-N-β-d-glucosamine-(3 → 1)-O-β-d-glucopyranosyl-(3 → 1)-O-β-d-glucopyranosyl-(6 → 1)-O-β-d-glucopyranoside tetrasaccharide moiety via an amido linkage with a phenyl polyene skeleton. Compounds 1 and 2 showed weak antibacterial activities against the Gram-positive bacteria Staphylococcus aureus Sau 16339 and Micrococcus luteus, respectively.

Establishing a coumarin production platform by protein and metabolic engineering

Coumarins are a vast family of natural products with diverse biological activities. Cinnamyl-CoA ortho-hydroxylases (CCHs) catalyze the gateway and rate-limiting step in coumarin biosynthesis. However, engineering CCHs is challenging due to the large size of the substrates and the vague structure-activity relationship. Herein, directed evolution and structure-guided engineering were performed to engineer a CCH (AtF6′H from Arabidopsis thaliana) using a fluorescence-based screening method, yielding the transplantable surface mutations and the substrate-specific pocket mutations with improved activity. Structural analysis and molecular dynamics simulations elucidated the conformational changes that led to increased catalytic efficiency. Applying appropriate variants with the optimized upstream biosynthetic pathways improved the titers of three simple coumarins by 5 to 22-fold. Further introducing glycosylation modules resulted in the production of four coumarin glucosides, among which the titer of aesculin was increased by 15.7-fold and reached 3 g/L in scale-up fermentation. This work unleashed the potential of CCHs and established an Escherichia coli platform for coumarins production.

Anodic Cyclization Reactions and an Approach to the Chrysosporazine Family of Natural Products

Electrochemistry has long offered new opportunities for the construction of biologically relevant molecules. Many of these opportunities are based on the ability that electrochemistry provides for reversing the polarity of known functional groups. For example, consider the brief retrosynthetic analysis shown in the Scheme below. The analysis proposes a route to the Chrysosporazine family of natural products (specifically Chrysosporazine D in this case) from readily available amino acid and lignin derived starting materials. The Chrysosporazines are efflux inhibitors that improve the effectiveness of chemotherapy agents.1 The plan calls for a coupling of the two starting materials to afford a cyclization substrate. However, the proposed cyclization reaction would combine a nucleophilic amide nitrogen with a nucleophile electron rich styrene derivative. In principle, an anodic oxidation reaction can solve this issue by either converting the amide nitrogen into a radical or oxidizing the styrene derivative to form a radical cation. Either mode would effectively reverse the polarity of one of the nucleophilic groups and allow for the cyclization. Previous efforts examining these cyclizations have focused on the generation of a N-radical intermediate.2 Yet while these efforts have been very successful, they require the presence of a phenyl ring on the amide nitrogen in order to lower its pKa. This is essential for the formation of an amide anion that is in turn oxidized to the desired N-radical. Such a plan is not compatible with the synthesis of the Chrysosporazines that simply do not have a phenyl ring on the nitrogen of the amide. Placing one there would prevent the intramolecular cyclization. So, how can this problem be resolved? There are two possible answers to this question. One involves a change in the overall strategy that converts the amide into an electron-rich olefin equivalent that can be oxidized to from a radical cation. The second involves finding an alternative method for dropping the pKa of the amide so that the anion and subsequent N-radical can still be made. In the chemistry to be discussed, both strategies will be examined and the propensity for the cyclic product to undergo over-oxidation shown to be the determining factor for which pathway is preferred; a preference that has led to a new approach to N-based radical cyclizations. 1. Elbanna, A. H., Khalil, Z. G., Bernhardt, P., V., Capon, R. J. Org. Lett. 2019, 21, 8097–8100. 2. Xiong, P.; Xu, H-C. Acc. Chem. Res. 2019, 52, 3339-3350. Figure 1

Systematic Route to Construct the 5-5-6 Tricyclic Core of Furanobutenolide-Derived Cembranoids and Norcembranoids.

Herein, we present a highly efficient method for constructing the intricate 5-5-6 fused ring system commonly found in the polycyclic furanobutenolide-derived cembranoid and norcembranoid natural product family with remarkable diastereoselectivity, utilizing an intramolecular Diels-Alder reaction as the cornerstone. Notably, employing a propargyl ether tether as the dienophile yields significant enhancements in the transformation process compared to its propargyl ester counterpart, as demonstrated in our previous total synthesis of havellockate. This advancement holds promising implications for future investigations, offering a streamlined pathway for rapidly assembling the tricyclic core characteristic of this diverse family of natural products.

A general strategy for the synthesis of taxane diterpenes.

The carbon skeleton of any organic molecule serves as the foundation for its three-dimensional structure, playing a pivotal role in determining its physical and biological properties1. As such, taxane diterpenes are one of the most well-known natural product families, primarily owing to the success of their most prominent compound, paclitaxel, an effective anticancer therapeutic for more than 25 years2-6. In contrast to classical taxanes, the bioactivity of cyclotaxanes (also referred to as complex taxanes) remains significantly underexplored. The carbon skeletons of these two groups of taxanes differ significantly, and so would typically their own distinct synthetic approaches. Here we report a versatile synthetic strategy based on the interconversion of complex molecular frameworks, providing general access to the wider taxane diterpene family. A range of classical and cyclotaxane frameworks was prepared including, among others, the total syntheses of taxinine K (2), canataxapropellane (5) and dipropellane C from a single advanced intermediate. The synthetic approach deliberately eschews biomimicry, emphasizing instead the power of stereoelectronic control in orchestrating the interconversion of polycyclic frameworks.

Cellulamides: A New Family of Marine-Sourced Linear Peptides from the Underexplored Cellulosimicrobium Genus.

Bioprospecting the secondary metabolism of underexplored Actinomycetota taxa is a prolific route to uncover novel chemistry. In this work, we report the isolation, structure elucidation, and bioactivity screening of cellulamides A and B (1 and 2), two novel linear peptides obtained from the culture of the macroalga-associated Cellulosimicrobium funkei CT-R177. The host of this microorganism, the Chlorophyta Codium tomentosum, was collected in the northern Portuguese coast and, in the scope of a bioprospecting study focused on its associated actinobacterial community, strain CT-R177 was isolated, taxonomically identified, and screened for the production of antimicrobial and anticancer compounds. Dereplication of a crude extract of this strain using LC-HRMS(/MS) analysis unveiled a putative novel natural product, cellulamide A (1), that was isolated following mass spectrometry-guided fractionation. An additional analog, cellulamide B (2) was obtained during the chromatographic process and chemically characterized. The chemical structures of the novel linear peptides, including their absolute configurations, were elucidated using a combination of HRMS, 1D/2D NMR spectroscopy, and Marfey's analysis. Cellulamide A (1) was subjected to a set of bioactivity screenings, but no significant biological activity was observed. The cellulamides represent the first family of natural products reported from the Actinomycetota genus Cellulosimicrobium, showcasing not only the potential of less-explored taxa but also of host-associated marine strains for novel chemistry discovery.

Investigation Into Novel Mukanadin B, Mukanadin D and Mukanadin F Derivatives as Antagonists of 5-HT1A Signalling.

Marine bromopyrrole alkaloids are a diverse family of natural products with a large array of biological applications. The mukanadin family is a group of molecules consisting of seven members (mukanadin A-G) that possess a range of biological activities. Inhibition of serotonergic signaling has been demonstrated by mukanadin B derivatives, presenting this chemical scaffold as a candidate for further SAR exploration. A library of thirteen novel mukanadin B and D derivatives with structural variation targeted at the pyrrole ring, central linker and hydantoin ring, were synthesized. These analogues were subsequently assessed for serotonergic antagonism, in addition to natural products, mukanadin B, D, F and 9-hydroxy mukanadin B. A collection of compounds exhibited significant 5-HT1A signaling, including five of the novel derivatives and two of the naturally occurring bromopyrroles, mukanadin B and F. Particular SAR information could be determined from these results, such as modification of the pyrrole ring being a well-tolerated strategy for improving serotonergic inhibition. Other changes to the pharmacophore led to significant reduction in activity such as saturation of the linker region, or no conclusive improvement in inhibitory activity such as a 9-OH group or replacement of the hydantoin ring with a triazole moiety.

Identification of a family of species-selective complex I inhibitors as potential anthelmintics

Soil-transmitted helminths (STHs) are major pathogens infecting over a billion people. There are few classes of anthelmintics and there is an urgent need for new drugs. Many STHs use an unusual form of anaerobic metabolism to survive the hypoxic conditions of the host gut. This requires rhodoquinone (RQ), a quinone electron carrier. RQ is not made or used by vertebrate hosts making it an excellent therapeutic target. Here we screen 480 structural families of natural products to find compounds that kill Caenorhabditis elegans specifically when they require RQ-dependent metabolism. We identify several classes of compounds including a family of species-selective inhibitors of mitochondrial respiratory complex I. These identified complex I inhibitors have a benzimidazole core and we determine key structural requirements for activity by screening 1,280 related compounds. Finally, we show several of these compounds kill adult STHs. We suggest these species-selective complex I inhibitors are potential anthelmintics.

Genome-Driven Discovery of Hygrocins in Streptomyces rapamycinicus.

Ansamycins, represented by the antituberculosis drug rifamycin, are an important family of natural products. To obtain new ansamycins, Streptomyces rapamycinicus IMET 43975 harboring an ansamycin biosynthetic gene cluster was fermented in a 50 L scale, and subsequent purification work led to the isolation of five known and four new analogues, where hygrocin W (2) belongs to benzoquinonoid ansamycins, and the other three hygrocins, hygrocins X-Z (6-8), are new seco-hygrocins. The structures of ansamycins (1-8) were determined by the analysis of spectroscopic (1D/2D NMR and ECD) and MS spectrometric data. The Baeyer-Villiger enzyme which catalyzed the ester formation in the ansa-ring was confirmed through in vivo CRISPR base editing. The discovery of these compounds further enriches the structural diversity of ansamycins.

A Cytochrome P450 Catalyzes Oxidative Coupling Formation of Insecticidal Dimeric Indole Piperazine Alkaloids.

Cytochrome P450 (CYP450)-catalyzed oxidative coupling is an efficient strategy for using simple building blocks to construct complex structural scaffolds of natural products. Among them, heterodimeric coupling between two different monomers is relatively scarce, and the corresponding CYP450s are largely undiscovered. In this study, we discovered a fungal CYP450 (CpsD) and its associated cps cluster from 37208 CYP450s of Pfam PF00067 family member database and subsequently identified a group of new skeleton indole piperazine alkaloids (campesines A-G) by combination of genome mining and heterologous synthesis. Importantly, CYP450 CpsD mainly catalyzes intermolecular oxidative heterocoupling of two different indole piperazine monomers to generate an unexpected 6/5/6/6/6/6/5/6 eight-ring scaffold through the formation of one C-C bond and two C-N bonds, illuminating its first dimerase role in this family of natural products. The proposed catalytic mechanism of CpsD was deeply investigated by diversified substrate derivatization. Moreover, dimeric campesine G shows good insecticidal activity against the global honeybee pest Galleria mellonella. Our study shows a representative example of discovering new skeleton monomeric and dimeric indole piperazine alkaloids from microbial resources, expands our knowledge of bond formation by CYP450s and supports further development of the newly discovered and engineered campesine family compounds as potential biopesticides.

A Cytochrome P450 Catalyzes Oxidative Coupling Formation of Insecticidal Dimeric Indole Piperazine Alkaloids

AbstractCytochrome P450 (CYP450)‐catalyzed oxidative coupling is an efficient strategy for using simple building blocks to construct complex structural scaffolds of natural products. Among them, heterodimeric coupling between two different monomers is relatively scarce, and the corresponding CYP450s are largely undiscovered. In this study, we discovered a fungal CYP450 (CpsD) and its associated cps cluster from 37208 CYP450s of Pfam PF00067 family member database and subsequently identified a group of new skeleton indole piperazine alkaloids (campesines A−G) by combination of genome mining and heterologous synthesis. Importantly, CYP450 CpsD mainly catalyzes intermolecular oxidative heterocoupling of two different indole piperazine monomers to generate an unexpected 6/5/6/6/6/6/5/6 eight‐ring scaffold through the formation of one C−C bond and two C−N bonds, illuminating its first dimerase role in this family of natural products. The proposed catalytic mechanism of CpsD was deeply investigated by diversified substrate derivatization. Moreover, dimeric campesine G shows good insecticidal activity against the global honeybee pest Galleria mellonella. Our study shows a representative example of discovering new skeleton monomeric and dimeric indole piperazine alkaloids from microbial resources, expands our knowledge of bond formation by CYP450s and supports further development of the newly discovered and engineered campesine family compounds as potential biopesticides.

Pepticinnamins N, O, and P, Nonribosomal Peptides from the Soil-Derived Streptomyces mirabilis P8-A2.

Cinnamoyl moiety containing nonribosomal peptides represented by pepticinnamin E are a growing family of natural products isolated from different Streptomyces species and possess diverse bioactivities. The soil bacterium Streptomyces mirabilis P8-A2 harbors a cryptic pepticinnamin biosynthetic gene cluster, producing azodyrecins as major products. Inactivation of the azodyrecin biosynthetic gene cluster by CRISPR-BEST base editing led to the activation and production of pepticinnamin E (1) and its analogues, pepticinnamins N, O, and P (2-4), the structures of which were determined by detailed NMR spectroscopy, HRMS data, and Marfey's reactions. These new compounds did not show a growth inhibitory effect against the LNCaP and C4-2B prostate cancer lines, respectively.

Recent Advances in Grayanane Diterpenes: Isolation, Structural Diversity, and Bioactivities from Ericaceae Family (2018-2024).

Diterpenes represent one of the most diverse and structurally complex families of natural products. Among the myriad of diterpenoids, grayanane diterpenes are particularly notable. These terpenes are characterized by their unique 5/7/6/5 tetracyclic system and are exclusive to the Ericaceae family of plants. Renowned for their complex structures and broad spectrum of bioactivities, grayanane diterpenes have become a primary focus in extensive phytochemical and pharmacological research. Recent studies, spanning from 2018 to January 2024, have reported a series of new grayanane diterpenes with unprecedented carbon skeletons. These compounds exhibit various biological properties, including analgesic, antifeedant, anti-inflammatory, and inhibition of protein tyrosine phosphatase 1B (PTP1B). This paper delves into the discovery of 193 newly identified grayanoids, representing 15 distinct carbon skeletons within the Ericaceae family. The study of grayanane diterpenes is not only a deep dive into the complexities of natural product chemistry but also an investigation into potential therapeutic applications. Their unique structures and diverse biological actions make them promising candidates for drug discovery and medicinal applications. The review encompasses their occurrence, distribution, structural features, and biological activities, providing invaluable insights for future pharmacological explorations and research.

Genome mining of labdane-related diterpenoids: Discovery of the two-enzyme pathway leading to (-)-sandaracopimaradiene in the fungus Arthrinium sacchari.

Labdane-related diterpenoids (LRDs) in fungi are a pharmaceutically important, but underexplored family of natural products. In the biosynthesis of fungal LRDs, bifunctional terpene cyclases (TCs) consisting of αβγ domains are generally used to synthesize the polycyclic skeletones of LRDs. Herein, we conducted genome mining of LRDs in our fungal genome database and identified a unique pair of TCs, AsPS and AsCPS, in the fungus Arthrinium sacchari. AsPS consists of catalytically active α and inactive β domains, whereas AsCPS contains βγ domains and a truncated α domain. Heterologous expression in Aspergillus oryzae and biochemical characterization of recombinant proteins demonstrated that AsCPS synthesized copalyl diphosphate and that AsPS then converted it to (-)-sandaracopimaradiene. Since AsPS and AsCPS have distinct domain organizations from those of known fungal TCs and are likely generated through fusion or loss of catalytic domains, our findings provide insight into the evolution of TCs in fungi.

Non-modular fatty acid synthases yield distinct N-terminal acylation in ribosomal peptides.

Recent efforts in genome mining of ribosomally synthesized and post-translationally modified peptides (RiPPs) have expanded the diversity of post-translational modification chemistries. However, RiPPs are rarely reported as hybrid molecules incorporating biosynthetic machinery from other natural product families. Here we report lipoavitides, a class of RiPP/fatty-acid hybrid lipopeptides that display a unique, putatively membrane-targeting 4-hydroxy-2,4-dimethylpentanoyl (HMP)-modified N terminus. The HMP is formed via condensation of isobutyryl-coenzyme A (isobutyryl-CoA) and methylmalonyl-CoA catalysed by a 3-ketoacyl-(acyl carrier protein) synthase III enzyme, followed by successive tailoring reactions in the fatty acid biosynthetic pathway. The HMP and RiPP substructures are then connected by an acyltransferase exhibiting promiscuous activity towards the fatty acyl and RiPP substrates. Overall, the discovery of lipoavitides contributes a prototype of RiPP/fatty-acid hybrids and provides possible enzymatic tools for lipopeptide bioengineering.

Full biosynthetic pathway of pyrrolobenzoxazines

Pyrrolobenzoxazines are a rare terpene-amino acid family of natural products with potent biological activities. Here, we reported the full biosynthetic pathway of paeciloxazine (1), a typical pyrrolobenzoxazine, with significant insecticidal activity. Base on heterologous expression, chemical complement experiment, and in vitro biochemical assays, we demonstrated the sesquiterpene portion of 1 derived from discontinuously oxidations of amorphdiene, in which P450 monooxygenase PaxH catalyzed a cascade of hydroxylation and epoxidation, while two flavin dependent monooxygenases are involved in the transformation of the esterified tryptophan into a pyrrolobenzoxazine core. Furthermore, a total of 15 compounds were generated through heterologous expression, of which 13, 17 and 20 showed potential antiepileptic activity. This study fully elucidated the biosynthetic pathway of paeciloxazine (1) and showed the diversity and complexity of constructing natural products by organisms.

Decylprodigiosin: a new member of the prodigiosin family isolated from a seaweed-associated Streptomyces.

Bioprospecting actinobacterial secondary metabolism from untapped marine sources may lead to the discovery of biotechnologically-relevant compounds. While studying the diversity and bioactive potential of Actinomycetota associated with Codium tomentosum, a green seaweed collected in the northern Portuguese cost, strain CT-F61, identified as Streptomyces violaceoruber, was isolated. Its extracts displayed a strong anticancer activity on breast carcinoma T-47D and colorectal carcinoma HCT116 cells, being effective as well against a panel of human and fish pathogenic bacteria. Following a bioactivity-guided isolation pipeline, a new analogue of the red-pigmented family of the antibiotics prodigiosins, decylprodigiosin (1), was identified and chemically characterized. Despite this family of natural products being well-known for a long time, we report a new analogue and the first evidence for prodigiosins being produced by a seaweed-associated actinomycete.