RESUMEN
Highly functionalized spirobisnaphthalenes, preussomerins N (1) and O (2), and simpler compounds, such as 2,3-α-epoxypalmarumycin CP18 (3), 3α-hydroxy-CJ-12,372 (4), and 16 known structurally related congeners, were isolated from a culture broth of Roussoella sp. KT4147. Structural analysis revealed that 1 was a dimer of preussomerin G (6), connected by a nitrogen atom, and 2 was a derivative of 6 with a macommelin substructure. Preussomerin N (1) was considered to be biosynthetically derived via the Michael-type 1,4-addition of ammonia to 6, followed by another Michael addition to another molecule of 6. Contrarily, 2 was suggested to be derived through an endo-Diels-Alder cycloaddition between a diene derived from the (E)-enol form of macommelinal via an ene-reaction and dienophile 6. Compounds 1 and 2 exhibited potent cytotoxicity against COLO-201 human colorectal cancer cells.
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Naftalenos , Compuestos de Espiro , Humanos , Estructura Molecular , Compuestos de Espiro/química , Compuestos de Espiro/aislamiento & purificación , Compuestos de Espiro/farmacología , Naftalenos/química , Naftalenos/farmacología , Naftalenos/aislamiento & purificación , Ensayos de Selección de Medicamentos Antitumorales , Antineoplásicos/farmacología , Antineoplásicos/química , Antineoplásicos/aislamiento & purificación , Reacción de Cicloadición , Línea Celular TumoralRESUMEN
In recent years, synthetic opioids have emerged as a predominant cause of drug-overdose-related fatalities, causing the "opioid crisis." To design safer therapeutic agents, we accidentally discovered µ-opioid receptor (MOR) antagonists based on fentanyl with a relatively uncomplicated chemical composition that potentiates structural modifications. Here, we showed the development of novel atropisomeric fentanyl analogues that exhibit more potent antagonistic activity against MOR than naloxone, a morphinan MOR antagonist. Derivatives displaying stable axial chirality were synthesized based on the amide structure of fentanyl. The aS- and aR-enantiomers exerted antagonistic and agonistic effects on the MOR, respectively, and each atropisomer interacted with the MOR by assuming a distinct binding mode through molecular docking. These findings suggest that introducing atropisomerism into fentanyl may serve as a key feature in the molecular design of future MOR antagonists to help mitigate the opioid crisis.
Asunto(s)
Fentanilo , Receptores Opioides mu , Receptores Opioides mu/antagonistas & inhibidores , Receptores Opioides mu/metabolismo , Fentanilo/farmacología , Fentanilo/análogos & derivados , Fentanilo/química , Estereoisomerismo , Humanos , Simulación del Acoplamiento Molecular , Relación Estructura-Actividad , Animales , Antagonistas de Narcóticos/química , Antagonistas de Narcóticos/farmacología , Conformación Molecular , Analgésicos Opioides/farmacología , Analgésicos Opioides/química , Analgésicos Opioides/síntesis química , Células CHO , CricetulusRESUMEN
This study focused on synthesizing the tricyclic hexasubstituted spirocyclopropane-core framework 2 of cyclohelminthol X (1), an antifungal cytotoxin isolated from Helminthosporium velutinum yone96 in a stereoselective manner. The synthesis features an SN2-type cyclopropanation of the quaternary chloride 23 generated via a retro-Michael-type ring-opening reaction of an 8-azatricyclo[4.3.0.12,5]deca-3,7,9-trione derivative 22. The successful synthesis confirmed the structure of 1, resolving the ambiguity from the absence of X-ray crystallographic analysis. The prepared models exhibited potent cytotoxicity.
RESUMEN
The relative configuration of the epoxide functionality in pinofuranoxin A (1), α-alkylidene-ß-hydroxy-γ-methyl-γ-butyrolactone with trans-epoxy side chain isolated by Evidente et al. in 2021, was revised by DFT-based spectral reinvestigations and stereo-controlled synthesis. The present investigation demonstrates the difficulty of the configurational elucidation of the stereogenic centers on the conformationally flexible acyclic side-chains. Sharpless's enantioselective epoxidations and dihydroxylations were quite effective in the reinvestigations of the configurations. As our syntheses made all diastereomers available, these would be quite effective in the next structure-biological activity relationship studies.
Asunto(s)
4-Butirolactona , Estereoisomerismo , Estructura Molecular , 4-Butirolactona/química , 4-Butirolactona/análogos & derivados , 4-Butirolactona/síntesis química , Relación Estructura-Actividad , Conformación MolecularRESUMEN
Paraphaeoketones A-C (1-3) were isolated from the culture broth of Paraphaeosphaeria sp. KT4192. Their structures and relative configurations were determined using spectroscopic analysis and verified through density functional theory (DFT)-based chemical shift calculations. The absolute configurations of these compounds were determined by comparing the experimental electronic circular dichroism (ECD) spectra with those based on DFT calculations. We also propose a plausible biosynthetic route to 1-3. While our prior studies on the isolation and structural elucidation of paraphaeolactones (e.g., 4) led us to suggest a Favorskii rearrangement for their biosynthesis, the isolation of 2 prompted the proposal of an alternative biosynthesis for 4, featuring a benzilic acid rearrangement of 2. Moreover, an in vitro conversion of 2 into 4 was achieved successfully, suggesting that a biosynthetic pathway for paraphaeolactones involving a benzilic acid rearrangement is more plausible than the previously presumed Favorskii rearrangement pathway. Arguments based on DFT calculations for these pathways are also described.
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Ascomicetos , Cetonas , Ascomicetos/química , Ascomicetos/metabolismo , Lactonas/química , Lactonas/metabolismo , Estructura Molecular , Cetonas/química , Cetonas/metabolismoRESUMEN
Oryzamutaic acids, possessing a nitrogen-containing heterocyclic skeleton, have been isolated and identified from a rice mutant. Although oryzamutaic acids are expected to be functional ingredients, their functionality is difficult to evaluate, because of their wide variety and presence in trace amounts. Furthermore, how oryzamutaic acid is synthesized in vivo is unclear. Therefore, we developed a simple enzymatic synthesis method for these compounds in vitro. We focused on L-lysine ε-dehydrogenase (LysDH) from Agrobacterium tumefaciens, which synthesizes α-aminoadipate-δ-semialdehyde-a precursor of oryzamutaic acids. LysDH was cloned and expressed in Escherichia coli. Analysis of activity revealed that LysDH catalyzed the synthesis of oryzamutaic acid H at neutral pH in vitro. We synthesized 1.6â¯mg oryzamutaic acid H from 100â¯mgâ¯L-lysine. The synthesized oryzamutaic acid H exhibited UVA absorption, stability of temperature, and stability at a wide pH range. To our knowledge, this study is the first to report the enzymatic synthesis of oryzamutaic acid H in vitro and provides a basis for understanding the mechanisms of oryzamutaic acid synthesis in vivo.
Asunto(s)
Agrobacterium tumefaciens , Aminoácido Oxidorreductasas , Agrobacterium tumefaciens/genética , Lisina , ÁcidosRESUMEN
Dihydroobionin B (1), a chiral congener of known obionin B, was isolated from Pseudocoleophoma sp. KT4119, a freshwater fungus collected from a submerged wood block in Kochi Prefecture, Japan, in 2020. The planar structure of 1 was characterized by mass and NMR spectral analysis and confirmed by density functional theory (DFT)-based chemical shift calculations. Its absolute structure was determined by electronic circular dichroism spectral analysis. Notably, 1 exhibited an extraordinarily large specific rotation [[α]20D +1080 (c 0.056, CHCl3)], which was verified by DFT-based specific rotation calculations. However, these calculations indicated that the sign of the specific rotation based on static analysis was insufficient to determine the absolute configuration in this case. Furthermore, Pseudocoleophoma KT4119 produced coleophomapyrones A (2) and B (3) and coleophomaldehyde A (4). While this is the first report of 2 isolated from a natural source, it has also been prepared previously using a synthetic approach. Compound 1 potently inhibited HIV type 1 integrase (IC50 = 0.44 µM) without significant cytotoxicity. Finally, docking experiments were conducted to propose a plausible mechanism for the behavior of 1.
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VIH-1 , Rotación , Hongos , Inhibidores de Integrasa , Japón , Estructura Molecular , Dicroismo CircularRESUMEN
In general, mushroom-forming fungi secrete liquid on the surface of mycelia just before fruiting-body formation. However, no researchers in mushroom science have paid attention to the liquid until now. We formulated a hypothesis that the liquid plays an important role(s) in the formation of the fruiting body and produces various bioactive compounds and named it the "fruiting liquid (FL)". Four novel compounds (1-4) were isolated from FL of Hypholoma lateritium and Hericium erinaceus. The structures of 1-4 except for their stereochemistry were determined by interpretation of MS and NMR data. The absolute configurations of compounds 1-4 were determined by quantum chemical calculation of the ECD spectrum, by single-crystal X-ray diffraction analyses, or by chemical syntheses. Compounds 1, 3, and 4 induced fruiting body formation of Flammulina velutipes. Compound 4 inhibited the activity of hypoxia-inducible factor, and compounds 2-4 suppressed receptor tyrosine kinase (Axl) expression.
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Agaricales , Ascomicetos , Flammulina , Cristalografía por Rayos X , FrutasRESUMEN
Paraphaeolactones A1, A2, B1, and B2 (1-4, respectively), known arthropsadiol D (5), massariphenone (6) and its positional isomer 7, and massarilactones E (8) and G (9) were isolated from the culture broth of Paraphaeosphaeria sp. KT4192. Although the structural resemblance between 1 and 2 implies that these comprised a diastereomeric pair at the C-2 stereogenic center, electronic circular dichroism (ECD) spectral analyses revealed that they were pseudo-enantiomers possessing the common (2R)-configuration. Paraphaeolactones B1 and B2 (3 and 4) were the derivatives of 2, which equipped the 3-(1-hydroxy-2-oxopropyl)-4-methylcatechol moiety via an acetal bond at C-10. The relative configurations of their acetal carbons were elucidated by NOE experiments, and those of C-8' were deduced independently by ECD spectral analysis. The present study disclosed that 1-5, 8, and 9 contain a methylcyclohexene substructure with the same absolute configuration. This prompted us to reinvestigate the absolute configurations of known structurally related fungal metabolites, allowing us to conclude that the methylcyclohexene moieties of these natural products have the same absolute configuration despite the variety of configurations of other stereogenic centers. The plausible biosynthetic routes for 1-9 are discussed on the basis of the above conclusion. We propose a Favorskii rearrangement as the key transformation for biosyntheses of 1-4.
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Acetales , Ascomicetos , Lactonas , Dicroismo Circular , Conformación Molecular , Estructura Molecular , Estereoisomerismo , Lactonas/químicaRESUMEN
Polysulfide plays an essential role in controlling various physiological activities in almost all organisms. We recently investigated the impact of polysulfide metabolic enzymes on the temporal dynamics of cellular polysulfide speciation and transcriptional regulation by the polysulfide-responsive transcription factor SqrR in Rhodobacter capsulatus. However, how the polysulfidation of thiol groups in SqrR is reduced remains unclear. In the present study, we examined the reduction of polysulfidated thiol residues by the thioredoxin system. TrxC interacted with SqrR in vitro and reduced the polysulfide crosslink between two cysteine residues in SqrR. Furthermore, we found that exogenous sulfide-induced SqrR de-repression during longer culture times is maintained upon disruption of the trxC gene. These results establish a novel signaling pathway in SqrR-mediated polysulfide-induced transcription, by which thioredoxin-2 restores SqrR to a transcriptionally repressed state via the reduction of polysulfidated thiol residues.
RESUMEN
Five integrasone derivatives, integrasone C (1), isointegrasone C (2), integrasone D1 (3), integrasone D2 (4), and integrasone E (5), were isolated from the culture broth of Lepteutypa sp. KT4162. Neither conventional NMR analyses nor DFT (density functional theory)-based computationally assisted chemical shift discussions were sufficient to elucidate the relative configuration of the 1,4-epoxydiol moiety. A combined analysis using the calculated nJCH values and HMBC spectra was helpful to establish the relative configuration. The absolute configurations of 1-5 were determined using DFT-based ECD (electronic circular dichroism) spectral analysis. Biological assays of these compounds revealed that 2 potently inhibits HIV-1 integrase without cytotoxicity.
Asunto(s)
Compuestos Bicíclicos Heterocíclicos con Puentes , Furanos , Estereoisomerismo , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Furanos/farmacología , Espectroscopía de Resonancia Magnética , Dicroismo Circular , Estructura MolecularRESUMEN
Sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) is a membrane protein on the endoplasmic reticulum (ER) that transports Ca2+ from the cytosol into the ER. As its function is associated with various biological phenomena, SERCA has been recognized as a promising druggable target. Here, we report the second-strongest SERCA-inhibitory compound known to date, which we isolated from the marine cyanobacterium Leptochromothrix valpauliae and named iezoside (1). The structure of iezoside (1) is fundamentally different from that of any other SERCA inhibitor, and its potency is the strongest among marine natural products (Ki 7.1 nM). In this article, we report our comprehensive analysis of iezoside (1), which covers its isolation, structural characterization supported by density functional theory (DFT) calculations and statistical analysis, total synthesis, and clarification of the mode of action of its potent antiproliferative activity (IC50 6.7 ± 0.4 nM against HeLa cells).
Asunto(s)
Calcio , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico , Calcio/metabolismo , Citosol/metabolismo , Retículo Endoplásmico/metabolismo , Células HeLa , Humanos , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico/metabolismoRESUMEN
Highly reducing polyketide synthases (HR-PKSs) produce structurally diverse polyketides (PKs). The PK diversity is constructed by a variety of factors, including the ß-keto processing, chain length, methylation pattern, and relative and absolute configurations of the substituents. We examined the stereochemical course of the PK processing for the synthesis of polyhydroxy PKs such as phialotides, phomenoic acid, and ACR-toxin. Heterologous expression of a HR-PKS gene, a trans-acting enoylreductase gene, and a truncated non-ribosomal peptide synthetase gene resulted in the formation of a linear PK with multiple stereogenic centers. The absolute configurations of the stereogenic centers were determined by chemical degradation followed by comparison of the degradation products with synthetic standards. A stereochemical rule was proposed to explain the absolute configurations of other reduced PKs and highlights an error in the absolute configurations of a reported structure. The present work demonstrates that focused functional analysis of functionally related HR-PKSs leads to a better understanding of the stereochemical course.
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Proteínas Fúngicas/química , Sintasas Poliquetidas/química , Policétidos/síntesis química , Ascomicetos/enzimología , Proteínas Fúngicas/genética , Mutación , Oxidación-Reducción , Sintasas Poliquetidas/genética , EstereoisomerismoRESUMEN
A series of cyclohumulanoids, i.e., tricocerapicanols A-C (1a-1c), tricoprotoilludenes A (2a) and B (3), tricosterpurol (4), and tricoilludins A-C (5-7) were isolated along with known violascensol (2b) and omphadiol (8) from the culture broth of Daedaleopsis tricolor, an inedible but not toxic mushroom. The structures were fully elucidated on the basis of NMR spectroscopic analysis, and the suggested relative structures were confirmed via density functional theory (DFT)-based chemical shift calculations involving a DP4 probability analysis. In the present study, the 1H chemical shifts were more informative than the 13C chemical shifts to distinguish the diastereomers at C-11. The absolute configurations of 1-5 were determined by comparing the experimental and calculated electronic circular dichroism (ECD) spectra. For 6 and 7, the same chirality was assigned according to their biosynthetic similarities with the other compounds. The successful assignment of some Cotton effects was achieved by utilizing DFT calculations using simple model compounds. The plausible biosynthesis of 1-7 was also discussed on the basis of the structural commonality and general cyclohumulanoid biosynthesis. Compounds 2a and 5 were found to simultaneously induce hyphal swelling and branching at 5.0 µg/mL against a test fungus Cochliobolus miyabeanus.
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Medios de Cultivo/química , Polyporaceae/crecimiento & desarrollo , Sesquiterpenos , Sesquiterpenos/química , Sesquiterpenos/aislamiento & purificación , Sesquiterpenos/farmacologíaRESUMEN
A potent antifungal fusicoccane dehydroxypericonicin A (1) was isolated from Roussoella sp. along with known allantofuranone (2). The relative structure of 1 was fully elucidated by NMR spectroscopic manner. The suggested relative structure was confirmed by density functional theory (DFT)-based 13C NMR chemical shift calculations. The absolute configuration was investigated by a spectral comparison of the experimental electronic circular dichroism spectrum with that based on the DFT calculations.
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Ascomicetos/química , Teoría Funcional de la Densidad , Estructura Molecular , Análisis Espectral/métodos , EstereoisomerismoRESUMEN
Seiridiasteriscane A is an asteriscane-type sesquiterpenoid bearing a trans-fused bicyclo[6.3.0]undecane skeleton. Although its biosynthesis has been proposed to involve a semipinacol rearrangement of a putative intermediary acetate bearing a bicyclo[7.2.0]undecane ring system (presumably derived from coisolated pestalotiopsin M) followed by epimerization of the resulting cis-fused seiridiasteriscane B, such a type of semipinacol rearrangement has never been reported so far. Our model study revealed that a 1-hydroxybicyclo[7.2.0]undecan-2-yl acetate underwent a smooth and stereospecific semipinacol rearrangement with the assistance of Et2AlCl to give the corresponding bicyclo[6.3.0]undecane-9-one. In addition, the resulting cis-fused 5,8-bicyclic ketone was partially epimerized to the corresponding trans-fused ketone by prolonged adsorption onto a silica gel plate. These results may support a recently proposed biosynthetic pathway of seiridiasteriscane A.
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Alcanos/química , Compuestos Bicíclicos con Puentes/química , Ciclohexenos/química , Modelos Químicos , Análisis Espectral/métodosRESUMEN
The effectiveness and limitations of density functional theory (DFT) calculations in the structural determination of complexed and conformationally flexible natural products were demonstrated using the cyclohelminthols CP-1 (1) CP-2 (2), CP-3 (3), and CP-4 (4) newly isolated from Helminthosporium velutinum yone96. Prior to DFT calculations, the structures were tentatively assigned using conventional spectroscopic analyses. The structures were verified with reference to DFT-derived 13C and 1H NMR chemical shifts, 3JHH and nJCH values, and electronic circular dichroism (ECD) spectra. The 13C chemical shift calculations were very effective for verifying the ring-structure moieties but less effective for verifying the geometry of the side chain in which the juncture asymmetric carbon (C-16) was apart from the ring-structure moiety. However, 1H chemical shift calculations compensated for the imperfection of the latter. ECD spectral calculations were used to determine the absolute configurations. Calculations for virtual simple model molecules enabled us to evaluate the reliability of the ECD spectral calculation and derive the chiral torsion responsible for the characteristic Cotton effects.
RESUMEN
To elucidate the biosynthesis of a fungicidal dimeric anhydride zopfiellin, the putative biosynthetic gene cluster was identified. We conducted heterologous expression of candidate genes for the synthesis of maleic anhydride and its dimerization and identified the two isomeric dimers with 9-membered rings as products. Notably, α-ketoglutarate-dependent dioxygenase ZopK oxidized one of the dimers, giving the 8-membered ring of zopfiellin. The mechanism of oxidative rearrangement is proposed by analyzing the incorporation of 13C-labeled precursors.
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Ciclooctanos/química , Dioxigenasas/metabolismo , Hongos/química , Anhídridos Maleicos/química , Anhídridos , Dimerización , Dioxigenasas/química , Hongos/metabolismo , Fungicidas Industriales , Estructura Molecular , Familia de Multigenes , Estrés OxidativoRESUMEN
Oxygenated cyclopentene systems are unique structural motifs found in fungal polyketides such as terrein, cyclohelminthols, and palmaenones. Here we report the identification of the biosynthetic gene clusters for cyclohelminthols and palmaenones and the functional characterization of the polyketide synthases and halogenases involved in the construction of 6-hydroxymellein derivatives. Heterologous expression in Aspergillus oryzae demonstrated that 6-hydroxymellein is a common biosynthetic intermediate and that chlorination occurs in the early stages of its products' biosynthesis. This was further confirmed by in vitro enzymatic reactions conducted in the presence of recombinant proteins. Plausible means of biogenesis of fungal polyketides from 6-hydroxymellein derivatives, additionally supported by the reported labeling patterns of terrein and structurally related fungal polyketides, are also discussed. This study sets the stage for elucidation of the biosynthetic machinery of fungal polyketides of this type.
Asunto(s)
Ciclopentanos/metabolismo , Ciclopropanos/metabolismo , Isocumarinas/metabolismo , Sintasas Poliquetidas/metabolismo , Aspergillus oryzae/enzimología , Ciclopentanos/química , Ciclopropanos/química , Isocumarinas/química , Estructura MolecularRESUMEN
Arundifungin (1) has been reported as a potent antifungal agent against Candida and Aspergillus spp; however, only its planar structure has been disclosed. This paper describes the assignment of the relative and absolute configuration of 1, which includes (a) determination of the relative configuration of the ABCD polycyclic ring moiety on the basis of detailed nuclear magnetic resonance (NMR) analysis, followed by the confirmation with density functional theory (DFT)-based 13 C NMR chemical shift calculations, (b) determination of the absolute configuration of the ABCD polycyclic ring moiety by observing a positive Cotton effect at 217 nm because of the C-8/C-9 tetrasubstituted double bond and its reproduction using DFT calculations, (c) determination of the configurational relationship between C-17 and C-20 by a combination of nuclear Overhauser effect (NOE) analysis and DFT-based conformational analysis, and (d) determination of the relative and absolute configuration of the C-24 and C-25 asymmetric centers on the acyclic side chain by a combination of chemical derivatization including modified Mosher's method and DFT-based conformational analysis, followed by electronic circular dichroism (ECD) spectral reproduction. Present study also discovered 26-deoxyarundifungin (2) of which relative structure was readily elucidated by 1 H and 13 C spectral comparison with those of 1. Since 2 exhibits slightly weaker antifungal activity against Cochliobolus miyabeanus than 1, the hydroxy group at C-26 moderately contributes to the activity.