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1.
Toxicon ; 203: 139-146, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34653444

RESUMO

The genus Brachycephalus includes small species of aposematic anurans known as microendemic, occurring in the mountains of the Atlantic Forest. Brachycephalus ephippium, B. nodoterga and B. pernix have been reported to contain the neurotoxin tetrodotoxin in skin and viscera. The biological conservation of several Brachycephalus species is currently threatened by climate change, deforestation, and the pandemic caused by the fungus Batrachochytrium dendrobatidis (Bd). Despite the well-known importance of amphibians' associated bacteria in the defensive role against pathogens, there is still a poor understanding of amphibian microbiome composition. The present study investigated the composition of B. pitanga microbial community and the presence of TTX in the host and in cultures of bacterial isolates, using a combination of metagenomics, bacterial culture isolation, mass spectrometry and metabolomic analyses. Results of culture-dependent and -independent analyses characterized the microbial communities associated with the skin and viscera of B. pitanga. Mass spectrometry analysis indicated the presence of TTX in host tissues, while bacterial production of TTX was not observed under the experimental conditions used in this investigation. This is the first report confirming the occurrence of TTX in B. pitanga.


Assuntos
Eugenia , Microbiota , Animais , Anuros , Bactérias , Tetrodotoxina/toxicidade
2.
Nat Prod Rep ; 2021 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-34647117

RESUMO

Covering period: up to 2019Water-soluble natural products constitute a relevant group of secondary metabolites notably known for presenting potent biological activities. Examples are aminoglycosides, ß-lactam antibiotics, saponins of both terrestrial and marine origin, and marine toxins. Although extensively investigated in the past, particularly during the golden age of antibiotics, hydrophilic fractions have been less scrutinized during the last few decades. This review addresses the possible reasons on why water-soluble metabolites are now under investigated and describes approaches and strategies for the isolation of these natural compounds. It presents examples of several classes of hydrosoluble natural products and how they have been isolated. Novel stationary phases and chromatography techniques are also reviewed, providing a perspective towards a renaissance in the investigation of water-soluble natural products.

4.
Chemistry ; 27(46): 11895-11903, 2021 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-34114710

RESUMO

The mycotoxin terrein is derived from the C10 -precursor 6-hydroxymellein (6-HM) via an oxidative ring contraction. Although the corresponding biosynthetic gene cluster (BGC) has been identified, details of the enzymatic oxidative transformations are lacking. Combining heterologous expression and in vitro studies we show that the flavin-dependent monooxygenase (FMO) TerC catalyzes the initial oxidative decarboxylation of 6-HM. The reactive intermediate is further hydroxylated by the second FMO TerD to yield a highly oxygenated aromatic species, but further reconstitution of the pathway was hampered. A related BGC was identified in the marine-derived Roussoella sp. DLM33 and confirmed by heterologous expression. These studies demonstrate that the biosynthetic pathways of terrein and related (polychlorinated) congeners diverge after oxidative decarboxylation of the lactone precursor that is catalyzed by a conserved FMO and further indicate that early dehydration of the side chain is an essential step.


Assuntos
Produtos Biológicos , Ciclopentanos , Oxirredução , Estresse Oxidativo
5.
Chem Commun (Camb) ; 57(47): 5758-5761, 2021 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-34002741

RESUMO

We report a selective, mild, and efficient C-H functionalization of tryptophan and tryptophan-containing peptides with activated α-bromo-carbonyl compounds under visible-light irradiation. The protocol efficiency is outlined by the wide substrate scope and excellent tolerance of sensitive functional groups present in the amino acid side chains. The method can be successfully extended to access pharmaco-peptide conjugate scaffolds.


Assuntos
Indóis/química , Peptídeos/química , Triptofano/química , Alquilação , Catálise , Estrutura Molecular , Processos Fotoquímicos
6.
Angew Chem Int Ed Engl ; 60(29): 15891-15898, 2021 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-33961724

RESUMO

Although swarming motility and biofilms are opposed collective behaviors, both contribute to bacterial survival and host colonization. Pseudovibrio bacteria have attracted attention because they are part of the microbiome of healthy marine sponges. Two-thirds of Pseudovibrio genomes contain a member of a nonribosomal peptide synthetase-polyketide synthase gene cluster family, which is also found sporadically in Pseudomonas pathogens of insects and plants. After developing reverse genetics for Pseudovibrio, we isolated heptapeptides with an ureido linkage and related nonadepsipeptides we termed pseudovibriamides A and B, respectively. A combination of genetics and imaging mass spectrometry experiments showed heptapetides were excreted, promoting motility and reducing biofilm formation. In contrast to lipopeptides widely known to affect motility/biofilms, pseudovibriamides are not surfactants. Our results expand current knowledge on metabolites mediating bacterial collective behavior.


Assuntos
Peptídeos/metabolismo , Poríferos/genética , Poríferos/metabolismo , Animais , Família Multigênica/genética , Peptídeo Sintases/genética , Peptídeo Sintases/metabolismo , Policetídeo Sintases/genética , Policetídeo Sintases/metabolismo , Simbiose
7.
mBio ; 12(1)2021 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-33563828

RESUMO

Penicillium digitatum is the most aggressive pathogen of citrus fruits. Tryptoquialanines are major indole alkaloids produced by P. digitatum It is unknown if tryptoquialanines are involved in the damage of citrus fruits caused by P. digitatum. To investigate the pathogenic roles of tryptoquialanines, we initially asked if tryptoquialanines could affect the germination of Citrus sinensis seeds. Exposure of the citrus seeds to tryptoquialanine A resulted in a complete inhibition of germination and an altered metabolic response. Since this phytotoxic effect requires the extracellular export of tryptoquialanine A, we investigated the mechanisms of extracellular delivery of this alkaloid in P. digitatum We detected extracellular vesicles (EVs) released by P. digitatum both in culture and during infection of citrus fruits. Compositional analysis of EVs produced during infection revealed the presence of a complex cargo, which included tryptoquialanines and the mycotoxin fungisporin. The EVs also presented phytotoxicity activity in vitro and caused damage to the tissues of citrus seeds. Through molecular networking, it was observed that the metabolites present in the P. digitatum EVs are produced in all of its possible hosts. Our results reveal a novel phytopathogenic role of P. digitatum EVs and tryptoquialanine A, implying that this alkaloid is exported in EVs during plant infection.IMPORTANCE During the postharvest period, citrus fruits can be affected by phytopathogens such as Penicillium digitatum, which causes green mold disease and is responsible for up to 90% of total citrus losses. Chemical fungicides are widely used to prevent green mold disease, leading to concerns about environmental and health risks. To develop safer alternatives to control phytopathogens, it is necessary to understand the molecular basis of infection during the host-pathogen interaction. In the P. digitatum model, the virulence strategies are poorly known. Here, we describe the production of phytotoxic extracellular vesicles (EVs) by P. digitatum during the infection of citrus fruits. We also characterized the secondary metabolites in the cargo of EVs and found in this set of molecules an inhibitor of seed germination. Since EVs and secondary metabolites have been related to virulence mechanisms in other host-pathogen interactions, our data are important for the comprehension of how P. digitatum causes damage to its primary hosts.


Assuntos
Alcaloides/metabolismo , Alcaloides/farmacologia , Citrus/microbiologia , Vesículas Extracelulares/química , Penicillium/patogenicidade , Sementes/crescimento & desenvolvimento , Alcaloides/biossíntese , Frutas/microbiologia , Fungicidas Industriais/farmacologia , Interações Hospedeiro-Patógeno , Doenças das Plantas/microbiologia , Metabolismo Secundário , Sementes/efeitos dos fármacos , Sementes/metabolismo , Sementes/microbiologia
8.
Nat Prod Rep ; 38(3): 586-667, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33021301

RESUMO

Covering: 2017-2019Guanidine natural products isolated from microorganisms, marine invertebrates and terrestrial plants, amphibians and spiders, represented by non-ribosomal peptides, guanidine-bearing polyketides, alkaloids, terpenoids and shikimic acid derived, are the subject of this review. The topics include the discovery of new metabolites, total synthesis of natural guanidine compounds, biological activity and mechanism-of-action, biosynthesis and ecological functions.


Assuntos
Anuros/metabolismo , Bactérias/metabolismo , Produtos Biológicos/química , Fungos/metabolismo , Guanidinas/metabolismo , Animais , Organismos Aquáticos/química , Organismos Aquáticos/metabolismo , Bactérias/química , Bactérias/genética , Produtos Biológicos/metabolismo , Fungos/química , Invertebrados/química , Invertebrados/metabolismo , Estrutura Molecular , Plantas/química , Plantas/metabolismo , Saxitoxina/química , Saxitoxina/metabolismo , Metabolismo Secundário , Aranhas/química , Aranhas/metabolismo , Tetrodotoxina/química , Tetrodotoxina/metabolismo
9.
J Nat Prod ; 84(3): 790-796, 2021 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-33371682

RESUMO

Metabolomics analysis detected tambjamine alkaloids in aqueous and EtOAc extracts of the marine invertebrates Virididentula dentata, Tambja stegosauriformis, Tambja brasiliensis, and Roboastra ernsti. Among several tambjamines, the new amino acid derivatives tambjamines M-O (17-19) were identified by Marfey's advanced analysis, UPLC-MS/MS analyses, and total synthesis. The tambjamine diversity increased from the bryozoan V. dentata to its nudibranch predators T. stegosauriformis and T. brasiliensis and attained a higher diversity in R. ernsti, the nudibranch that preys upon T. stegosauriformis and T. brasiliensis. The total tambjamine content also increases among the trophic levels, probably due to biomagnification. Tambjamines A (1), C (3), and D (4) are the major metabolites in the tissues of V. dentata, T. stegosauriformis, T. brasiliensis, and R. ernsti and are likely the main chemical defenses of these marine invertebrates.

10.
Microbiol Res ; 244: 126667, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33338969

RESUMO

It is known that different plant species select specific microbes to live inside their tissues in a process determined by the host genotype, phenotype and geographic location, which can introduce discussion on plant endemism and the assembly of specific microbial communities. Herein, we report the results of an investigation relating the geographic distribution of plant species and the composition of microbial communities associated with plant hosts. The bacterial and fungal community associated with Anthurium plant leaves was mapped to assess the diversity and ecology of the endophytic community associated with Anthurium spp. collected on islands and on the Brazilian mainland. Twenty-six Anthurium specimens were surveyed, distributed throughout the São Paulo state coastline, including Alcatrazes Island, some coastal islands and distinct mainland environments. Bacterial and fungal endophytes were obtained from the leaves of A. alcatrazense, A. loefgrenii, A. penthaphyllum, A. urvellianum and A. intermedium and subjected to massive bacterial 16S rRNA and fungal ITS sequencing. The results indicated that A. alcatrazense, endemic to Alcatrazes Island, hosted a specific bacterial community structure, while its fungal community was similar to that of Anthurium species from other locations. Betaproteobacteria showed a high differential occurrence in A. alcatrazense. Some groups of fungi were found mainly inhabiting A. loefgrenii plants. While Alphaproteobacteria, Gammaproteobacteria, Actinobacteria and Sordariomycetes, Dothiodeomycetes and Tremellomycetes composed the core microbial community among Anthurium plants. The results suggest crucial role for the bacterial communities to endemic plants, while endophytic fungal diversity is less specifically distributed among endemic and nonendemic plant species.


Assuntos
Araceae/microbiologia , Bactérias/isolamento & purificação , Fungos/isolamento & purificação , Microbiota , Micobioma , Bactérias/classificação , Bactérias/genética , Biodiversidade , Brasil , Fungos/classificação , Fungos/genética , Folhas de Planta/microbiologia
11.
Bioorg Chem ; 105: 104402, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33130347

RESUMO

ATP-Binding Cassette (ABC) transporters are the main class of transmembrane transporters involved in pathogenic fungal resistance against chemotherapeutic agents. Herein we report results which show that batzelladine D (1) and norbatzelladine L (2) reverse the fluconazole resistance phenotype mediated by Pdr5p transporter on Saccharomyces cerevisiae. Both alkaloids were able to chemosensitize the Pdr5p-overexpressing strain by synergistic interaction with fluconazole. Both compounds also showed an inhibitory effect on the catalytic activity and on the intracellular accumulation of rhodamine 6G, and did not show significant in vitro mammalian cells toxicity.


Assuntos
Alcaloides/farmacologia , Fluconazol/farmacologia , Poríferos/química , Pirimidinas/farmacologia , Rodaminas/antagonistas & inibidores , Transportadores de Cassetes de Ligação de ATP/metabolismo , Alcaloides/química , Alcaloides/isolamento & purificação , Animais , Células Cultivadas , Relação Dose-Resposta a Droga , Camundongos , Estrutura Molecular , Pirimidinas/química , Pirimidinas/isolamento & purificação , Rodaminas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Relação Estrutura-Atividade
12.
J Nat Prod ; 83(6): 1784-1793, 2020 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-32525315

RESUMO

Herein reported are results of the chemical and biological investigation of red propolis collected at the Brazilian Northeast coastline. New propolones A-D (1-4), with a 3-{3-[(2-phenylbenzofuran-3-yl)methyl]phenyl}chromane skeleton; propolonones A-C (5-7), with a 3-[3-(3-benzylbenzofuran-2-yl)phenyl]chromane skeleton; and propolol A (8), with a 6-(3-benzylbenzofuran-2-yl)-3-phenylchromane skeleton, were isolated as constituents of Brazilian red propolis by cytotoxicity-guided assays and structurally identified by analysis of their spectroscopic data. Propolone B (2) and propolonone A (5) display significant cytotoxic activities against an ovarian cancer cell line expressing a multiple drug resistance phenotype when compared with doxorubicin.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Própole/química , Antibióticos Antineoplásicos/farmacologia , Brasil , Linhagem Celular Tumoral , Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Neoplasias Ovarianas/tratamento farmacológico
13.
Bioorg Chem ; 100: 103921, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32464403

RESUMO

Enterococci are gram-positive, widespread nosocomial pathogens that in recent years have developed resistance to various commonly employed antibiotics. Since finding new infection-control agents based on secondary metabolites from organisms has proved successful for decades, natural products are potentially useful sources of compounds with activity against enterococci. Herein are reported the results of a natural product library screening based on a whole-cell assay against a gram-positive model organism, which led to the isolation of a series of anacardic acids identified by analysis of their spectroscopic data and by chemical derivatizations. Merulinic acid C was identified as the most active anacardic acid derivative obtained against antibiotic-resistant enterococci. Fluorescence microscopy analyses showed that merulinic acid C targets the bacterial membrane without affecting the peptidoglycan and causes rapid cellular ATP leakage from cells. Merulinic acid C was shown to be synergistic with gentamicin against Enterococcus faecium, indicating that this compound could inspire the development of new antibiotic combinations effective against drug-resistant pathogens.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Enterococcus faecium/efeitos dos fármacos , Gentamicinas/farmacologia , Sinergismo Farmacológico , Enterococcus faecium/crescimento & desenvolvimento , Enterococcus faecium/metabolismo , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Infecções por Bactérias Gram-Positivas/microbiologia , Humanos , Hidroxibenzoatos/farmacologia
14.
Chemosphere ; 242: 125211, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31896201

RESUMO

While tetrodotoxin (TTX) is commonly found in pufferfish tissues, it is unclear if bacterial symbionts isolated from pufferfish tissues can produce TTX. In this investigation, UPLC qTOF-MS/MS analysis of tissue extracts obtained from Sphoeroides spengleri and Canthigaster figuereidoi identified TTX in their composition, indicating their consumption is unsafe. UPLC qTOF-MS/MS analysis coupled with Molecular Networking indicated new TTX analogs (methyl-TTX, TTX-acetate, hydroxypropyl-TTX and glycerol-TTX). Bacterial extracts from sixteen strains revealed a compound with a [M+H]+ ion at m/z 320.1088, identical to TTX. However, TTX itself was not detected in these cultures by UPLC-MS/MS. Neurotoxicity of Vibrio A665 purified fraction 2 (with precursor [M+H]+ ion at m/z 320.1088) was significant in human neural stem cells (hNSCs), but the Nav blockage activity was not confirmed by the veratridine/ouabain essays, indicating a possible difference in the mechanism of action between the bacterium A665 purified fraction 2 and TTX. Vibrios symbionts of pufferfish point out involving in the production of TTX precursors.


Assuntos
Microbiota , Tetraodontiformes/fisiologia , Tetrodotoxina/metabolismo , Animais , Brasil , Cromatografia Líquida , Humanos , Espectrometria de Massas em Tandem , Tetraodontiformes/microbiologia , Vibrio
15.
J Nat Prod ; 83(1): 55-65, 2020 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-31895573

RESUMO

A new method of screening was developed to generate 770 organic and water-soluble fractions from extracts of nine species of marine sponges, from the growth media of 18 species of marine-derived fungi, and from the growth media of 13 species of endophytic fungi. The screening results indicated that water-soluble fractions displayed significant bioactivity in cytotoxic, antibiotic, anti-Leishmania, anti-Trypanosoma cruzi, and inhibition of proteasome assays. Purification of water-soluble fractions from the growth medium of Penicillium solitum IS1-A provided the new glutamic acid derivatives solitumine A (1), solitumine B (2), and solitumidines A-D (3-6). The structures of compounds 1-6 have been established by analysis of spectroscopic data, chemical derivatizations, and vibrational circular dichroism calculations. Although no biological activity could be observed for compounds 1-6, the new structures reported for 1-6 indicate that the investigation of water-soluble natural products represents a relevant strategy in finding new secondary metabolites.


Assuntos
Glutamatos/química , Regiões Antárticas , Fungos/química , Estrutura Molecular , Penicillium/química , Água
16.
Rev. bras. farmacogn ; 29(6): 715-719, Nov.-Dec. 2019. tab, graf
Artigo em Inglês | LILACS-Express | LILACS | ID: biblio-1057861

RESUMO

ABSTRACT Chemical investigation of the aqueous fraction of the ethanol extract from the Brazilian endemic marine sponge Clathria (Clathria) nicoleae Vieira de Barros, Santos & Pinheiro, 2013, Microcionidae, sampled from a 55 m deep rhodolith bed at the Amazon River mouth, led to the isolation of a new hexapeptide, clathriamide (1). HP-20 resin was used to capture compound 1 from the aqueous fraction, which was purified by additional chromatographic steps. The absolute configuration of the amino acids of 1 was determined by advanced Marfey's analysis using 5-fluoro-2,4-dinitrophenyl-Nα-L-tryptophanamide. The amino acid derivatives analyzed by ultra-performance liquid chromatography coupled to a mass spectrometry using a C8 column enabled a good chromatographic resolution of L-Ile and L-allo-Ile, previously unfeasible using C18 column.

17.
Nat Prod Rep ; 36(7): 981-1004, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31049509

RESUMO

Covering: up to 2019The discovery of new bioactive natural products gained momentum during the last few decades, resulting from instrumentation advances, from the expansion of genome mining and regulation, as well as by exploration of untapped biological sources. However, water-soluble, volatile, minor and photosensitive natural products are yet poorly known. This review discusses the literature reporting the isolation strategies for some of these metabolites. Analysis of minor metabolites at sub-milligram level are also presented, since analytical instrumentation enabling structure assignment in minute quantities is now routine. Major trends related to natural products discovery are discussed, under the light of further developments in biodiscovery.


Assuntos
Produtos Biológicos/isolamento & purificação , Produtos Biológicos/química , Produtos Biológicos/efeitos da radiação , Interações Hidrofóbicas e Hidrofílicas , Luz , Estrutura Molecular , Compostos Orgânicos Voláteis/química , Compostos Orgânicos Voláteis/isolamento & purificação , Compostos Orgânicos Voláteis/efeitos da radiação
18.
J Nat Prod ; 81(10): 2296-2300, 2018 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-30281303

RESUMO

The new pyrrole-imidazole and pyrrole-guanidine alkaloids 4-debromooroidin (1), 4-debromougibohlin (2), 5-debromougibohlin (3), and 5-bromopalau'amine (4), along with the known hymenidin (5) and (+)-monobromoisophakellin (6), have been isolated from a Dictyonella sp. marine sponge, collected at the Amazon River mouth. The bromine-substitution pattern observed for compounds 1, 2 and 4 is unusual among bromopyrrole alkaloids isolated from marine sponges. The 20S proteasome inhibitory activities of compounds 1-6 have been recorded, with 5-bromopalau'amine (4) being the most active in this series.


Assuntos
Poríferos/química , Inibidores de Proteassoma/química , Inibidores de Proteassoma/farmacologia , Pirróis/química , Pirróis/farmacologia , Animais , Brasil , Estrutura Molecular , Complexo de Endopeptidases do Proteassoma , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética
19.
Nat Chem ; 10(9): 938-945, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30061613

RESUMO

Studies of secondary metabolites (natural products) that cover their isolation, chemical synthesis and bioactivity investigation present myriad opportunities for discovery. For example, the isolation of novel secondary metabolites can inspire advances in chemical synthesis strategies to achieve their practical preparation for biological evaluation. In the process, chemical synthesis can also provide unambiguous structural characterization of the natural products. Although the isolation, chemical synthesis and bioactivity studies of natural products are mutually beneficial, they are often conducted independently. Here, we demonstrate the benefits of a collaborative study of the phomactins, diterpenoid fungal metabolites that serve as antagonists of the platelet activating factor receptor. Our isolation of novel phomactins has spurred the development of a bioinspired, unified approach that achieves the total syntheses of six congeners. We also demonstrate in vitro the beneficial effects of several phomactins in suppressing the rate of repopulation of tumour cells following gamma radiation therapy.


Assuntos
Produtos Biológicos/síntese química , Terpenos/química , Produtos Biológicos/isolamento & purificação , Produtos Biológicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Fungos/química , Fungos/metabolismo , Raios gama , Humanos , Concentração Inibidora 50 , Glicoproteínas da Membrana de Plaquetas/antagonistas & inibidores , Glicoproteínas da Membrana de Plaquetas/metabolismo , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Acoplados a Proteínas G/metabolismo , Estereoisomerismo , Relação Estrutura-Atividade , Terpenos/isolamento & purificação , Terpenos/farmacologia
20.
Org Biomol Chem ; 16(35): 6450-6459, 2018 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-30141817

RESUMO

Antimicrobial and anti-proliferative meleagrin and oxaline are roquefortine C-derived alkaloids produced by fungi of the genus Penicillium. Tandem O-methylations complete the biosynthesis of oxaline from glandicoline B through meleagrin. Currently, little is known about the role of these methylation patterns in the bioactivity profile of meleagrin and oxaline. To establish the structural and mechanistic basis of methylation in these pathways, crystal structures were determined for two late-stage methyltransferases in the oxaline and meleagrin gene clusters from Penicillium oxalicum and Penicillium chrysogenum. The homologous enzymes OxaG and RoqN were shown to catalyze penultimate hydroxylamine O-methylation to generate meleagrin in vitro. Crystal structures of these enzymes in the presence of methyl donor S-adenosylmethionine revealed an open active site, which lacks an apparent base indicating that catalysis is driven by proximity effects. OxaC was shown to methylate meleagrin to form oxaline in vitro, the terminal pathway product. Crystal structures of OxaC in a pseudo-Michaelis complex containing sinefungin and meleagrin, and in a product complex containing S-adenosyl-homocysteine and oxaline, reveal key active site residues with His313 serving as a base that is activated by Glu369. These data provide structural insights into the enzymatic methylation of these alkaloids that include a rare hydroxylamine oxygen acceptor, and can be used to guide future efforts towards selective derivatization and structural diversification and establishing the role of methylation in bioactivity.


Assuntos
Imidazóis/metabolismo , Metiltransferases/metabolismo , Ovomucina/biossíntese , Metiltransferases/química , Modelos Moleculares , Penicillium/enzimologia , Penicillium/metabolismo , Conformação Proteica
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