RESUMEN
Zamamiphidins B (1) and C (2), two new manzamine-related alkaloids with an unprecedented fused diazahexacyclic ring system, were isolated from an Amphimedon sp. marine sponge collected in Okinawa. The structures of zamamiphidins B (1) and C (2) including the relative configurations were elucidated on the basis of spectroscopic data.
Asunto(s)
Alcaloides , Poríferos , Alcaloides/química , Alcaloides/aislamiento & purificación , Alcaloides/farmacología , Animales , Línea Celular Tumoral , Estructura Molecular , Poríferos/químicaRESUMEN
BACKGROUND: Some dinoflagellates cause harmful algal blooms, releasing toxic secondary metabolites, to the detriment of marine ecosystems and human health. Our understanding of dinoflagellate toxin biosynthesis has been hampered by their unusually large genomes. To overcome this challenge, for the first time, we sequenced the genome, microRNAs, and mRNA isoforms of a basal dinoflagellate, Amphidinium gibbosum, and employed an integrated omics approach to understand its secondary metabolite biosynthesis. RESULTS: We assembled the ~ 6.4-Gb A. gibbosum genome, and by probing decoded dinoflagellate genomes and transcriptomes, we identified the non-ribosomal peptide synthetase adenylation domain as essential for generation of specialized metabolites. Upon starving the cells of phosphate and nitrogen, we observed pronounced shifts in metabolite biosynthesis, suggestive of post-transcriptional regulation by microRNAs. Using Iso-Seq and RNA-seq data, we found that alternative splicing and polycistronic expression generate different transcripts for secondary metabolism. CONCLUSIONS: Our genomic findings suggest intricate integration of various metabolic enzymes that function iteratively to synthesize metabolites, providing mechanistic insights into how dinoflagellates synthesize secondary metabolites, depending upon nutrient availability. This study provides insights into toxin production associated with dinoflagellate blooms. The genome of this basal dinoflagellate provides important clues about dinoflagellate evolution and overcomes the large genome size, which has been a challenge previously.
Asunto(s)
Dinoflagelados/metabolismo , Genoma de Protozoos , MicroARNs/análisis , Isoformas de ARN/análisis , ARN Protozoario/análisis , Metabolismo Secundario , Dinoflagelados/genética , ARN de Algas/análisisRESUMEN
Two new bromotyrosine alkaloids, ceratinadins E (1) and F (2), were isolated from an Okinawan marine sponge Pseudoceratina sp. as well as a known bromotyrosine alkaloid, psammaplysin F (3). The gross structures of 1 and 2 were elucidated on the basis of spectroscopic data. The absolute configurations of 1 and 2 were assigned by comparison of the NMR and ECD data with those of a known related bromotyrosine alkaloid, psammaplysin A (4). Ceratinadins E (1) and F (2) are new bromotyrosine alkaloids possessing an 8,10-dibromo-9-methoxy-1,6-dioxa-2-azaspiro[4.6]undeca-2,7,9-trien-4-ol unit with two or three 11-N-methylmoloka'iamine units connected by carbonyl groups, respectively. Ceratinadin E (1) exhibited antimalarial activities against a drug-resistant and a drug-sensitive strains of Plasmodium falciparum (K1 and FCR3 strains, respectively).
Asunto(s)
Alcaloides/farmacología , Antimaláricos/farmacología , Plasmodium falciparum/efectos de los fármacos , Poríferos , Tirosina/análogos & derivados , Alcaloides/química , Alcaloides/aislamiento & purificación , Animales , Antimaláricos/química , Antimaláricos/aislamiento & purificación , Dicroismo Circular , Espectroscopía de Resonancia Magnética , Estructura Molecular , Compuestos de Espiro/química , Compuestos de Espiro/aislamiento & purificación , Compuestos de Espiro/farmacología , Tirosina/química , Tirosina/aislamiento & purificación , Tirosina/farmacologíaRESUMEN
The investigation of the Vietnamese marine sponge Spongia sp. led to the isolation of three new sesquiterpene phenols, langconols A-C (1-3), and one new sesquiterpene hydroxyquinone, langcoquinone C (4), together with two known meroterpenoids (5 and 6). Their structures were determined on the basis of spectroscopic analyses and comparisons with published data. Furthermore, the antibacterial assays of the isolates 1-6 suggested that 4 and 6 had significant antibacterial activities against Bacillus subtilis and Staphylococcus aureus, with MICs ranging from 6.25 to 25.0µM, while 1 and 3 possessed significant antibacterial activities against B. subtilis with MICs of 12.5 and 25.0µM, respectively. In contrast, cytotoxic assays of the isolated compounds 1-6, as well as compounds 7-15 previously isolated from this sponge, indicated that 1 and the previously reported anti-B. subtilis and anti-S. aureus sesquiterpene phenol 9 lacked cytotoxic activities against three human cancer cell lines (A549, lung cancer; MCF7, breast cancer; HeLa, cervix cancer) and a human normal cell line (WI-38 fibroblast).
Asunto(s)
Antibacterianos/química , Poríferos/química , Sesquiterpenos/química , Células A549 , Animales , Antibacterianos/aislamiento & purificación , Antibacterianos/farmacología , Antineoplásicos/química , Antineoplásicos/aislamiento & purificación , Bacillus subtilis/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células HeLa , Humanos , Células MCF-7 , Espectroscopía de Resonancia Magnética , Conformación Molecular , Poríferos/metabolismo , Sesquiterpenos/aislamiento & purificación , Sesquiterpenos/farmacología , Staphylococcus aureus/efectos de los fármacosRESUMEN
A new manzamine alkaloid, zamamidine D (1), was isolated from an Okinawan Amphimedon sp. marine sponge. The structure of zamamidine D (1) including the relative configuration was elucidated on the basis of spectroscopic data. Zamamidine D (1) is the first manzamine alkaloid possessing a 2,2'-methylenebistryptamine unit as the aromatic moiety instead of a ß-carboline unit. Zamamidine D (1) showed antimicrobial activity against several bacteria and fungi.
Asunto(s)
Alcaloides/aislamiento & purificación , Carbolinas/aislamiento & purificación , Poríferos/química , Alcaloides/química , Alcaloides/farmacología , Animales , Carbazoles/química , Carbolinas/química , Carbolinas/farmacología , Ensayos de Selección de Medicamentos Antitumorales , Japón , Biología Marina , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Resonancia Magnética Nuclear BiomolecularRESUMEN
Two distinct bacterial and eukaryotic serine racemases (SRs) have been identified based on phylogenetic and biochemical characteristics. Although some reports have suggested that marine heterotrophic bacteria have the potential to produce d-serine, the gene encoding bacterial SRs is not found in those bacterial genomes. In this study, using in-depth genomic analysis, we found that eukaryotic SR homologues were distributed widely in various bacterial genomes. Additionally, we selected a eukaryotic SR homologue from a marine heterotrophic bacterium, Roseobacter litoralis Och 149 (RiSR), and constructed an RiSR gene expression system in Escherichia coli for studying the properties of the enzyme. Among the tested amino acids, the recombinant RiSR exhibited both racemization and dehydration activities only towards serine, similar to many eukaryotic SRs. Mg2+ and MgATP enhanced both activities of RiSR, whereas EDTA abolished these enzymatic activities. The enzymatic properties and domain structure of RiSR were similar to those of eukaryotic SRs, particularly mammalian SRs. However, RiSR showed lower catalytic efficiency for L-serine dehydration (kcat/Km=0.094 min(-1) mM(-1)) than those of eukaryotic SRs reported to date (kcat/Km=0.6-21 min(-1) mM(-1)). In contrast, the catalytic efficiency for L-serine racemization of RiSR (kcat/Km=3.14 min(-1) mM(-1)) was 34-fold higher than that of l-serine dehydration. These data suggested that RiSR primarily catalysed serine racemization rather than dehydration.
Asunto(s)
Proteínas Bacterianas/química , Eucariontes/enzimología , Racemasas y Epimerasas/química , Roseobacter/enzimología , Secuencia de Aminoácidos , Bacterias/química , Bacterias/clasificación , Bacterias/enzimología , Bacterias/genética , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Eucariontes/química , Eucariontes/clasificación , Eucariontes/genética , Expresión Génica , Humanos , Cinética , Datos de Secuencia Molecular , Filogenia , Estructura Terciaria de Proteína , Transporte de Proteínas , Racemasas y Epimerasas/genética , Racemasas y Epimerasas/metabolismo , Roseobacter/química , Roseobacter/genética , Alineación de Secuencia , Especificidad por SustratoRESUMEN
The total synthesis of 7,10-epimer of the proposed structure of amphidinolideâ N was accomplished. The requisite chiral C17-C29 subunit was assembled stereoselectively via Keck allylation, Shi epoxidation, diastereoselective 1,3-reduction, and a later oxidative synthesis of the THF framework. The C1-C13 and C17-C29 subunits were successfully coupled using a Enders RAMP "linchpin" as the C14-C16 three carbon unit, thereby controlling the chirality at C14 and C16. The labile allyl epoxy moiety was successfully constructed by Grieco-Nishizawa olefination at a final stage of the synthesis.
RESUMEN
Amphidinolide N, the structure of which has been recently revised, is a 26-membered macrolide featuring allyl epoxide and tetrahydropyran moieties with 13 chiral centers. Due to its challenging structure and extraordinary potent cytotoxicity, amphidinolide N is a highly attractive target of total synthesis. During our total synthesis studies of the 7,10-epimer of the proposed structure of amphidinolide N, we have synthesized the C1-C13 subunit enantio- and diastereoselectively. Key reactions include an l-proline catalyzed enantioselective intramolecular aldol reaction, Evans aldol reaction, Sharpless asymmetric epoxidation and Tamao-Fleming oxidation. To aid late-stage manipulations, we also developed the 4-(N-benzyloxycarbonyl-N-methylamino)butyryl group as a novel ester protective group for the C9 alcohol.
RESUMEN
A novel, anaerobic bacterium, strain MO-SEDIT, was isolated from a methanogenic microbial community, which was originally obtained from marine subsurface sediments collected from off the Shimokita Peninsula of Japan. Cells were Gram-stain-negative, non-motile, non-spore-forming rods, 0.4-1.4 µm long by 0.4-0.6 µm wide. The cells also formed long filaments of up to about 11 µm. The strain grew on amino acids (i.e. valine, leucine, isoleucine, methionine, glycine, phenylalanine, tryptophan, lysine and arginine), pyruvate and melezitose in the presence of yeast extract. Growth was observed at 4-37 °C (optimally at 30 °C), at pH 6.0 and 8.5 (optimally at 7.0-7.5) and in 0-60âg l- 1 NaCl (optimally 20âg NaCl l- 1). The G+C content of the DNA was 32.0âmol%. The polar lipids of strain MO-SEDIT were phosphatidylglycerol, phosphatidyl lipids and unknown lipids. The major cellular fatty acids (>10 % of the total) were C14 : 0, C16 : 1ω9 and C16 : 0 dimethyl aldehyde. Comparative sequence analysis of the 16S rRNA gene showed that strain MO-SEDIT was affiliated with the genus Sedimentibacter within the phylum Firmicutes. It was related most closely to the type strain of Sedimentibacter saalensis (94 % sequence similarity). Based on the phenotypic and genetic characteristics, strain MO-SEDIT is considered to represent a novel species of the genus Sedimentibacter, for which the name Sedimentibacter acidaminivorans sp. nov. is proposed. The type strain is MO-SEDIT ( = JCM 17293T = DSM 24004T).
RESUMEN
A ß-1,3-glucanase (LpGluA) of deep subseafloor Laceyella putida JAM FM3001 was purified to homogeneity from culture broth. The molecular mass of the enzyme was around 36 kDa as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). LpGluA hydrolyzed curdlan optimally at pH 4.2 and 80 °C. In spite of the high optimum temperature, LpGluA showed relatively low thermostability, which was stabilized by adding laminarin, xylan, colloidal chitin, pectin, and its related polysaccharides. The gene for LpGluA cloned by using degenerate primers was composed of 1236 bp encoding 411 amino acids. Production of both LpGluA and a chitinase (LpChiA; Shibasaki et al. Appl Microbiol Biotechnol 98, 7845-7853, 2014) was induced by adding N-acetylglucosamine (GluNAc) to a culture medium of strain JAM FM3001. Construction of expression vectors containing the gene for LpGluA and its flanking regions showed the existence of a putative repressor protein.
Asunto(s)
Bacillales/enzimología , Bacillales/genética , Glucano 1,3-beta-Glucosidasa/genética , Glucano 1,3-beta-Glucosidasa/metabolismo , Acetilglucosamina/metabolismo , Secuencia de Aminoácidos , Bacillales/aislamiento & purificación , Secuencia de Bases , Clonación Molecular , Medios de Cultivo/química , Electroforesis en Gel de Poliacrilamida , Estabilidad de Enzimas , Sedimentos Geológicos , Glucano 1,3-beta-Glucosidasa/química , Glucano 1,3-beta-Glucosidasa/aislamiento & purificación , Concentración de Iones de Hidrógeno , Hidrólisis , Datos de Secuencia Molecular , Peso Molecular , Temperatura , Activación Transcripcional , beta-Glucanos/metabolismoRESUMEN
The biogenetic origins of amphidinin A (1) and amphidinolide P (2) were investigated by feeding experiments with (13)C-labeled acetates. (13)C-NMR data of (13)C-enriched samples revealed that the all carbons of 1 and 2 were derived from acetates. The polyketide chain of 1 was formed from one triketide chain, two diketide chains, and three unusual isolated C1 units derived from C-2 of cleaved acetates, while the polyketide chain of 2 was formed from one pentaketide chain, two acetate units, and three unusual isolated C1 units derived from C-2 of cleaved acetates. The all branched C1 units of 1 and 2 were derived from C-2 of cleaved acetates.
Asunto(s)
Furanos/metabolismo , Macrólidos/metabolismo , Dinoflagelados/química , Furanos/química , Furanos/aislamiento & purificación , Macrólidos/química , Macrólidos/aislamiento & purificación , Conformación MolecularRESUMEN
New bisindole alkaloids, hyrtinadines C (1) and D (2), have been isolated from an Okinawan marine sponge Hyrtios sp. The structures of hyrtinadines C (1) and D (2) were elucidated based on analyses of the spectral data. Hyrtinadines C (1) and D (2) were the relatively rare alkaloids possessing a 3,4-fused azepinoindole skeleton. Hyrtinadines C (1) and D (2) showed antimicrobial activity.
Asunto(s)
Antibacterianos/farmacología , Antifúngicos/farmacología , Alcaloides Indólicos/farmacología , Poríferos/química , Animales , Antibacterianos/química , Antibacterianos/aislamiento & purificación , Antifúngicos/química , Antifúngicos/aislamiento & purificación , Aspergillus niger/efectos de los fármacos , Bacillus subtilis/efectos de los fármacos , Candida albicans/efectos de los fármacos , Cryptococcus neoformans/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Escherichia coli/efectos de los fármacos , Alcaloides Indólicos/química , Alcaloides Indólicos/aislamiento & purificación , Pruebas de Sensibilidad Microbiana , Micrococcus luteus/efectos de los fármacos , Estructura Molecular , Staphylococcus aureus/efectos de los fármacos , Relación Estructura-Actividad , Trichophyton/efectos de los fármacosRESUMEN
Two new bromotyrosine alkaloids, tyrokeradines G (1) and H (2), have been isolated from an Okinawan marine sponge of the order Verongida. The structures of 1 and 2 were elucidated on the basis of spectroscopic data. Tyrokeradine G (1) is the first bromotyrosine alkaloid possessing a ß-alanine unit, while tyrokeradine H (2) is a rare bromotyrosine alkaloid possessing a N-substituted pyridinium ring. Tyrokeradines G (1) and H (2) showed antifungal activity.
Asunto(s)
Alcaloides/química , Alcaloides/aislamiento & purificación , Antifúngicos/química , Antifúngicos/aislamiento & purificación , Tirosina/análogos & derivados , Alcaloides/farmacología , Animales , Antifúngicos/farmacología , Aspergillus niger/efectos de los fármacos , Cryptococcus neoformans/efectos de los fármacos , Poríferos , Tirosina/química , Tirosina/aislamiento & purificación , Tirosina/farmacologíaRESUMEN
Speradine A is a derivative of cyclopiazonic acid (CPA) found in culture of an Aspergillus tamarii isolate. Heterologous expression of a predicted methyltransferase gene, cpaM, in the cpa biosynthesis gene cluster of A. tamarii resulted in the speradine A production in a 2-oxoCPA producing A. oryzae strain, indicating cpaM is involved in the speradine A biosynthesis.
Asunto(s)
Aspergillus/genética , Aspergillus/metabolismo , Indoles/metabolismo , Familia de Multigenes/genética , Secuencia de Aminoácidos , Secuencia de Bases , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Datos de Secuencia MolecularRESUMEN
The absolute configurations at six stereogenic centers in amphidinin A (1), a cytotoxic linear polyketide isolated from a symbiotic marine dinoflagellate, Amphidinium sp., were elucidated to be 2R, 4R, 6S, 9R 11R, and 12S by the combination of J-based configuration analysis, modified Mosher's method, and density-functional theory calculations.
Asunto(s)
Furanos/química , Policétidos/química , Dinoflagelados/química , Biología Marina , Estructura Molecular , Resonancia Magnética Nuclear Biomolecular , Policétidos/aislamiento & purificación , EstereoisomerismoRESUMEN
Iejimalides (IEJLs) A-D are 24-membered macrolides isolated from a tunicate Eudistoma cf. rigida, and exhibit potent cytotoxicity in vitro and antitumor activity in vivo. We previously reported that the molecular target of IEJL-A and -B was the vacuolar-type H(+)-ATPases (V-ATPases). However IEJL-C and -D, which are sulfonylated IEJL-A and -B, respectively, show more potent antitumor activity, and their molecular targets remain to be discovered. Here, we report that IEJL-C is also a potent V-ATPase inhibitor by binding in a site similar to the bafilomycin-binding site. Two-hour treatment with IEJL-C resulted in the complete disappearance of acidic organelles in HeLa cells. Interestingly, after 24-h treatment, small actin aggregates were observed instead of actin fibers. The same actin reorganization was also observed in cells treated with another V-ATPase inhibitor, bafilomycin A1. Because IEJLs did not inhibit actin polymerization in vitro, these results suggest that the primary target of IEJL-C, as well as IEJL-A and -B, is V-ATPase, and actin reorganizations are probably caused by the disruption of pH homeostasis via V-ATPase inhibition.
Asunto(s)
Actinas/química , Antineoplásicos/farmacología , Carbamatos/farmacología , Macrólidos/farmacología , ATPasas de Translocación de Protón Vacuolares/antagonistas & inhibidores , Actinas/metabolismo , Antineoplásicos/química , Carbamatos/química , Células HeLa , Humanos , Macrólidos/química , Estructura Molecular , LevadurasRESUMEN
New dimeric bromopyrrole-imidazole alkaloids, nagelamide I (1) and 2,2'-didebromonagelamide B (2), have been isolated from an Okinawan marine sponge Agelas species. The structures of 1 and 2 were elucidated based on analyses of the spectral data. Nagelamide I (1) was the first symmetric dimeric bromopyrrole-imidazole alkaloid consisting of two subunits connected with a single bond.
Asunto(s)
Agelas/química , Alcaloides/química , Imidazoles/química , Alcaloides/aislamiento & purificación , Animales , Halogenación , Imidazoles/aislamiento & purificación , Pirroles/química , Pirroles/aislamiento & purificaciónRESUMEN
New meroterpenoids, nakijiquinone S (1) and nakijinol C (2), have been isolated from an Okinawan marine sponge of the family Spongiidae. The gross structures and relative stereochemistries of 1 and 2 were elucidated on the basis of their spectral data. Nakijiquinone S (1) and nakijinol C (2) were new meroterpenoids consisting of a clerodane-type decalin ring connected to a 2-butoxy-5-hydroxy-benzoquinone unit or methyl 2,3,4-trihydroxybenzoate unit through a methylene, respectively. Nakijiquinone S (1) and nakijinol C (2) showed antimicrobial activities against several bacteria and fungi.
Asunto(s)
Antiinfecciosos/química , Antiinfecciosos/farmacología , Benzoquinonas/química , Benzoquinonas/farmacología , Poríferos/química , Pirogalol/análogos & derivados , Sesquiterpenos/química , Sesquiterpenos/farmacología , Animales , Antiinfecciosos/aislamiento & purificación , Bacterias/efectos de los fármacos , Infecciones Bacterianas/tratamiento farmacológico , Benzoquinonas/aislamiento & purificación , Hongos/efectos de los fármacos , Modelos Moleculares , Micosis/tratamiento farmacológico , Pirogalol/química , Pirogalol/aislamiento & purificación , Pirogalol/farmacología , Sesquiterpenos/aislamiento & purificaciónRESUMEN
Metachromins are a series of sesquiterpenoid quinones isolated from Okinawan marine sponges. Inhibitory effects of metachromins L-Q (1-6), sesquiterpenoid quinones with an amino acid residue, and their related analogs (7-18) prepared from metachromins A (19) and C (20) against receptor tyrosine kinases EGFR and HER2 were investigated. Two analogs 11 and 12 showed relatively stronger inhibitory activity against EGFR, while metachromins L-Q (1-6) and seven analogs (8, 10, 11, and 15-18) showed inhibitory activities against HER2.
Asunto(s)
Receptores ErbB/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Receptor ErbB-2/antagonistas & inhibidores , Sesquiterpenos/química , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Receptores ErbB/metabolismo , Humanos , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/toxicidad , Receptor ErbB-2/metabolismo , Sesquiterpenos/síntesis química , Sesquiterpenos/toxicidad , Relación Estructura-ActividadRESUMEN
Three new polyketides, manzamenones L-N (1-3), have been isolated from an Okinawan marine sponge of the genus Plakortis. The structures of 1-3 were elucidated on the basis of spectroscopic data. Manzamenones L-N (1-3) were new dimeric fatty-acid derivatives consisting of a tetrahydroindenone with three carboxy groups and two hexadecanyl chains. Manzamenones M (2) and N (3) showed antimicrobial activity against several bacteria and fungi.