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1.
Biosci Biotechnol Biochem ; 86(8): 1013-1023, 2022 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-35648459

RESUMO

Simplified analogs of aplysiatoxin (ATX) such as 10-Me-aplog-1 exhibit potent antiproliferative activity toward human cancer cell lines by activating protein kinase C (PKC). However, the synthesis of 10-Me-aplog-1 involved a 23-step longest linear sequence (LLS). Therefore, we have been working toward the development of a more synthetically accessible analog of ATX. In this study, we designed a new analog of ATX wherein a cyclic ketal moiety derived from (R)-(-)-carvone replaced the spiroketal moiety in 18-deoxy-aplog-1. The new analog's synthesis proceeded in an 8-step LLS. Although the configuration at position 3 of the cyclic ketal in the (R)-(-)-carvone-based analog was opposite to those of ATX and 18-deoxy-aplog-1, the antiproliferative activity toward human cancer cell lines of the carvone-based analog was comparable with that of 18-deoxy-aplog-1. The obtained results indicate the potential of the carvone-based analog as a basis for discovering PKC-targeting molecules requiring a decreased number of synthetic steps.


Assuntos
Antineoplásicos , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Monoterpenos Cicloexânicos , Humanos , Toxinas de Lyngbya , Proteína Quinase C/metabolismo , Relação Estrutura-Atividade
3.
Appl Microbiol Biotechnol ; 106(4): 1521-1530, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-35138454

RESUMO

Streptomyces clavuligerus is an industrially important producer of clavulanic acid (CA), a ß-lactamase inhibitor which is used together with amoxicillin in one of the most widely prescribed antibacterial medicines, the co-amoxiclav. In a mid-eighties ATCC vial of S. clavuligerus ATCC 27064 culture, we have found a new genotype, which was apparently lost from the subsequent ATCC collection stocks, and has remained obscure to the scientific community. Most importantly, this genotype harbors teleocidin (lyngbyatoxin) biosynthetic genes, which are located on an enigmatic 138 kb chromosomal region and support accumulation of significant amounts of these highly toxic, tumor-promoting secondary metabolites in cultures of S. clavuligerus. While this genomic region is completely absent from all published sequences for S. clavuligerus ATCC strain, at least one of the industrial strains for commercial production of CA, originating from ATCC 27064, retained the genetic potential for production of teleocidins. The origin of teleocidin biosynthetic cluster can now be traced back to early S. clavuligerus stocks at the ATCC. Our work provides a genome sequence and a deposited monoisolate of this genotype. Given the scale of industrial use of S. clavuligerus world-wide and toxicity of teleocidins, we also discuss the environmental and safety implications and provide a method of abolishing teleocidin production without affecting productivity of CA. KEY POINTS: • Early stocks of S. clavuligerus ATCC 27064 produce toxic teleocidins • Teleocidin biosynthetic genes were found within a distinct S. clavuligerus genotype • The genotype has been passed on to some industrial clavulanic acid producer strains.


Assuntos
Toxinas de Lyngbya , Streptomyces , Ácido Clavulânico , Genótipo , Toxinas de Lyngbya/metabolismo , Streptomyces/genética , Streptomyces/metabolismo
4.
Molecules ; 27(3)2022 Jan 26.
Artigo em Inglês | MEDLINE | ID: mdl-35164089

RESUMO

Liver cancer is a leading cause of cancer death globally. Marine mollusc-derived drugs have gained attention as potential natural-based anti-cancer agents to overcome the side effects caused by conventional chemotherapeutic drugs during cancer therapy. Using liquid chromatography-mass spectrometry, the main biomolecules in the purple ink secretion released by the sea hare, named Bursatella leachii (B. leachii), were identified as hectochlorin, malyngamide X, malyngolide S, bursatellin and lyngbyatoxin A. The cytotoxic effects of B. leachii ink concentrate against human hepatocarcinoma (HepG2) cells were determined to be dose- and time-dependent, and further exploration of the underlying mechanisms causing the programmed cell death (apoptosis) were performed. The expression of cleaved-caspase-8 and cleaved-caspase-3, key cysteine-aspartic proteases involved in the initiation and completion of the apoptosis process, appeared after HepG2 cell exposure to the B. leachii ink concentrate. The gene expression levels of pro-apoptotic BAX, TP53 and Cyclin D1 were increased after treatment with the B. leachii ink concentrate. Applying in silico approaches, the high scores predicted that bioactivities for the five compounds were protease and kinase inhibitors. The ADME and cytochrome profiles for the compounds were also predicted. Altogether, the B. leachii ink concentrate has high pro-apoptotic potentials, suggesting it as a promising safe natural product-based drug for the treatment of liver cancer.


Assuntos
Antineoplásicos/farmacologia , Produtos Biológicos/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Gastrópodes/química , Neoplasias Hepáticas/tratamento farmacológico , Amidas/química , Amidas/isolamento & purificação , Amidas/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Apoptose/efeitos dos fármacos , Produtos Biológicos/química , Produtos Biológicos/isolamento & purificação , Células Hep G2 , Humanos , Lactonas/química , Lactonas/isolamento & purificação , Lactonas/farmacologia , Toxinas de Lyngbya/química , Toxinas de Lyngbya/isolamento & purificação , Toxinas de Lyngbya/farmacologia , Pirrolidinonas/química , Pirrolidinonas/isolamento & purificação , Pirrolidinonas/farmacologia , Tiazóis/química , Tiazóis/isolamento & purificação , Tiazóis/farmacologia
5.
Chembiochem ; 23(3): e202100574, 2022 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-34850512

RESUMO

Indolactam alkaloids are activators of protein kinase C (PKC) and are of pharmacological interest for the treatment of pathologies involving PKC dysregulation. The marine cyanobacterial nonribosomal peptide synthetase (NRPS) pathway for lyngbyatoxin biosynthesis, which we previously expressed in E. coli, was studied for its amenability towards the biosynthesis of indolactam variants. Modification of culture conditions for our E. coli heterologous expression host and analysis of pathway products suggested the native lyngbyatoxin pathway NRPS does possess a degree of relaxed specificity. Site-directed mutagenesis of two positions within the adenylation domain (A-domain) substrate-binding pocket was performed, resulting in an alteration of substrate preference between valine, isoleucine, and leucine. We observed relative congruence of in vitro substrate activation by the LtxA NRPS to in vivo product formation. While there was a preference for isoleucine over leucine, the substitution of alternative tailoring domains may unveil the true in vivo effects of the mutations introduced herein.


Assuntos
Toxinas de Lyngbya/biossíntese , Peptídeo Sintases/metabolismo , Toxinas de Lyngbya/química , Estrutura Molecular , Mutagênese Sítio-Dirigida , Peptídeo Sintases/genética
6.
Mar Drugs ; 19(11)2021 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-34822501

RESUMO

Potassium channel Kv1.5 has been considered a key target for new treatments of atrial tachyarrhythmias, with few side effects. Four new debromoaplysiatoxin analogues with a 6/6/12 fused ring system were isolated from marine cyanobacterium Lyngbya sp. Their planar structures were elucidated by HRESIMS, 1D and 2D NMR. The absolute configuration of oscillatoxin J (1) was determined by single-crystal X-ray diffraction, and the absolute configurations of oscillatoxin K (2), oscillatoxin L (3) and oscillatoxin M (4) were confirmed on the basis of GIAO NMR shift calculation followed by DP4 analysis. The current study confirmed the absolute configuration of the pivotal chiral positions (7S, 9S, 10S, 11R, 12S, 15S, 29R and 30R) at traditional ATXs with 6/12/6 tricyclic ring system. Compound 1, 2 and 4 exhibited blocking activities against Kv1.5 with IC50 values of 2.61 ± 0.91 µM, 3.86 ± 1.03 µM and 3.79 ± 1.01 µM, respectively. However, compound 3 exhibited a minimum effect on Kv1.5 at 10 µM. Furthermore, all of these new debromoaplysiatoxin analogs displayed no apparent activity in a brine shrimp toxicity assay.


Assuntos
Canal de Potássio Kv1.5/efeitos dos fármacos , Toxinas de Lyngbya/farmacologia , Lyngbya , Animais , Organismos Aquáticos , Artemia , Humanos , Concentração Inibidora 50 , Canal de Potássio Kv1.5/antagonistas & inibidores , Toxinas de Lyngbya/química , Camundongos , Relação Estrutura-Atividade
7.
J Nat Med ; 75(3): 467-474, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33675456

RESUMO

The teleocidin B family members are terpene indole compounds isolated from Streptomyces bacteria, and they strongly activate protein kinase C (PKC). Their unique structures have attracted many researchers in the natural product chemistry and pharmacology fields, and numerous isolation and bioactivity studies have been conducted. The accumulated information has facilitated the identification of the enzymatic reactions in teleocidin biosynthesis, and new developments in structural biology have strongly aided efforts to clarify the finer points of these reactions. This review describes the recent biochemical and structural biological studies to reveal their reaction mechanisms, with a primary focus on the terpene cyclization triggered by the C-N bond formation by P450 oxygenase (TleB), the prenyltransferase (TleC), and the methyltransferase (TleD). This new knowledge will benefit future engineering studies to create unnatural PKC activators.


Assuntos
Indóis/metabolismo , Toxinas de Lyngbya/biossíntese , Streptomyces/enzimologia , Terpenos/metabolismo , Proteínas de Bactérias/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Dimetilaliltranstransferase/metabolismo , Metiltransferases/metabolismo , Estrutura Molecular , Família Multigênica
8.
Biosci Biotechnol Biochem ; 85(1): 168-180, 2021 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-33577665

RESUMO

10-Methyl-aplog-1 (1), a simplified analog of debromoaplysiatoxin, exhibits a high binding affinity for protein kinase C (PKC) isozymes and potent antiproliferative activity against several cancer cells with few adverse effects. A recent study has suggested that its phenol group in the side chain is involved in hydrogen bonding and CH/π interactions with the binding cleft-forming loops in the PKCδ-C1B domain. To clarify the effects of the side chain length on these interactions, four analogs of 1 with various lengths of side chains (2-5) were prepared. The maximal PKC binding affinity and antiproliferative activity were observed in 1. Remarkably, the introduction of a bromine atom into the phenol group of 2 increased not only these activities but also proinflammatory activity. These results indicated that 1 has the optimal side chain length as an anticancer seed. This conclusion was supported by docking simulations of 1-5 to the PKCδ-C1B domain.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Toxinas de Lyngbya/química , Toxinas de Lyngbya/farmacologia , Proteína Quinase C-delta/metabolismo , Antineoplásicos/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Inflamação/induzido quimicamente , Toxinas de Lyngbya/metabolismo , Modelos Moleculares , Ligação Proteica , Domínios Proteicos , Proteína Quinase C-delta/química , Relação Estrutura-Atividade
9.
Chemistry ; 27(9): 2963-2972, 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-32996659

RESUMO

Teleocidins are potent protein kinase C activators, and possess a unique indole-fused nine-membered lactam structure. Teleocidin biosynthesis starts from the formation of a dipeptide by non-ribosomal peptide synthetase (NRPS), followed by oxidative C-N bond formation by a cytochrome P450 oxidase, reverse-prenylation by a prenyltransferase, and methylation-initiated terpene cyclization by a C-methyltransferase. This minireview focuses on recent research progress toward the elucidation of the molecular basis for the remarkable P450-catalyzed intramolecular C-N bond-forming reaction, which is challenging in synthetic chemistry, to generate the indolactam scaffold. In addition, precursor-directed biosynthesis with the promiscuous P450 enzymes led to the formation of a series of unnatural and novel molecular scaffolds, including a sulfur-substituted indolactam with a different conformation from that of indolactam V.


Assuntos
Sistema Enzimático do Citocromo P-450/metabolismo , Indóis/metabolismo , Lactamas/metabolismo , Toxinas de Lyngbya/biossíntese , Toxinas de Lyngbya/química , Animais , Ciclização , Humanos
10.
Pest Manag Sci ; 77(4): 1607-1615, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32954637

RESUMO

BACKGROUND: Pine wilt disease (PWD) is a plant disease that causes serious damage to pine trees. PWD occurs when the host plant is infected with pinewood nematode (PWN), Bursaphelenchus xylophilus. In this study, a compound with nematicidal activity was isolated from actinomycetes and its efficacy was investigated in vitro. RESULT: We screened and selected Streptomyces sp. 680560, which had nematicidal activity against B. xylophilus. Based on 16S rRNA sequence analysis, it showed 99.93% similarity with Streptomyces blastmyceticus NRRB-5480T . Furthermore, the active compound was isolated and identified as teleocidin B4. Teleocidin B4 at concentrations ranging from 6.25 to 100 µM had low nematicidal activity after 24 and 36 h against adult and stage juveniles (J2) of B. xylophilus, but after 48 h nematicidal activity exceeded 95%. The rate of inhibition of egg hatching for Teleocidin B4 6.25, 12.5, 25, 50, and 100 µM was confirmed to be dose-dependently inhibited after 48 h of treatment. Teleocidin B4 is not only toxic to hatched B. xylophilus, but also affects egg hatching. CONCLUSION: This study was carried out to isolate actinomycete metabolites from pine tree endophytes from various natural environments for control of PWD. A compound with nematicidal activity was isolated from a selected strain and its structure was identified as teleocidin B4. The nematicidal effect of the isolated active substance, teleocidin B4, was confirmed. This is the first report of the effect of teleocidin B4 on B. xylophilus, suggesting its possibility as a PWD control agent. © 2020 Society of Chemical Industry.


Assuntos
Pinus , Streptomyces , Tylenchida , Animais , Toxinas de Lyngbya , Doenças das Plantas , RNA Ribossômico 16S , Xylophilus
11.
Toxins (Basel) ; 12(11)2020 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-33238397

RESUMO

Since 1970s, aplysiatoxins (ATXs), a class of biologically active dermatoxins, were identified from the marine mollusk Stylocheilus longicauda, whilst further research indicated that ATXs were originally metabolized by cyanobacteria. So far, there have been 45 aplysiatoxin derivatives discovered from marine cyanobacteria with various geographies. Recently, we isolated two neo-debromoaplysiatoxins, neo-debromoaplysiatoxin G (1) and neo-debromoaplysiatoxin H (2) from the cyanobacterium Lyngbya sp. collected from the South China Sea. The freeze-dried cyanobacterium was extracted with liquid-liquid extraction of organic solvents, and then was subjected to multiple chromatographies to yield neo-debromoaplysiatoxin G (1) (3.6 mg) and neo-debromoaplysiatoxin H (2) (4.3 mg). They were elucidated with spectroscopic methods. Moreover, the brine shrimp toxicity of the aplysiatoxin derivatives representing differential structural classifications indicated that the debromoaplysiatoxin was the most toxic compound (half inhibitory concentration (IC50) value = 0.34 ± 0.036 µM). While neo-aplysiatoxins (neo-ATXs) did not exhibit apparent brine shrimp toxicity, but showed potent blocking action against potassium channel Kv1.5, likewise, compounds 1 and 2 with IC50 values of 1.79 ± 0.22 µM and 1.46 ± 0.14 µM, respectively. Therefore, much of the current knowledge suggests the ATXs with different structure modifications may modulate multiple cellular signaling processes in animal systems leading to the harmful effects on public health.


Assuntos
Toxinas de Lyngbya/química , Toxinas de Lyngbya/toxicidade , Lyngbya , Bloqueadores dos Canais de Potássio/química , Bloqueadores dos Canais de Potássio/toxicidade , Animais , Artemia/efeitos dos fármacos , Células CHO , Cricetulus , Canal de Potássio Kv1.5/antagonistas & inibidores , Canal de Potássio Kv1.5/genética , Canal de Potássio Kv1.5/fisiologia
12.
Bioorg Med Chem Lett ; 30(24): 127657, 2020 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-33130291

RESUMO

Debromoaplysiatoxin (DAT) is a potent protein kinase C (PKC) activator with tumor-promoting and pro-inflammatory activities. Irie and colleagues have found that 10-methyl-aplog-1 (1), a simplified analog of DAT, has strong anti-proliferative activity against several cancer cell lines with few adverse effects. Therefore, 1 is a potential lead compound for cancer therapy. We synthesized a new derivative 2 which has a naphthalene ring at the side chain terminal position instead of a benzene ring, to increase CH/π interactions with Pro-241 of the PKCδ-C1B domain. Based on the synthetic route of 1, 2 was convergently synthesized in 26 linear steps from 6-hydroxy-1-naphthoic acid with an overall yield of 0.18%. Although the anti-proliferative activity of 2 was more potent than that of 1, the binding potency of 2 to the PKCδ-C1B domain did not exceed that of 1. Molecular dynamics simulation indicated the capability of 2 to simultaneously form hydrogen bonds and CH/π interactions with the PKCδ-C1B domain. Focusing on the hydrogen bonds, their geometry in the binding modes involving the CH/π interactions seemed to be sub-optimal, which may explain the slightly lower affinity of 2 compared to 1. This study could be of help in optimizing such interactions and synthesizing a promising lead cancer compound.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Toxinas de Lyngbya/química , Toxinas de Lyngbya/farmacologia , Antineoplásicos/síntese química , Linhagem Celular Tumoral , Humanos , Toxinas de Lyngbya/síntese química , Modelos Moleculares , Neoplasias/tratamento farmacológico , Relação Estrutura-Atividade
13.
Molecules ; 25(3)2020 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-31978978

RESUMO

A new aplysiatoxin derivative, neo-aplysiatoxin A (1), along with seven known compounds, neo-debromoaplysiatoxin A (2), dolastatin 3 (3), lyngbic acid (4), malyngamide M (5), hermitamide A (6), (-)-loliolide (7), and (+)-epiloliolide (8), was isolated from the Okinawan cyanobacterium Moorea producens. Their structures were elucidated on the basis of spectroscopic data, including high-resolution mass spectrometry and nuclear magnetic resonance. The compounds were evaluated for cytotoxic and diatom growth inhibition activities.


Assuntos
Cianobactérias/metabolismo , Toxinas de Lyngbya/metabolismo , Depsipeptídeos/metabolismo , Ácidos Graxos Monoinsaturados/metabolismo , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Estrutura Molecular
14.
Nat Prod Res ; 34(15): 2151-2156, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30835553

RESUMO

Neo-debromoaplysiatoxin C (1), a new member of the aplysiatoxin family, was isolated from the marine cyanobacterium Lyngbya sp. The structure of 1 was elucidated based on spectroscopic data, and its stereochemistry was determined from NOESY spectrum and biosynthetic considerations. This new compound presents an intriguing 10-membered lactone ring skeleton derived from debromoaplysiatoxin by structural rearrangement, which is the first example in the aplysiatoxin family. Its biological properties were evaluated for cytotoxicity, PKCδ activation and inhibitory effects on potassium channel.


Assuntos
Cianobactérias/química , Toxinas de Lyngbya/química , Citotoxinas/farmacologia , Lactonas/química , Lactonas/farmacologia , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Bloqueadores dos Canais de Potássio/farmacologia , Proteína Quinase C-delta/efeitos dos fármacos , Alga Marinha/química
15.
Nat Prod Rep ; 37(3): 425-463, 2020 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-31650156

RESUMO

Covering: up to July 2019 Terpene synthases (TSs) are responsible for generating much of the structural diversity found in the superfamily of terpenoid natural products. These elegant enzymes mediate complex carbocation-based cyclization and rearrangement cascades with a variety of electron-rich linear and cyclic substrates. For decades, two main classes of TSs, divided by how they generate the reaction-triggering initial carbocation, have dominated the field of terpene enzymology. Recently, several novel and unconventional TSs that perform TS-like reactions but do not resemble canonical TSs in sequence or structure have been discovered. In this review, we identify 12 families of non-canonical TSs and examine their sequences, structures, functions, and proposed mechanisms. Nature provides a wide diversity of enzymes, including prenyltransferases, methyltransferases, P450s, and NAD+-dependent dehydrogenases, as well as completely new enzymes, that utilize distinctive reaction mechanisms for TS chemistry. These unique non-canonical TSs provide immense opportunities to understand how nature evolved different tools for terpene biosynthesis by structural and mechanistic characterization while affording new probes for the discovery of novel terpenoid natural products and gene clusters via genome mining. With every new discovery, the dualistic paradigm of TSs is contradicted and the field of terpene chemistry and enzymology continues to expand.


Assuntos
Alquil e Aril Transferases/química , Alquil e Aril Transferases/metabolismo , Aminofenóis/química , Aminofenóis/metabolismo , Canabinoides/química , Canabinoides/metabolismo , Ciclização , Sistema Enzimático do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/metabolismo , Dimetilaliltranstransferase/química , Dimetilaliltranstransferase/metabolismo , Iridoides/química , Iridoides/metabolismo , Toxinas de Lyngbya/química , Toxinas de Lyngbya/metabolismo , Estrutura Molecular , Fenazinas/química , Fenazinas/metabolismo , Compostos Policíclicos/química , Compostos Policíclicos/metabolismo , Conformação Proteica , Terpenos/química , Terpenos/metabolismo
16.
Angew Chem Int Ed Engl ; 59(10): 3988-3993, 2020 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-31886618

RESUMO

C-S bond formation reactions are widely distributed in the biosynthesis of biologically active molecules, and thus have received much attention over the past decades. Herein, we report intramolecular C-S bond formation by a P450 monooxygenase, TleB, which normally catalyzes a C-N bond formation in teleocidin biosynthesis. Based on the proposed reaction mechanism of TleB, a thiol-substituted substrate analogue was synthesized and tested in the enzyme reaction, which afforded the unprecedented sulfur-containing thio-indolactam V, in addition to an unusual indole-fused 6/5/8-tricyclic product whose structure was determined by the crystalline sponge method. Interestingly, conformational analysis revealed that the SOFA conformation is stable in thio-indolactam V, in sharp contrast to the major TWIST form in indolactam V, resulting in differences in their biological activities.


Assuntos
Sistema Enzimático do Citocromo P-450/metabolismo , Toxinas de Lyngbya/biossíntese , Biocatálise , Cristalografia por Raios X , Sistema Enzimático do Citocromo P-450/química , Toxinas de Lyngbya/química , Conformação Molecular , Simulação de Dinâmica Molecular , Pseudomonas putida/enzimologia , Especificidade por Substrato
17.
ACS Synth Biol ; 9(1): 63-75, 2020 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-31846576

RESUMO

Cyanobacteria are prolific producers of natural products, and genome mining has shown that many orphan biosynthetic gene clusters can be found in sequenced cyanobacterial genomes. New tools and methodologies are required to investigate these biosynthetic gene clusters, and here we present the use of Anabaena sp. strain PCC 7120 as a host for combinatorial biosynthesis of natural products using the indolactam natural products (lyngbyatoxin A, pendolmycin, and teleocidin B-4) as a test case. We were able to successfully produce all three compounds using codon optimized genes from Actinobacteria. We also introduce a new plasmid backbone based on the native Anabaena 7120 plasmid pCC7120ζ and show that production of teleocidin B-4 can be accomplished using a two-plasmid system, which can be introduced by coconjugation.


Assuntos
Alcaloides/biossíntese , Anabaena/genética , Anabaena/metabolismo , Produtos Biológicos/metabolismo , Toxinas de Lyngbya/biossíntese , Engenharia Metabólica/métodos , Proteínas de Bactérias/genética , Códon/genética , Genes Bacterianos , Família Multigênica , Plasmídeos/genética
18.
Harmful Algae ; 90: 101700, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31806161

RESUMO

Mass spectrometric methods for the quantitative and qualitative analyses of algal biotoxins are often complicated by co-eluting compounds that present analytically as interferences. This issue is particularly critical for organic polyamines, where co-eluting materials can suppress the formation of cations during electrospray ionization. Here we present an extraction procedure designed specifically to overcome matrix-derived ion suppression of algal toxins in samples of Lyngbya wollei, a filamentous benthic algae known to produce several saxitoxin analogues. Lyngbya wollei samples were collected from a large, persistent harmful algal bloom in Lake Wateree, SC. Six known Lyngbya wollei-specific toxins (LWT1-6) were successfully resolved and quantified against saxitoxin using hydrophilic interaction liquid chromatography coupled with triple quadrupole and quadrupole time-of-flight mass spectrometry. The parent ions [M2+ - H+]+ were observed for LWTs 1-6 and the [M]2+ ion was observed for LWT5. High resolution mass spectra and unique fragmentation ions were obtained for LWTs 1-6. A dilution factor of 50 resulted in a linear calibration of saxitoxin in the algae matrix. Ion suppression was resolved by sample dilution, which led to linear, positive correlations between peak area and mass of the extracted sample (R2 > 0.96). Optimized sample extraction method and instrument parameters are presented.


Assuntos
Cianobactérias , Toxinas de Lyngbya , Proliferação Nociva de Algas , Espectrometria de Massas , Saxitoxina
19.
Mar Drugs ; 17(12)2019 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-31766406

RESUMO

A pair of stereoisomers possessing novel structures with 6/6/5 fused-ring systems, neo-debromoaplysiatoxin E (1) and neo-debromoaplysiatoxin F (2), were isolated from the marine cyanobacterium Lyngbya sp. Their structures were elucidated using various spectroscopic techniques including high resolution electrospray ionization mass spectroscopy (HRESIMS) and nuclear magnetic resonance (NMR). The absolute stereochemistry was determined by calculated electronic circular dichroism (ECD) and gauge-independent atomic orbital (GIAO) NMR shift calculation followed by DP4+ analysis. Significantly, this is the first report on aplysiatoxin derivatives with different absolute configurations at C9-C12 (1: 9S, 10R, 11S, 12S; 2: 9R, 10S, 11R, 12R). Compounds 1 and 2 exhibited potent blocking activities against Kv1.5 with IC50 values of 1.22 ± 0.22 µM and 2.85 ± 0.29 µM, respectively.


Assuntos
Organismos Aquáticos/química , Cianobactérias/química , Canal de Potássio Kv1.5/antagonistas & inibidores , Toxinas de Lyngbya/farmacologia , Animais , Células CHO , Dicroísmo Circular , Cricetulus , Canal de Potássio Kv1.5/metabolismo , Toxinas de Lyngbya/química , Toxinas de Lyngbya/isolamento & purificação , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Estereoisomerismo
20.
Toxins (Basel) ; 11(6)2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31234410

RESUMO

Cyanobacteria have been shown to produce a number of bioactive compounds, including toxins. Some bioactive compounds obtained from a marine cyanobacterium Moorea producens (formerly Lyngbya majuscula) have been recognized as drug leads; one of these compounds is aplysiatoxin. We have isolated various aplysiatoxin derivatives from a M. producens sample obtained from the Okinawan coastal area. The frozen sample was extracted with organic solvents. The ethyl acetate layer was obtained from the crude extracts via liquid-liquid partitioning, then separated by HPLC using a reversed-phase column. Finally, 1.1 mg of the compound was isolated. The chemical structure of the isolated compound was elucidated with spectroscopic methods, using HR-MS and 1D and 2D NMR techniques, and was revealed to be oscillatoxin I, a new member of the aplysiatoxin family. Oscillatoxin I showed cytotoxicity against the L1210 mouse lymphoma cell line and diatom growth-inhibition activity against the marine diatom Nitzschia amabilis.


Assuntos
Toxinas Bacterianas/isolamento & purificação , Cianobactérias , Toxinas de Lyngbya/isolamento & purificação , Animais , Toxinas Bacterianas/toxicidade , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Diatomáceas/efeitos dos fármacos , Diatomáceas/crescimento & desenvolvimento , Toxinas de Lyngbya/toxicidade , Camundongos
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