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1.
Ann Clin Microbiol Antimicrob ; 11: 10, 2012 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-22549052

RESUMO

BACKGROUND: The global burden of bacterial infections is high and has been further aggravated by increasing resistance to antibiotics. In the search for novel antibacterials, three medicinal plants: Peperomia vulcanica, Peperomia fernandopoioana (Piperaceae) and Scleria striatinux (Cyperaceae), were investigated for antibacterial activity and toxicity. METHODS: Crude extracts of these plants were tested by the disc diffusion method against six bacterial test organisms followed by bio-assay guided fractionation, isolation and testing of pure compounds. The minimum inhibitory (MIC) and minimum bactericidal (MBC) concentrations were measured by the microdilution method. The acute toxicity of the active extracts and cytotoxicity of the active compound were performed in mice and mammalian cells, respectively. RESULTS: The diameter of the zones of inhibition (DZI) of the extracts ranged from 7-13 mm on Escherichia coli and Staphylococcus aureus of which the methylene chloride:methanol [1:1] extract of Scleria striatinux recorded the highest activity (DZI = 13 mm). Twenty-nine pure compounds were screened and one, Okundoperoxide, isolated from S. striatinux, recorded a DZI ranging from 10-19 mm on S. aureus. The MICs and MBCs indicated that the Peperomias had broad-spectrum bacteriostatic activity. Toxicity tests showed that Okundoperoxide may have a low risk of toxicity with an LC50 of 46.88 µg/mL. CONCLUSIONS: The antibacterial activity of these plants supports their use in traditional medicine. The pure compound, Okundoperoxide, may yield new antibacterial lead compounds following medicinal chemistry exploration.


Assuntos
Antibacterianos/farmacologia , Cyperaceae/química , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Peperomia/química , Extratos Vegetais/farmacologia , Animais , Antibacterianos/isolamento & purificação , Antibacterianos/toxicidade , Bioensaio , Linhagem Celular , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão , Descoberta de Drogas , Farmacorresistência Bacteriana Múltipla , Feminino , Bactérias Gram-Negativas/crescimento & desenvolvimento , Bactérias Gram-Positivas/crescimento & desenvolvimento , Dose Letal Mediana , Masculino , Metanol , Cloreto de Metileno , Camundongos , Camundongos Endogâmicos BALB C , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/toxicidade , Sesquiterpenos/isolamento & purificação , Sesquiterpenos/toxicidade
2.
Protein Expr Purif ; 63(1): 33-9, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18824106

RESUMO

UapA, the uric acid-xanthine permease from the filamentous ascomycete Aspergillus nidulans, is one of the most thoroughly characterized purine/H(+) transporters in eukaryotes. Detailed studies have addressed its regulation of expression, at both the transcriptional and post-translational levels, in response to physiological and developmental signals. An extensive kinetic profile towards a plethora of purines and mutational analyses have established models on how UapA recognizes the purine ring and revealed specific amino acid residues involved in proper folding, topogenesis, function and specificity. The present work describes for the first time the purification of the UapA transporter of A. nidulans through overexpression via the strong, ethanol-inducible, glucose-repressible, alcA promoter. Purification, almost to homogeneity, was achieved by Ni(2+) affinity chromatography using a functional His-tagged UapA protein version. It is subsequently shown, by Circular Dichroism (CD) spectroscopy, that the purified protein is structured with a high alpha-helical content, as expected from the in silico predictions. The result of this work opens the way for further, analytical and biochemical studies on UapA at the protein level.


Assuntos
Aspergillus nidulans/enzimologia , Proteínas Fúngicas , Proteínas de Membrana Transportadoras , Cromatografia de Afinidade , Cromatografia em Gel , Dicroísmo Circular , Clonagem Molecular , Etanol/metabolismo , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/isolamento & purificação , Proteínas Fúngicas/metabolismo , Glucose/metabolismo , Immunoblotting , Proteínas de Membrana Transportadoras/química , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/isolamento & purificação , Proteínas de Membrana Transportadoras/metabolismo , Regiões Promotoras Genéticas , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas Recombinantes de Fusão/metabolismo , Análise Espectral
3.
Fungal Genet Biol ; 44(7): 627-40, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17126042

RESUMO

In this article we study the cellular expression of UapA and AzgA, the two major purine transporters of Aspergillus nidulans, by constructing strains expressing, from their native promoters, fully functional fluorescent (UapA-sGFP, AzgA-sGFP) or immunological (UapA-His) chimeric transporters. Epifluorescence microscopy and immunodetection showed that under different physiological conditions and during Aspergillus development: (i) UapA and AzgA expression in the plasma membrane becomes evident early during germination and remains at a significant basal level in mycelium, (ii) Neither of the two transporters is expressed in the stalk, the vesicle, the phialides and the conidiospores, but surprisingly, UapA is specifically and strongly expressed in the periphery of metulae, (iii) Both transporters are expressed in ascogenous hyphae and in hülle cells but not in cleistothecia or ascospores, (iv) Purine induction leads to approximately 4-fold increase in UapA and AzgA protein content in mycelium, compatible with an analogous increase at the transcriptional level, (v) Ammonium leads to removal of UapA, but not of AzgA, from the plasma membrane by sorting of the protein to the vacuole.


Assuntos
Aspergillus nidulans/metabolismo , Proteínas de Transporte/metabolismo , Proteínas Fúngicas/metabolismo , Purinas/metabolismo , Adenina/metabolismo , Aspergillus nidulans/crescimento & desenvolvimento , Transporte Biológico , Membrana Celular/metabolismo , Meios de Cultura , Regulação para Baixo , Guanina/metabolismo , Hifas/metabolismo , Hipoxantina/metabolismo , Immunoblotting , Proteínas de Membrana Transportadoras/metabolismo , Compostos de Amônio Quaternário
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