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1.
São Paulo; s.n; s.n; 2016. 201 p. tab, graf, ilus.
Tese em Português | LILACS | ID: biblio-847477

RESUMO

Esticolisinas I e II, citolisinas purificadas da anêmona marinha Stichodactyla helianthus, agem lisando membranas biológicas e modelo. O mecanismo de ação proposto consiste na formação de um poro toroidal com o envolvimento do domínio N-terminal. Diferentes aspectos da interação entre peptídeos derivados do N-terminal das toxinas (StI1-31 and StI12-31 SELAGTIIDGASLTFEVLDKVLGELGKVSRK, e StII1-30 and StII11-30 ALAGTIIAGASLTFQVLDKVLEELGKVSRK) com membranas modelo - micelas e bicamadas - foram estudados com o objetivo de contribuir para a elucidação do mecanismo de ação das toxinas em nível molecular. O emprego dos peptídeos teve como base a hipótese de que fragmentos proteicos podem ser capazes de mimetizar a estrutura e atividade das proteínas inteiras. O análogo contendo o aminoácido paramagnético TOAC (N-TOAC-StII11-30) também foi estudado. Estudos conformacionais foram realizados empregando-se as técnicas espectroscópicas de dicroísmo circular (CD), ressonância paramagnética eletrônica (EPR) e fluorescência. Foram ainda realizados estudos de predição de estrutura e modelagem molecular. Espectros de CD mostraram que os peptídeos adquirem conformação em α-hélice ao interagir com membranas modelo, de acordo com a conformação observada nessa região para as toxinas. Variando a composição lipídica das membranas modelo estudadas, foi possível investigar a contribuição de forças eletrostáticas de de interações hidrofóbicas para a ligação do peptídeo. Ensaios de supressão de fluorescência de lípidos contendo grupamentos fluorescentes em diferentes posições pelo resíduo paramagnético TOAC e espectros de ressonância paramagnética eletrônica (EPR) permitiram localizar o resíduo TOAC na interface membrana-água, corroborando o modelo proposto do poro toroidal. A análise dos espectros de CD e EPR também permitiu obter as constantes de ligação dos peptídeos com micelas e bicamadas. Os peptídeos também foram capazes de mimetizar as toxinas do ponto de vista funcional, como mostrado por testes de vazamento de carboxifluoresceína e atividade hemolítica. Peptídeos curtos, contendo partes da sequência de StII1-30, sintetizados com o objetivo de examinar uma eventual atividade antimicrobiana, demonstraram baixa atividade, bem como ausência de atividade hemolítica e de toxicidade para células humanas


Sticholysins I and II, cytolysins purified from the sea anemone Stichodactyla helianthus, act by lysing biological and model membranes. The proposed mechanism of action consists in the formation of a toroidal pore with the involvement of the N-terminal domain [1]. Different aspects of the interaction between peptides from the toxins' N-termini (StI1-31 and StI12-31 SELAGTIIDGASLTFEVLDKVLGELGKVSRK, and StII1-30 and StII11-30 ALAGTIIAGASLTFQVLDKVLEELGKVSRK) and model membranes - micelles and bilayers - have been studied to contribute to the elucidation of the toxins mechanism of action at the molecular level. The use of peptides was based on the hypothesis that potein fragments can eventually mimic the structure and activity of the whole protein. An analogue containing the paramagnetic amino acid TOAC (N-TOAC-StII11-30) was also studied. Conformational studies were performed making use of the spectroscopic techniques circular dichroism (CD), electron paramagnetic resonance (EPR), and fluorescence. Studies of structure prediction and molecular modeling were also performed. CD spectra showed that the peptides acquired α-helical conformation upon interaction with model lipid membranes, in agreement with the conformation found for these segments in the whole proteins. Making use of membranes of variable lipid composition, it was possible to assess the contribution of electrostatic and hydrophobic interactions for peptide binding. Fluorescence quenching of labeled lipids by paramagnetic TOAC and EPR spectra allowed us to locate the TOAC residue at the membrane-water interface, corroborating the proposed model of the toroidal pore. The CD and EPR studies also allowed us to obtain the binding constants for the peptide-micelle and peptide-bilayer interaction. The peptides were also capable of mimicking the toxins function, as shown by assays of carboxyfluorescein leakage and hemolytic activity. Short peptides containing parts of StII1-30's sequence were synthesized with the aim of testing their antimicrobial activity. The peptides displayed low antimicrobial activity, as well as lack of hemolytic activity and toxicity against human cells


Assuntos
Espectroscopia de Ressonância de Spin Eletrônica/métodos , Peptídeos/análise , Análise Espectral/métodos , Infecções Bacterianas/prevenção & controle , Dicroísmo Circular/instrumentação , Modelos Estruturais , Relação Estrutura-Atividade , Tomografia Computadorizada Espiral
2.
Indian J Exp Biol ; 2015 Jun; 53(6): 380-387
Artigo em Inglês | IMSEAR | ID: sea-158514

RESUMO

Melanins are enigmatic pigments produced by a wide variety of microorganisms including bacteria and fungi. Here, we have isolated and characterized extracellular melanin from mushroom fungus, Schizophyllum commune. The extracellular dark pigment produced by the broth culture of S. commune, after 21 days of incubation was recovered by hot acid-alkali treatment. The melanin nature of the pigment was characterized by biochemical tests and further, confirmed by UV, IR, EPR, NMR and MALDI-TOF Mass Spectra. Extracellular melanin, at 100 µg/ml, showed significant antibacterial activity against Escherichia coli, Bacillus subtilis, Klebsiella pneumoniae and Pseudomonas fluorescens and antifungal activity against Trichophyton simii and T. rubrum. At a concentration of 50 µg/ml, melanin showed high free radical scavenging activity of DPPH (2,2-diphenyl-1-picrylhydrazyl) indicating its antioxidant potential. It showed concentration dependent inhibition of cell proliferation of Human Epidermoid Larynx Carcinoma Cell Line (HEP-2). This study has demonstrated characterization of melanin from basidiomycetes mushroom fungus, Schizophyllum commune and its applications.


Assuntos
Antibacterianos/farmacologia , Antifúngicos/farmacologia , Antineoplásicos/farmacologia , Antioxidantes/farmacocinética , Basidiomycota/química , Espectroscopia de Ressonância de Spin Eletrônica/métodos , Fungos , Melaninas/biossíntese , Melaninas/isolamento & purificação , Melaninas/farmacocinética , Melaninas/metabolismo , Schizophyllum/química , Schizophyllum/classificação
3.
Electron. j. biotechnol ; 7(3): 11-12, Dec. 2004. ilus, tab, graf
Artigo em Inglês | LILACS | ID: lil-448768

RESUMO

This preliminary work shows ESR (Electron Spin Resonance) can be used to detect biofilms, particularly from Fe-metabolising bacteria. A film was detected by ESR as early as 1 day, hence possibly more sensitively than by fluorescent methods. Films can probably be detected as early as one hour. Spectra contain a very broad peak at g=2.13, probably due to ferrihydrite. Results of field experiments from streams and ponds in New Zealand and Japan, particularly the Minoh River, showed a general increase of ferrihydrite with time. Loss by exfoliation was later than 20 days. The rate of accumulation was faster in a nutrient-rich stagnant pond. Hematite (g=4.3) was often observed, magnetite (g=9) once, and usually small amounts of a common bacterial decay product. The latter was detected for at least 18 months film storage. ESR is a particularly good tool for observing the growth of oxic biofilms containing Fe-metabolising bacteria, and should be just as sensitive for observing Mn-metabolising bacteria in reducing conditions.


Assuntos
Bactérias , Biofilmes , Espectroscopia de Ressonância de Spin Eletrônica/métodos , Compostos Férricos , Óxido Ferroso-Férrico
4.
Rev. chil. radiol ; 6(4): 141-7, 2000. ilus
Artigo em Espanhol | LILACS | ID: lil-295369

RESUMO

Los avances tecnológicos de los últimos años en la resonancia magnética (RM) y la utilización de medio de contraste paramagnético han significado un gran impacto en la imagenología mamaria, especialmente en la evaluación preoperatoria del cáncer y en el seguimiento de los pacientes tratados por el cáncer de mama. En este artículo se resumen los aspectos técnicos de la realización de una RM mamaria, se detallan las principales indicaciones y se mencionan las futuras aplicaciones posibles


Assuntos
Humanos , Feminino , Neoplasias da Mama/diagnóstico , Espectroscopia de Ressonância de Spin Eletrônica/métodos , Implantes de Mama/efeitos adversos , Neoplasias da Mama , Quimioterapia Adjuvante , Espectroscopia de Ressonância de Spin Eletrônica , Mamografia , Sensibilidade e Especificidade
5.
Indian J Biochem Biophys ; 1992 Jun; 29(3): 245-50
Artigo em Inglês | IMSEAR | ID: sea-27917

RESUMO

X-band electron paramagnetic resonance (epr) spectra of the binary systems, BSA-copper(II) (1:1 and 2:1), and the ternary systems, BSA-Cu(II)-aminoacid (1:1:1), are described. In the binary system, two distinct epr features have been observed. One of the features (towards the low pH), showing broad and overlapping epr signals, has been attributed to non-specific bonding of copper(II) to the albumin and other feature (towards higher pH), showing sharp intense epr signals, has been attributed to the specific bonding. The change from non-specific to specific binding is favoured by increase in pH as well as by increase in protein concentration. Specific binding of copper(II) in BSA-Cu(II) has been suggested to be similar to that in HSA-Cu(II). Spectra of BSA-Cu(II)-aminoacid (1:1:1) show simultaneous presence of binary BSA-Cu(II) and ternary BSA-Cu(II)-aminoacid.


Assuntos
Aminoácidos/metabolismo , Sítios de Ligação , Cobre/metabolismo , Espectroscopia de Ressonância de Spin Eletrônica/métodos , Humanos , Conformação Proteica , Albumina Sérica/química , Soroalbumina Bovina/química
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