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1.
Biochim Biophys Acta Biomembr ; 1861(7): 1375-1387, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-30926365

RESUMO

Infections caused by Gram-negative bacteria, Escherichia coli and Pseudomonas aeruginosa foremost among them, constitute a major worldwide health problem. Bioinformatics methodologies are being used to rationally design new antimicrobial peptides, a potential alternative for treating these infections. One of the algorithms used to develop antimicrobial peptides is the Joker, which was used to design the peptide PaDBS1R6. This study evaluates the antibacterial activities of PaDBS1R6 in vitro and in vivo, characterizes the peptide interaction to target membranes, and investigates the PaDBS1R6 structure in contact with mimetic vesicles. Moreover, we demonstrate that PaDBS1R6 exhibits selective antimicrobial activity against Gram-negative bacteria. In the presence of negatively charged and zwitterionic lipids the structural arrangement of PaDBS1R6 transits from random coil to α-helix, as characterized by circular dichroism. The tertiary structure of PaDBS1R6 was determined by NMR in zwitterionic dodecylphosphocholine (DPC) micelles. In conclusion, PaDBS1R6 is a candidate for the treatment of nosocomial infections caused by Gram-negative bacteria, as template for producing other antimicrobial agents.


Assuntos
Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bactérias Gram-Negativas/efeitos dos fármacos , Animais , Camundongos , Camundongos Endogâmicos C57BL , Testes de Sensibilidade Microbiana
2.
Biochim Biophys Acta Biomembr ; 1861(1): 178-190, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30463701

RESUMO

Antimicrobial peptides (AMPs) are promising candidates for the development of future antibiotics. In an attempt to increase the efficacy of therapeutic AMPs, computer-based design methods appear as a reliable strategy. In this study, we evaluated the antimicrobial efficiency and mechanism of action of a novel designed AMP named PaDBS1R1, previously designed by means of the Joker algorithm, using a fragment of the ribosomal protein L39E from the archaeon Pyrobaculum aerophilum as a template. PaDBS1R1 displayed low micromolar broad-spectrum antimicrobial activity against Gram-negative (MIC of 1.5 µM) and Gram-positive (MIC of 3 µM) bacteria, including carbapenem-resistant Klebsiella pneumoniae (MIC of 6.25 µM) and methicillin-resistant Staphylococcus aureus (MIC of 12.5 µM), without cytotoxicity towards HEK-293 cells. In addition, membrane permeabilization and depolarization assays, combined with time-kill studies and FEG-SEM imaging, indicated a fast membrane permeation and further leakage of intracellular content. Biophysical studies with lipid vesicles show a preference of PaDBS1R1 for Gram-negative bacteria-like membranes. We investigated the three-dimensional structure of PaDBS1R1 by CD and NMR analyses. Our results suggest that PaDBS1R1 adopts an amphipathic α-helix upon interacting with hydrophobic environments, after an initial electrostatic interaction with negative charges, suggesting a membrane lytic effect. This study reveals that PaDBS1R1 has potential application in antibiotic therapy.


Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Membrana Celular/efeitos dos fármacos , Antibacterianos/farmacologia , Membrana Celular/metabolismo , Permeabilidade da Membrana Celular/efeitos dos fármacos , Dicroísmo Circular , Bactérias Gram-Negativas , Células HEK293 , Humanos , Interações Hidrofóbicas e Hidrofílicas , Luz , Lipídeos/química , Espectroscopia de Ressonância Magnética , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Micelas , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de Varredura , Conformação Proteica em alfa-Hélice , Espalhamento de Radiação
3.
Nat Commun ; 9(1): 1490, 2018 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-29662055

RESUMO

Plants are extensively used in traditional medicine, and several plant antimicrobial peptides have been described as potential alternatives to conventional antibiotics. However, after more than four decades of research no plant antimicrobial peptide is currently used for treating bacterial infections, due to their length, post-translational modifications or  high dose requirement for a therapeutic effect . Here we report the design of antimicrobial peptides derived from a guava glycine-rich peptide using a genetic algorithm. This approach yields guavanin peptides, arginine-rich α-helical peptides that possess an unusual hydrophobic counterpart mainly composed of tyrosine residues. Guavanin 2 is characterized as a prototype peptide in terms of structure and activity. Nuclear magnetic resonance analysis indicates that the peptide adopts an α-helical structure in hydrophobic environments. Guavanin 2 is bactericidal at low concentrations, causing membrane disruption and triggering hyperpolarization. This computational approach for the exploration of natural products could be used to design effective peptide antibiotics.


Assuntos
Antibacterianos/química , Peptídeos Catiônicos Antimicrobianos/química , Proteínas de Plantas/química , Infecções por Pseudomonas/tratamento farmacológico , Pseudomonas aeruginosa/efeitos dos fármacos , Psidium/química , Algoritmos , Sequência de Aminoácidos , Animais , Antibacterianos/biossíntese , Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/biossíntese , Peptídeos Catiônicos Antimicrobianos/genética , Peptídeos Catiônicos Antimicrobianos/farmacologia , Membrana Celular/química , Membrana Celular/efeitos dos fármacos , Técnicas de Química Combinatória , Desenho de Fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Interações Hidrofóbicas e Hidrofílicas , Camundongos , Modelos Moleculares , Ressonância Magnética Nuclear Biomolecular , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Proteínas de Plantas/farmacologia , Estrutura Secundária de Proteína , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/química , Pseudomonas aeruginosa/crescimento & desenvolvimento , Psidium/metabolismo , Pele/efeitos dos fármacos , Pele/microbiologia , Relação Estrutura-Atividade
4.
Sci Rep ; 6: 27128, 2016 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-27292548

RESUMO

Clavanins is a class of peptides (23aa) histidine-rich, free of post-translational modifications. Clavanins have been studied largely for their ability to disrupt bacterial membranes. In the present study, the interaction of clavanin A with membranes was assessed by dynamic light scattering, zeta potential and permeabilization assays. We observed through those assays that clavanin A lysis bacterial cells at concentrations corresponding to its MIC. Further, the structure and function of clavanin A was investigated. To better understand how clavanin interacted with bacteria, its NMR structure was elucidated. The solution state NMR structure of clavanin A in the presence of TFE-d3 indicated an α-helical conformation. Secondary structures, based on circular dichroism measurements in anionic sodium dodecyl sulfate (SDS) and TFE (2,2,2-trifluorethanol), in silico lipid-peptide docking and molecular simulations with lipids DPPC and DOPC revealed that clavanin A can adopt a variety of folds, possibly influencing its different functions. Microcalorimetry assays revealed that clavanin A was capable of discriminating between different lipids. Finally, clavanin A was found to eradicate bacterial biofilms representing a previously unrecognized function.


Assuntos
Bactérias/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Proteínas Sanguíneas/química , Bicamadas Lipídicas/metabolismo , Urocordados/metabolismo , Animais , Fenômenos Fisiológicos Bacterianos/efeitos dos fármacos , Proteínas Sanguíneas/farmacologia , Membrana Celular/efeitos dos fármacos , Dicroísmo Circular , Difusão Dinâmica da Luz , Hemócitos/química , Hemócitos/metabolismo , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Estrutura Secundária de Proteína , Urocordados/química
5.
Protein Pept Lett ; 22(8): 719-26, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26059694

RESUMO

Antimicrobial peptides (AMPs) appear as a promising therapeutic candidate against multiresistant pathogens, because they are able to kill microorganisms and have low toxicity of resistance cells. Hylin a1 (Hy-a1, IFGAILPLALGALKNLIK-NH2) is a peptide extracted from the skin secretion of the frog Hypsiboas albopunctatus, which displays antimicrobial and hemolytic activities. We report here structural studies of Hy-a1 using different techniques such as fluorescence, CD and NMR. Our data showed that Hy-a1 acquires a well defined amphipathic α-helix when interacting with a membrane-like environment. Furthermore, Hy-a1 presented different affinity when compared to membranes of zwitterionic or anionic lipid composition. Finally, we proposed a molecular interaction model of this peptide with micelles.


Assuntos
Proteínas de Anfíbios/química , Proteínas de Anfíbios/metabolismo , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/metabolismo , Membranas Artificiais , Micelas , Animais , Modelos Moleculares , Ressonância Magnética Nuclear Biomolecular , Porosidade , Ranidae
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