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1.
Artículo en Inglés | MEDLINE | ID: mdl-38926900

RESUMEN

OBJECTIVE: Because 66/68 joint counts are not always performed in routine care, we aimed to determine which of the modified 28-joint disease activity index for psoriatic arthritis (DAPSA28) or 28-joint disease activity score with C-reactive protein (DAS28-CRP) should be preferred for monitoring disease activity in psoriatic arthritis (PsA) when the original DAPSA (66/68 joints) is not available. METHODS: Prospectively collected real-world data of European bionaive patients with PsA initiating a first tumor necrosis factor inhibitor were pooled. Remission and response status were evaluated at 6 months by remission (DAPSA ≤ 4, DAPSA28 ≤ 4, and DAS28-CRP < 2.6), response (75% improvement for DAPSA and DAPSA28), and combined EULAR good/moderate responses for DAS28-CRP. Logistic regression analyses on multiple imputed data were used to identify baseline predictors. RESULTS: Remission and response cohorts included 3,159 and 1,866 patients, respectively. The 6-month proportions achieving remission/response were DAPSA (27%/44%), DAPSA28 (28%/44%), and DAS28-CRP (59%/80%). Of 14 possible baseline predictors, 11 predicted both DAPSA and DAPSA28 remission (8 of which also predicted their response, indicated by "*"): longer disease duration*, male sex*, and higher CRP* were positive, whereas older age*, higher body mass index*, patient fatigue*, and global, physician global, health assessment questionnaire score*, and tender and swollen* joint counts were negative predictors. Eight and five of these predicted DAS28-CRP remission and response, respectively. CONCLUSION: In patients with PsA, DAPSA28 should be preferred over DAS28-CRP as a substitute for DAPSA when 66/68 joint counts are not available because of the large overlap in remission and response status and in predictors between DAPSA and DAPSA28.

2.
Biomol NMR Assign ; 17(2): 239-242, 2023 12.
Artículo en Inglés | MEDLINE | ID: mdl-37589820

RESUMEN

Molecular chaperones aid proteins to fold and assemble without modifying their final structure, requiring, in several folding processes, the interplay between members of the Hsp70 and Hsp40 families. Here, we report the NMR chemical shift assignments for 1 H, 15 N, and 13 C nuclei of the backbone and side chains of the J-domain of the class B Hsp40 from Saccharomyces cerevisiae, Sis1, complexed with the C-terminal EEVD motif of Hsp70. The data revealed information on the structure and backbone dynamics that add significantly to the understanding of the J-domain-Hsp70-EEVD mechanism of interaction.


Asunto(s)
Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Humanos , Saccharomyces cerevisiae/metabolismo , Proteínas de Choque Térmico/química , Proteínas de Choque Térmico/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Secuencia de Aminoácidos , Unión Proteica , Resonancia Magnética Nuclear Biomolecular , Proteínas HSP70 de Choque Térmico/química , Proteínas HSP70 de Choque Térmico/metabolismo , Péptidos/química
3.
Biochim Biophys Acta Biomembr ; 1863(1): 183449, 2021 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-32828849

RESUMEN

Studies have suggested that antimicrobial peptides act by different mechanisms, such as micellisation, self-assembly of nanostructures and pore formation on the membrane surface. This work presents an extensive investigation of the membrane interactions of the 14 amino-acid antimicrobial peptide hylaseptin P1-NH2 (HSP1-NH2), derived from the tree-frog Hyla punctata, which has stronger antifungal than antibacterial potential. Biophysical and structural analyses were performed and the correlated results were used to describe in detail the interactions of HSP1-NH2 with zwitterionic and anionic detergent micelles and phospholipid vesicles. HSP1-NH2 presents similar well-defined helical conformations in both zwitterionic and anionic micelles, although NMR spectroscopy revealed important structural differences in the peptide N-terminus. 2H exchange experiments of HSP1-NH2 indicated the insertion of the most N-terminal residues (1-3) in the DPC-d38 micelles. A higher enthalpic contribution was verified for the interaction of the peptide with anionic vesicles in comparison with zwitterionic vesicles. The pore formation ability of HSP1-NH2 (examined by dye release assays) and its effect on the size and surface charge as well as on the lipid acyl chain ordering (evaluated by Fourier-transform infrared spectroscopy) of anionic phospholipid vesicles showed membrane disruption even at low peptide-to-phospholipid ratios, and the effect increases proportionately to the peptide concentration. On the other hand, these biophysical investigations showed that a critical peptide-to-phospholipid ratio around 0.6 is essential for promoting disruption of zwitterionic membranes. In conclusion, this study demonstrates that the binding process of the antimicrobial HSP1-NH2 peptide depends on the membrane composition and peptide concentration.


Asunto(s)
Proteínas Anfibias/química , Membranas Artificiales , Proteínas Citotóxicas Formadoras de Poros/química , Animales , Anuros , Conformación Proteica en Hélice alfa
4.
FASEB J ; 34(1): 365-385, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31914616

RESUMEN

Structural conversion of cellular prion protein (PrPC) into scrapie PrP (PrPSc) and subsequent aggregation are key events associated with the onset of transmissible spongiform encephalopathies (TSEs). Experimental evidence supports the role of nucleic acids (NAs) in assisting this conversion. Here, we asked whether PrP undergoes liquid-liquid phase separation (LLPS) and if this process is modulated by NAs. To this end, two 25-mer DNA aptamers, A1 and A2, were selected against the globular domain of recombinant murine PrP (rPrP90-231) using SELEX methodology. Multiparametric structural analysis of these aptamers revealed that A1 adopts a hairpin conformation. Aptamer binding caused partial unfolding of rPrP90-231 and modulated its ability to undergo LLPS and fibrillate. In fact, although free rPrP90-231 phase separated into large droplets, aptamer binding increased the number of droplets but noticeably reduced their size. Strikingly, a modified A1 aptamer that does not adopt a hairpin structure induced formation of amyloid fibrils on the surface of the droplets. We show here that PrP undergoes LLPS, and that the PrP interaction with NAs modulates phase separation and promotes PrP fibrillation in a NA structure and concentration-dependent manner. These results shed new light on the roles of NAs in PrP misfolding and TSEs.


Asunto(s)
Amiloide/metabolismo , Aptámeros de Nucleótidos/química , Aptámeros de Nucleótidos/metabolismo , Extracción Líquido-Líquido/métodos , Enfermedades por Prión/patología , Proteínas Priónicas/química , Proteínas Priónicas/metabolismo , Animales , Ratones , Conformación de Ácido Nucleico , Enfermedades por Prión/metabolismo , Proteínas Priónicas/aislamiento & purificación , Unión Proteica , Conformación Proteica , Pliegue de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Técnica SELEX de Producción de Aptámeros
5.
Biochim Biophys Acta Biomembr ; 1861(7): 1375-1387, 2019 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-30926365

RESUMEN

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.


Asunto(s)
Antibacterianos/farmacología , Péptidos Catiónicos Antimicrobianos/farmacología , Bacterias Gramnegativas/efectos de los fármacos , Animales , Ratones , Ratones Endogámicos C57BL , Pruebas de Sensibilidad Microbiana
6.
Biochim Biophys Acta Biomembr ; 1861(1): 178-190, 2019 01.
Artículo en Inglés | MEDLINE | ID: mdl-30463701

RESUMEN

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.


Asunto(s)
Péptidos Catiónicos Antimicrobianos/farmacología , Membrana Celular/efectos de los fármacos , Antibacterianos/farmacología , Membrana Celular/metabolismo , Permeabilidad de la Membrana Celular/efectos de los fármacos , Dicroismo Circular , Bacterias Gramnegativas , Células HEK293 , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Luz , Lípidos/química , Espectroscopía de Resonancia Magnética , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Micelas , Pruebas de Sensibilidad Microbiana , Microscopía Electrónica de Rastreo , Conformación Proteica en Hélice alfa , Dispersión de Radiación
7.
Nat Commun ; 9(1): 1490, 2018 04 16.
Artículo en Inglés | MEDLINE | ID: mdl-29662055

RESUMEN

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.


Asunto(s)
Antibacterianos/química , Péptidos Catiónicos Antimicrobianos/química , Proteínas de Plantas/química , Infecciones por Pseudomonas/tratamiento farmacológico , Pseudomonas aeruginosa/efectos de los fármacos , Psidium/química , Algoritmos , Secuencia de Aminoácidos , Animales , Antibacterianos/biosíntesis , Antibacterianos/farmacología , Péptidos Catiónicos Antimicrobianos/biosíntesis , Péptidos Catiónicos Antimicrobianos/genética , Péptidos Catiónicos Antimicrobianos/farmacología , Membrana Celular/química , Membrana Celular/efectos de los fármacos , Técnicas Químicas Combinatorias , Diseño de Fármacos , Escherichia coli/efectos de los fármacos , Escherichia coli/crecimiento & desarrollo , Interacciones Hidrofóbicas e Hidrofílicas , Ratones , Modelos Moleculares , Resonancia Magnética Nuclear Biomolecular , Proteínas de Plantas/biosíntesis , Proteínas de Plantas/genética , Proteínas de Plantas/farmacología , Estructura Secundaria de Proteína , Infecciones por Pseudomonas/microbiología , Pseudomonas aeruginosa/química , Pseudomonas aeruginosa/crecimiento & desarrollo , Psidium/metabolismo , Piel/efectos de los fármacos , Piel/microbiología , Relación Estructura-Actividad
9.
Sci Rep ; 6: 21385, 2016 Feb 26.
Artículo en Inglés | MEDLINE | ID: mdl-26916401

RESUMEN

Due to the growing concern about antibiotic-resistant microbial infections, increasing support has been given to new drug discovery programs. A promising alternative to counter bacterial infections includes the antimicrobial peptides (AMPs), which have emerged as model molecules for rational design strategies. Here we focused on the study of Pa-MAP 1.9, a rationally designed AMP derived from the polar fish Pleuronectes americanus. Pa-MAP 1.9 was active against Gram-negative planktonic bacteria and biofilms, without being cytotoxic to mammalian cells. By using AFM, leakage assays, CD spectroscopy and in silico tools, we found that Pa-MAP 1.9 may be acting both on intracellular targets and on the bacterial surface, also being more efficient at interacting with anionic LUVs mimicking Gram-negative bacterial surface, where this peptide adopts α-helical conformations, than cholesterol-enriched LUVs mimicking mammalian cells. Thus, as bacteria present varied physiological features that favor antibiotic-resistance, Pa-MAP 1.9 could be a promising candidate in the development of tools against infections caused by pathogenic bacteria.


Asunto(s)
Antiinfecciosos/farmacología , Bacterias/efectos de los fármacos , Infecciones Bacterianas/tratamiento farmacológico , Lenguado , Péptidos/farmacología , Animales , Clima Frío , Humanos
10.
Antimicrob Agents Chemother ; 60(1): 580-91, 2016 01.
Artículo en Inglés | MEDLINE | ID: mdl-26574006

RESUMEN

Attempts to isolate novel antimicrobial peptides from microbial sources have been on the rise recently, despite their low efficacy in therapeutic applications. Here, we report identification and characterization of a new efficient antimicrobial peptide from a bacterial strain designated A3 that exhibited highest identity with Paenibacillus ehimensis. Upon purification and subsequent molecular characterization of the antimicrobial peptide, referred to as penisin, we found the peptide to be a bacteriocin-like peptide. Consistent with these results, RAST analysis of the entire genome sequence revealed the presence of a lantibiotic gene cluster containing genes necessary for synthesis and maturation of a lantibiotic. While circular dichroism and one-dimension nuclear magnetic resonance experiments confirmed a random coil structure of the peptide, similar to other known lantibiotics, additional biochemical evidence suggests posttranslational modifications of the core peptide yield six thioether cross-links. The deduced amino acid sequence of the putative biosynthetic gene penA showed approximately 74% similarity with elgicin A and 50% similarity with the lantibiotic paenicidin A. Penisin effectively killed methicillin-resistant Staphylococcus aureus (MRSA) and did not exhibit hemolysis activity. Unlike other lantibiotics, it effectively inhibited the growth of Gram-negative bacteria. Furthermore, 80 mg/kg of body weight of penisin significantly reduced bacterial burden in a mouse thigh infection model and protected BALB/c mice in a bacteremia model entailing infection with Staphylococcus aureus MTCC 96, suggesting that it could be a promising new antimicrobial peptide.


Asunto(s)
Antibacterianos/química , Bacteriocinas/química , Genoma Bacteriano , Paenibacillus/genética , Procesamiento Proteico-Postraduccional , Infecciones Estafilocócicas/tratamiento farmacológico , Secuencia de Aminoácidos , Animales , Antibacterianos/biosíntesis , Antibacterianos/farmacología , Bacteriocinas/biosíntesis , Bacteriocinas/genética , Bacteriocinas/farmacología , Expresión Génica , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Staphylococcus aureus Resistente a Meticilina/crecimiento & desarrollo , Ratones , Ratones Endogámicos BALB C , Datos de Secuencia Molecular , Familia de Multigenes , Paenibacillus/química , Paenibacillus/metabolismo , Estructura Secundaria de Proteína , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Infecciones Estafilocócicas/microbiología , Infecciones Estafilocócicas/mortalidad , Infecciones Estafilocócicas/patología , Análisis de Supervivencia , Resultado del Tratamiento
11.
FEBS Lett ; 589(5): 639-44, 2015 Feb 27.
Artículo en Inglés | MEDLINE | ID: mdl-25639464

RESUMEN

Multifunctional and promiscuous antimicrobial peptides (AMPs) can be used as an efficient strategy to control pathogens. However, little is known about the structural properties of plant promiscuous AMPs without disulfide bonds. CD and NMR were used to elucidate the structure of the promiscuous peptide Cn-AMP1, a disulfide-free peptide isolated from green coconut water. Data here reported shows that peptide structure is transitory and could be different according to the micro-environment. In this regard, Cn-AMP1 showed a random coil in a water environment and an α-helical structure in the presence of SDS-d25 micelles. Moreover, deuterium exchange experiments showed that Gly4, Arg5 and Met9 residues are less accessible to solvent, suggesting that flexibility and cationic charges seem to be essential for Cn-AMP1 multiple activities.


Asunto(s)
Antiinfecciosos/química , Cocos/química , Disulfuros/química , Péptidos/química , Secuencia de Aminoácidos , Dicroismo Circular , Espectroscopía de Resonancia Magnética
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