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1.
Bioorg Med Chem ; 25(7): 2260-2265, 2017 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-28284867

RESUMO

Glycosylation by simple sugars is a drug discovery alternative that has been explored with varying success for enhancing the potency and bioavailability of opioid peptides. Long ago we described two O-glycosides having either ß-Glucose and ß-Galactose of (d-Met2, Pro5)-enkephalinamide showing one of the highest antinociceptive activities known. Here, we report the resynthesis of these two analogs and the preparation of three novel neoglycopeptide derivatives (α-Mannose, ß-Lactose and ß-Cellobiose). Binding studies to cloned zebrafish opioid receptors showed very small differences of affinity between the parent compound and the five glycopeptides thus suggesting that the nature of the carbohydrate moiety plays a minor role in determining the binding mode. Indeed, NMR conformational studies, combined with molecular mechanics calculations, indicated that all glycopeptides present the same major conformation either in solution or membrane-like environment. The evidences provided here highlight the relevance for in vivo activity of the conjugating bond between the peptide and sugar moieties in opioid glycopeptides.


Assuntos
Carboidratos/química , Encefalinas/química , Glicopeptídeos/metabolismo , Receptores Opioides/metabolismo , Animais , Glicopeptídeos/química , Glicosilação , Espectroscopia de Ressonância Magnética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Conformação Proteica , Relação Estrutura-Atividade
2.
Chemistry ; 21(32): 11408-16, 2015 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-26177718

RESUMO

Detection of molecular recognition processes requires robust, specific, and easily implementable sensing methods, especially for screening applications. Here, we propose the difluoroacetamide moiety (an acetamide bioisoster) as a novel tag for detecting by NMR analysis those glycan-protein interactions that involve N-acetylated sugars. Although difluoroacetamide has been used previously as a substituent in medicinal chemistry, here we employ it as a specific sensor to monitor interactions between GlcNAc-containing glycans and a model lectin (wheat germ agglutinin). In contrast to the widely employed trifluoroacetamide group, the difluoroacetamide tag contains geminal (1) H and (19) F atoms that allow both (1) H and (19) F NMR methods for easy and robust detection of molecular recognition processes involving GlcNAc- (or GalNAc-) moieties over a range of binding affinities. The CHF2 CONH- moiety behaves in a manner that is very similar to that of the natural acetamide fragment in the involved aromatic-sugar interactions, providing analogous binding energy and conformations, whereas the perfluorinated CF3 CONH- analogue differs more significantly.


Assuntos
Acetamidas/química , Flúor/química , Fluoracetatos/química , Polissacarídeos/química , Ligação de Hidrogênio , Lectinas/metabolismo , Espectroscopia de Ressonância Magnética , Modelos Moleculares
3.
Bioorg Med Chem Lett ; 25(22): 5190-3, 2015 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-26463133

RESUMO

The dual inhibitory action of the pain related peptide opiorphin (H-Gln-Arg-Phe-Ser-Arg-OH) against neutral endopeptidase (NEP) and aminopeptidase N (AP-N) was further investigated by a SAR study involving minor modifications on the polar side chains of Arg residues and glycosylation with monosaccharides at Ser. None of them exerted dual or individual inhibitory potency superior than opiorphin. However, the correlations deduced offer further proof for the key role of these residues upon the binding and bioactive conformational stabilization of opiorphin. NMR conformational studies on the glycopeptides suggest that they are still very flexible compounds that may attain their respective bioactive conformations.


Assuntos
Antígenos CD13/antagonistas & inibidores , Neprilisina/antagonistas & inibidores , Oligopeptídeos/química , Proteínas e Peptídeos Salivares/química , Acetilgalactosamina/química , Acetilglucosamina/química , Substituição de Aminoácidos , Arginina/química , Glicopeptídeos/química , Humanos , Modelos Moleculares , Estrutura Terciária de Proteína , Serina/química , Relação Estrutura-Atividade
4.
Nat Prod Rep ; 28(6): 1118-25, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21487589

RESUMO

In this review, we present applications of NMR spectroscopy as a potent tool for the study of molecular interactions. It is clear that a variety of NMR methods may be employed to deduce key features of ligand­receptor molecular recognition processes, looking at the process from the perspective of the receptor or the ligand. We have not provided an exhaustive review, but we have tried to focus on describing the different aspects within this research topic. We have therefore selected examples accordingly, depending on the particular problem under study or the application/development of protocols to circumvent the technical problems that may be found when working in this field.


Assuntos
Produtos Biológicos , Ressonância Magnética Nuclear Biomolecular/métodos , Produtos Biológicos/química , Produtos Biológicos/metabolismo , Estrutura Molecular
5.
Org Biomol Chem ; 9(17): 6133-42, 2011 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-21773621

RESUMO

To examine if the biological activity of the N/OFQ peptide, which is the native ligand of the pain-related and viable drug target NOP receptor, could be modulated by glycosylation and if such effects could be conformationally related, we have synthesized three N/OFQ glycopeptide analogues, namely: [Thr(5)-O-α-D-GalNAc-N/OFQ] (glycopeptide 1), [Ser(10)-O-α-D-GalNAc]-N/OFQ (glycopeptide 2) and [Ser(10)-O-ß-D-GlcNAc]-N/OFQ] (glycopeptide 3). They were tested for biological activity in competition binding assays using the zebrafish animal model in which glycopeptide 2 exhibited a slightly improved binding affinity, whereas glycopeptide 1 showed a remarkably reduced binding affinity compared to the parent compound and glycopeptide 3. The structural analysis of these glycopeptides and the parent N/OFQ peptide by NMR and circular dichroism indicated that their aqueous solutions are mainly populated by random coil conformers. However, in membrane mimic environments a certain proportion of the molecules of all these peptides exist as α-helix structures. Interestingly, under these experimental conditions, glycopeptide 1 (glycosylated at Thr-5) exhibited a population of folded hairpin-like geometries. From these facts it is tempting to speculate that nociceptin analogues showing linear helical structures are more complementary and thus interact more efficiently with the native NOP receptor than folded structures, since glycopeptide 1 showed a significantly reduced binding affinity for the NOP receptor.


Assuntos
Glicopeptídeos/química , Glicopeptídeos/farmacologia , Peptídeos Opioides/química , Peptídeos Opioides/farmacologia , Receptores Acoplados a Proteínas G/metabolismo , Sequência de Aminoácidos , Animais , Ligação Competitiva , Linhagem Celular , Glicopeptídeos/síntese química , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Peptídeos Opioides/síntese química , Ligação Proteica , Receptores Opioides/agonistas , Peixe-Zebra , Nociceptina
7.
J Med Chem ; 55(3): 1181-8, 2012 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-22224710

RESUMO

Toward developing new potential analgesics, this first structure-activity relationship study of opiorphin (H-Gln-Arg-Phe-Ser-Arg-OH), a human peptide inhibiting enkephalin degradation, was performed. A systematic Ala scanning proved that Phe(3) is a key residue for neprilysin and aminopeptidase N (AP-N) ectoenkephalinase inhibition. A series of Phe(3)-halogenated analogues revealed that halogen bonding based optimization strategies are not applicable to this residue. Additional substituted Phe(3) derivatives showed that replacing l-Phe(3) for d-Phe(3) increased the AP-N inhibition potency by 1 order of magnitude. NMR studies and molecular mechanics calculations indicated that the improved potency may be due to CH-π stacking interactions between the aromatic ring of d-Phe(3) and the Hγ protons of Arg(2). This structural motif is not possible for the native opiorphin and may be useful for the design of further potent and metabolically stable analogues.


Assuntos
Analgésicos/síntese química , Antígenos CD13/antagonistas & inibidores , Neprilisina/antagonistas & inibidores , Oligopeptídeos/síntese química , Inibidores de Proteases/síntese química , Proteínas e Peptídeos Salivares/síntese química , Analgésicos/química , Antígenos CD13/química , Ensaios Enzimáticos , Humanos , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Neprilisina/química , Oligopeptídeos/química , Inibidores de Proteases/química , Teoria Quântica , Proteínas e Peptídeos Salivares/química , Técnicas de Síntese em Fase Sólida , Soluções , Relação Estrutura-Atividade
8.
J Agric Food Chem ; 60(4): 1028-35, 2012 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-22229884

RESUMO

Bifidobacteria are natural members of the human intestinal microbiota and some strains are being used as probiotics. Adaptation to bile can allow them to increase survival in gastrointestinal conditions, thus improving their viability. Bifidobacterium longum NB667 and the cholate-resistant strain B. longum IPLA B667dCo produced exopolysaccharides (EPS) that were partially characterized. Analysis by size exclusion chromatography-multiangle laser light scattering indicated that the EPS crude fractions of both strains contained two polymer peaks of different molar mass. On the basis of chromatographic techniques both peaks appeared to be heteropolysaccharides. The smaller peak was mainly composed of glucose, galactose and rhamnose whose molar ratios and linkage types showed slight variations between the EPS fractions of both strains. The bigger peak consisted of glucose and galactose; the monosaccharide composition was identical in the EPS fractions of the two microorganisms, but their infrared spectra presented some differences regarding compounds other than carbohydrates that seem to be associated to the polymer. Differences in the composition of EPS fractions did not affect the capability of crude EPS from B. longum to be fermented by the human intestinal microbiota in fecal batch cultures.


Assuntos
Bifidobacterium/metabolismo , Colatos , Farmacorresistência Bacteriana , Polissacarídeos Bacterianos/biossíntese , Bifidobacterium/efeitos dos fármacos , Cromatografia em Gel , Fezes/microbiologia , Fermentação , Galactose/análise , Glucose/análise , Humanos , Intestinos/microbiologia , Polissacarídeos Bacterianos/química
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