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1.
Molecules ; 29(7)2024 Apr 03.
Artículo en Inglés | MEDLINE | ID: mdl-38611875

RESUMEN

Mamey (Mammea americana L.) is a tropical fleshy fruit native from the West Indies and northern South America. It is very appreciated for its flavor and color but has been little described. The present study investigates the composition and histochemistry of the pulp cell walls of three mamey accessions readily available in Martinique. The impact of pulp processing into puree on cell wall composition is evaluated. The histology and rheology of mamey puree are assessed considering these characterizations. Mamey pulp cell wall composition is dominated by highly methyl-esterified pectins (DM: 66.2-76.7%) of high molecular weight, and show few hemicelluloses, mainly xyloglucans. Processing reduced methyl-esterified uronic acid contents and gave purees with significantly different viscosities. Mamey puree was composed of polydisperse particles (20-2343 µm), which size distributions were different depending on the accession: Ti Jacques was dominated by smaller particles (50% had approximated diameters lower than 160 µm), Sonson's by larger particles (50% had approximated diameters higher than 900 µm), and Galion's had an intermediate profile. This new knowledge on mamey pulp is valuable for future works on mamey processing into new food products, even more so for those including cell wall polysaccharide-degrading enzymes.


Asunto(s)
Mammea , Pared Celular , Alimentos , Histocitoquímica , Peso Molecular
2.
J Biol Chem ; 283(44): 30112-20, 2008 Oct 31.
Artículo en Inglés | MEDLINE | ID: mdl-18687680

RESUMEN

Codakine is an abundant 14-kDa mannose-binding C-type lectin isolated from the gills of the sea bivalve Codakia orbicularis. Binding studies using inhibition of hemagglutination indicated specificity for mannose and fucose monosaccharides. Further experiments using a glycan array demonstrated, however, a very fine specificity for N-linked biantennary complex-type glycans. An unusually high affinity was measured by titration microcalorimetry performed with a biantennary Asn-linked nonasaccharide. The crystal structure of the native lectin at 1.3A resolution revealed a new type of disulfide-bridged homodimer. Each monomer displays three intramolecular disulfide bridges and contains only one calcium ion located in the canonical binding site that is occupied by a glycerol molecule. The structure of the complex between Asn-linked nonasaccharide and codakine has been solved at 1.7A resolution. All residues could be located in the electron density map, except for the capping beta1-4-linked galactosides. The alpha1-6-linked mannose binds to calcium by coordinating the O3 and O4 hydroxyl groups. The GlcNAc moiety of the alpha1,6 arm engages in several hydrogen bonds with the protein, whereas the GlcNAc on the other antenna is stacked against Trp(108), forming an extended binding site. This is the first structural report for a bivalve lectin.


Asunto(s)
Calorimetría/métodos , Lectinas/química , Polisacáridos/química , Secuencia de Aminoácidos , Animales , Bivalvos , Conformación de Carbohidratos , Secuencia de Carbohidratos , Cristalografía por Rayos X/métodos , Dimerización , Disulfuros/química , Cinética , Conformación Molecular , Datos de Secuencia Molecular , Homología de Secuencia de Aminoácido
3.
Fish Shellfish Immunol ; 22(5): 498-509, 2007 May.
Artículo en Inglés | MEDLINE | ID: mdl-17169576

RESUMEN

This work relates to the characterisation of the predominant gill protein of the white clam, Codakia orbicularis (Linné, 1758), which harbours endosymbiotic sulphur-oxidising chemoautotrophic bacteria. Total RNA was extracted from the clam to perform 3'rapid amplification of cDNA ends (3'RACE) using degenerate oligonucleotides prepared from a partial sequence of a predominant protein of about 14kDa, termed codakine. The partial peptide sequence was obtained by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and Edman degradation. Clones isolated from the cDNA library and containing the gene of interest were used in polymerase chain reaction (PCR). The PCR and RACE-PCR products were sequenced to determine the entire coding DNA sequence of codakine. BLAST analysis revealed about 23% to 29% sequence identity between codakine and various animal C-type lectins that are often involved in symbiosis and immune defences. Codakine also contains the motifs and domains of Ca(2+)-dependent C-type lectins, and in particular the tripeptide EPN motif frequently found in mannose-binding lectins (MBLs). Analysis of the protein by affinity chromatography on a mannose-agarose column is consistent with the findings that codakine is a dimeric Ca(2+)-dependent MBL of about 29kDa. Based on the present results, it is hypothesised that this novel C. orbicularis gill protein is involved in the recognition of symbiotic and pathogenic bacteria.


Asunto(s)
Bivalvos/genética , Lectina de Unión a Manosa/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Cromatografía de Afinidad/veterinaria , Cartilla de ADN/química , ADN Complementario/química , Electroforesis en Gel de Poliacrilamida/veterinaria , Biblioteca de Genes , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa/veterinaria , Alineación de Secuencia/veterinaria , Homología de Secuencia de Aminoácido , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/veterinaria
4.
Appl Biochem Biotechnol ; 125(1): 41-52, 2005 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-15834161

RESUMEN

We used the recombinant phage display antibody system (RPAS) to obtain chimeric single-chain fragment variable (ScFv) antibodies to gill proteins of the white clam Codakia orbicularis (Linné, 1758). After three rounds of selection on immunotubes loaded with total gill protein extract, recombinant phages exhibiting antibodies to gill proteins were isolated and tested by enzyme-linked immunosorbent assay (ELISA). Clones exhibiting a high affinity for the mollusk proteins were selected for production of soluble ScFv antibodies, which were purified for subsequent analysis. ScFv antibodies exhibited a reaction specific for a protein whose molecular mass was about 15,000 Daltons and that was detected by the antigen capture technique followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blotting.


Asunto(s)
Bacteriófagos/inmunología , Bivalvos/química , Branquias/química , Proteínas/análisis , Animales , Anticuerpos Antivirales/inmunología , Anticuerpos Antivirales/aislamiento & purificación , Electroforesis en Gel de Poliacrilamida , Biblioteca Genómica , Inmunoensayo/métodos , Región Variable de Inmunoglobulina/inmunología , Recombinación Genética
5.
Prep Biochem Biotechnol ; 32(4): 341-53, 2002 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-12455827

RESUMEN

Marine organisms inhabiting the coastal environment of the Caribbean islands have attracted the attention of a number of research scientists. These organisms generally live in areas of high environmental stress and may, therefore, contain specialized proteins and enzymes which exhibit valuable biotechnological applications. Among them, a small number of clams have been studied. Our work relates to the clam Codakia orbicularis. This bivalve lives in areas of high hydrogen sulfide concentrations on the Caribbean island of Guadeloupe. Its enzymatic system must, therefore, have evolved to allow its adaptation to this high-stress environment. C. orbicularis also contains endosymbiotic bacteria which are housed in the bacteriocytes of the gills. Its protein content can, therefore, be expected to have an impact on this symbiotic relationship. We have analysed gill protein extracts of this clam by various biochemical techniques: SDS-PAGE, IEF, PAS, and Western blotting using a panel of lectins, in order to establish its protein and glycoprotein profiles. This biochemical analysis, the first of its kind, constitutes an important step in separating and characterizing the proteins involved in the biochemical pathways of this organism whose stock is in decline in Guadeloupe. Our results show the presence of three major proteins whose molecular weights vary between 14,000 and 24,000 Daltons, and some of which are glycoproteins with predominantly alpha-mannose and N-acetylgalactosamine moieties. Their pI values are in the range between 4.5 and 5.6. These protein profiles are different from those observed for Lucina pectinata, a clam which has been the subject of earlier studies in the literature.


Asunto(s)
Bivalvos/química , Branquias/química , Proteínas/química , Animales , Electroforesis , Electroforesis en Gel de Poliacrilamida , Glicoproteínas/química , Focalización Isoeléctrica , Reacción del Ácido Peryódico de Schiff
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