Purification, Characterization, and Preliminary X-Ray Diffraction Analysis of a Lactose-Specific Lectin from Cymbosema roseum Seeds
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Data
2009-03-01
Autores
Rocha, Bruno A. M.
Moreno, Frederico B. M. B. [UNESP]
Delatorre, Plinio
Souza, Emmanuel P.
Marinho, Emmanuel S.
Benevides, Raquel G.
Rodrigues Rustiguel, Joane Kathelen [UNESP]
Souza, Luis A. G.
Nagano, Celso S.
Debray, Henri
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Humana Press Inc
Resumo
The unique carbohydrate-binding property of lectins makes them invaluable tools in biomedical research. Here, we report the purification, partial primary structure, carbohydrate affinity characterization, crystallization, and preliminary X-ray diffraction analysis of a lactose-specific lectin from Cymbosema roseum seeds (CRLII). Isolation and purification of CRLII was performed by a single step using a Sepharose-4B-lactose affinity chromatography column. The carbohydrate affinity characterization was carried using assays for hemagglutination activity and inhibition. CRLII showed hemagglutinating activity toward rabbit erythrocytes. O-glycoproteins from mucine mucopolysaccharides showed the most potent inhibition capacity at a minimum concentration of 1.2 A mu g mL(-1). Protein sequencing by mass spectrometry was obtained by the digestion of CRLII with trypsin, Glu-C, and AspN. CRLII partial protein sequence exhibits 46% similarity with the ConA-like alpha chain precursor. Suitable protein crystals were obtained with the hanging-drop vapor-diffusion method with 8% ethylene glycol, 0.1 M Tris-HCl pH 8.5, and 11% PEG 8,000. The monoclinic crystals belong to space group P2(1) with unit cell parameters a = 49.4, b = 89.6, and c = 100.8 A....
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Cymbosema roseum, Crystallization, Tandem mass spectrometry, Lectins, Lactose-specific lectin
Como citar
Applied Biochemistry and Biotechnology. Totowa: Humana Press Inc, v. 152, n. 3, p. 383-393, 2009.