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Purification, Biochemical Characterization, and Bioactive Properties of a Lectin Purified from the Seeds of White Tepary Bean (Phaseolus Acutifolius Variety Latifolius)
Institute of Health Sciences, Universidad Autónoma del Estado de Hidalgo, Ex-Hacienda de la Concepción, Tilcuautla, CP 42080 Pachuca de Soto, Hgo, Mexico
Center for Food Research and Development, A. C. Carretera a la Victoria Km 0.6 C.P. 83304. Hermosillo, Sonora, Mexico
Ezose Sciences Inc, Pine Brook, NJ 07058, USA
Basic Science and Engineering Institute, Universidad Autónoma del Estado de Hidalgo, Carr. A-Pachuca-Tulancingo Km 4.5 Cd Universitaria, CP 42184, Mineral de la Reforma, Hgo, Mexico
Laboratory of Biochemistry and Experimental Medicine, Division of Biomedical Research, National Medical Center “20 de Noviembre”, ISSSTE, México D.F., Mexico
* Author to whom correspondence should be addressed.
Received: 19 January 2011; in revised form: 17 March 2011 / Accepted: 17 March 2011 / Published: 21 March 2011
Abstract: The present work shows the characterization of Phaseolus acutifolius variety latifolius, on which little research has been published, and provides detailed information on the corresponding lectin. This protein was purified from a semi-domesticated line of white tepary beans from Sonora, Mexico, by precipitation of the aqueous extract with ammonium sulfate, followed by affinity chromatography on an immobilized fetuin matrix. MALDI TOF analysis of Phaseolus acutifolius agglutinin (PAA) showed that this lectin is composed of monomers with molecular weights ranging between 28 and 31 kDa. At high salt concentrations, PAA forms a dimer of 63 kDa, but at low salt concentrations, the subunits form a tetramer. Analysis of PAA on 2D-PAGE showed that there are mainly three types of subunits with isoelectric points of 4.2, 4.4, and 4.5. The partial sequence obtained by LC/MS/MS of tryptic fragments from the PAA subunits showed 90–100% identity with subunits from genus Phaseolus lectins in previous reports. The tepary bean lectin showed lower hemagglutination activity than Phaseolus vulgaris hemagglutinin (PHA-E) toward trypsinized human A and O type erythrocytes. The hemagglutination activity was inhibited by N-glycans from glycoproteins. Affinity chromatography with the immobilized PAA showed a high affinity to glycopeptides from thyroglobulin, which also has N-glycans with a high content of N-acetylglucosamine. PAA showed less mitogenic activity toward human lymphocytes than PHA-L and Con A. The cytotoxicity of PAA was determined by employing three clones of the 3T3 cell line, demonstrating variability among the clones as follows: T4 (DI50 51.5 µg/mL); J20 (DI50 275 µg/mL), and N5 (DI50 72.5 µg/mL).
Keywords: tepary bean; Phaseolus acutifolius; hemagglutinins; oligosaccharide specificity
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Valadez-Vega, C.; Guzmán-Partida, A.M.; Soto-Cordova, F.J.; Álvarez-Manilla, G.; Morales-González, J.A.; Madrigal-Santillán, E.; Villagómez-Ibarra, J.R.; Zúñiga-Pérez, C.; Gutiérrez-Salinas, J.; Becerril-Flores, M.A. Purification, Biochemical Characterization, and Bioactive Properties of a Lectin Purified from the Seeds of White Tepary Bean (Phaseolus Acutifolius Variety Latifolius). Molecules 2011, 16, 2561-2582.
Valadez-Vega C, Guzmán-Partida AM, Soto-Cordova FJ, Álvarez-Manilla G, Morales-González JA, Madrigal-Santillán E, Villagómez-Ibarra JR, Zúñiga-Pérez C, Gutiérrez-Salinas J, Becerril-Flores MA. Purification, Biochemical Characterization, and Bioactive Properties of a Lectin Purified from the Seeds of White Tepary Bean (Phaseolus Acutifolius Variety Latifolius). Molecules. 2011; 16(3):2561-2582.
Valadez-Vega, Carmen; Guzmán-Partida, Ana María; Soto-Cordova, Francisco Javier; Álvarez-Manilla, Gerardo; Morales-González, José A.; Madrigal-Santillán, Eduardo; Villagómez-Ibarra, José Roberto; Zúñiga-Pérez, Clara; Gutiérrez-Salinas, José; Becerril-Flores, Marco A. 2011. "Purification, Biochemical Characterization, and Bioactive Properties of a Lectin Purified from the Seeds of White Tepary Bean (Phaseolus Acutifolius Variety Latifolius)." Molecules 16, no. 3: 2561-2582.