TY - JOUR
T1 - Purification, biochemical characterization, and bioactive properties of a lectin purified from the seeds of white tepary bean (phaseolus acutifolius variety latifolius)
AU - Valadez-Vega, Carmen
AU - Guzmán-Partida, Ana María
AU - Soto-Cordova, Francisco Javier
AU - Álvarez-Manilla, Gerardo
AU - Morales-González, José A.
AU - Madrigal-Santillán, Eduardo
AU - Villagómez-Ibarra, José Roberto
AU - Zúniga-Pérez, Clara
AU - Gutiérrez-Salinas, José
AU - Becerril-Flores, Marco A.
PY - 2011/3
Y1 - 2011/3
N2 - 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).
AB - 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).
KW - Hemagglutinins
KW - Oligosaccharide specificity
KW - Phaseolus acutifolius
KW - Tepary bean
UR - http://www.scopus.com/inward/record.url?scp=79953070831&partnerID=8YFLogxK
U2 - 10.3390/molecules16032561
DO - 10.3390/molecules16032561
M3 - Artículo
C2 - 21441861
AN - SCOPUS:79953070831
SN - 1420-3049
VL - 16
SP - 2561
EP - 2582
JO - Molecules
JF - Molecules
IS - 3
ER -