| Autor: |
Cruz Rodríguez A; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México., Sánchez Esperanza FA; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México., Pérez-Campos E; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México., Hernández-Huerta MT; CONACyT Facultad de Medicina y Cirugía, Universidad Autónoma Benito Juárez de Oaxaca, Oaxaca 68020, México., Pérez-Campos Mayoral L; Centro de Investigación Facultad de Medicina UNAM-UABJO, Facultad de Medicina y Cirugía, Universidad Autónoma 'Benito Juárez' de Oaxaca, Oaxaca 68020, México., Matias-Cervantes CA; CONACyT Facultad de Medicina y Cirugía, Universidad Autónoma Benito Juárez de Oaxaca, Oaxaca 68020, México., Martínez Barras A; Facultad de Ingeniería, Universidad Autónoma de Querétaro, 76017, México., Mayoral-Andrade G; Centro de Investigación Facultad de Medicina UNAM-UABJO, Facultad de Medicina y Cirugía, Universidad Autónoma 'Benito Juárez' de Oaxaca, Oaxaca 68020, México., Santos Pineda LÁ; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México., Díaz Barrita AJ; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México., Zenteno E; Facultad de Medicina de la Universidad Nacional Autónoma de México, Ciudad de México 04510, México., Romero Díaz C; Centro de Investigación Facultad de Medicina UNAM-UABJO, Facultad de Medicina y Cirugía, Universidad Autónoma 'Benito Juárez' de Oaxaca, Oaxaca 68020, México., Martínez Cruz R; Centro de Investigación Facultad de Medicina UNAM-UABJO, Facultad de Medicina y Cirugía, Universidad Autónoma 'Benito Juárez' de Oaxaca, Oaxaca 68020, México., Pérez-Campos Mayoral E; Centro de Investigación Facultad de Medicina UNAM-UABJO, Facultad de Medicina y Cirugía, Universidad Autónoma 'Benito Juárez' de Oaxaca, Oaxaca 68020, México., Bernabé Pérez EA; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México., Pérez Santiago AD; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México., Pina-Canseco MDS; Centro de Investigación Facultad de Medicina UNAM-UABJO, Facultad de Medicina y Cirugía, Universidad Autónoma 'Benito Juárez' de Oaxaca, Oaxaca 68020, México., Martínez Cruz M; Tecnológico Nacional de México/IT de Oaxaca, Oaxaca 68030, México. |
| Abstrakt: |
The presence of isoforms of β-glucosidase has been reported in some grasses such as sorghum, rice and maize. This work aims to extract and characterize isoform II in β-glucosidase from S. edule . A crude extract was prepared without buffer solution and adjusted to pH 4.6. Contaminating proteins were precipitated at 4 °C for 24 h. The supernatant was purified by chromatography on carboxymethyl cellulose (CMC) column, molecular exclusion on Sephacryl S-200HR, and exchange anionic on QFF column. Electrophoretic analyzes revealed a purified enzyme with aggregating molecular complex on SDS-PAGE, Native-PAGE, and AU-PAGE. Twelve peptides fragments were identified by nano liquid chromatography-tandem mass spectrometry (nano LC-ESI-MS/MS), which presented as 61% identical to Cucurbita moschata β-glucosidase and 55.74% identical to β-glucosidase from Cucumis sativus , another Cucurbitaceous member. The relative masses which contained 39% hydrophobic amino acids ranged from 982.49 to 2,781.26. The enzyme showed a specificity to β-d-glucose with a K m of 4.59 mM, a V max value of 104.3 μM∙min -1 and a k cat of 10,087 μM∙min -1 using p -nitrophenyl-β-D-glucopyranoside. The presence of molecular aggregates can be attributed to non-polar amino acids. This property is not mediated by a β-glucosidase aggregating factor (BGAF) as in grasses (maize and sorghum). The role of these aggregates is discussed. |
