Aggregation and Molecular Properties of ?-Glucosidase Isoform II in Chayote (Sechium edule).

ABSTRACT: The presence of isoforms of ?-glucosidase has been reported in some grasses such as sorghum, rice and maize. This work aims to extract and characterize isoform II in ?-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 ?-glucosidase and 55.74% identical to ?-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 ?-d-glucose with a Km of 4.59 mM, a Vmax value of 104.3 ?M?min-1 and a kcat of 10,087 ?M?min-1 using p-nitrophenyl-?-D-glucopyranoside. The presence of molecular aggregates can be attributed to non-polar amino acids. This property is not mediated by a ?-glucosidase aggregating factor (BGAF) as in grasses (maize and sorghum). The role of these aggregates is discussed.

SUBMITTER: Cruz Rodriguez A 

PROVIDER: S-EPMC7180739 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature