Molecular characterization of sucrose: sucrose 1-fructosyltransferase (1-SST) from Agave tequilana Weber var. azul.

A full-length cDNA encoding for 1-SST in Agave tequilana Weber var. azul. was isolated, cloned and expressed in Pichia pastoris. The heterologous protein, with a molecular mass of 75 kDa, shows identity with different plant fructosyltransferases and invertases, which belong to the glycoside hydrolase 32 family. When sucrose was used as substrate, only the transference products 1-kestose and glucose were identified, while from 1-kestose, nystose and fructose were obtained as transference and hydrolysis products respectively, with sucrose as a secondary product. The enzyme has a low turnover number (260 min−1) at 30 °C, with optimal stability at 25 °C and pH 5.5. In order to identify the probable residues involved in the active site, a three-dimensional model was built using the fructan 1-exohydrolase IIa from Cichorium intybus (PDB entry 1st8) as template. This is the first report concerning cloning and expression of an Agave fructosyltransferase.