RESUMEN
This work reports the characterization of an amylolytic enzyme from the bacteria Massilia timonae CTI-57. A gene encoding this protein was expressed from the pTrcHis2B plasmid in Escherichia coli BL21 Star™ (DE3). The purified protein had 64 kDa, and its modeled structure showed a monomer with the conserved α-amylases structure composed of the domain A with the characteristic (ß/α)8-barrel, the small domain B, and the domain C with an antiparallel beta-sheet. Phylogenetic analysis demonstrated that the expressed protein belongs to the GH13_19 subfamily of glycoside hydrolases. The ions Ca2+, Mn2+, Na+, Mg2+, Mo6+, and K+ did activate the purified enzyme, while EDTA and the ions Fe2+, Hg2+, Zn2+, and Cu2+ were strong inhibitors. SDS was also a strong inhibitor. The enzyme's optimal pH and temperature were 7.0 and 45 °C, respectively, and its Tm was 62.2 °C. The KM of the purified enzyme for starch was 13 mg/mL, and the Vmax was 0.24 µmol of reducing sugars released per min. The characterized enzyme presented higher specificity for maltodextrin and starch and produced maltose as the main starch hydrolysis product. This is the first characterized maltose-forming amylolytic enzyme from the GH13_19 subfamily. The purified enzyme produced ß-cyclodextrin from starch and maltodextrin and could be considered a cyclodextrin glucanotransferase (CGTase). This is the first report of a GH13_19 subfamily enzyme with CGTase activity.