Sequencing, cloning and heterologous expression of cyclomaltodextrin glucanotransferase of Bacillus firmus strain 37 in Bacillus subtilis WB800

COSTA, HERNÁN; FERRAROTTI SUSANA ALICIA; ALVEZ DE LIMA NETO QUIRINO; MATIOLI, GRACIETTE; GREGOLIN GIMENEZ, GABRIELA; FERNANDEZ MARÍA APRECIADA

BIOPROCESS AND BIOSYSTEMS ENGINEERING

SPRINGER

Lugar: Berlin; Año: 2018

1615-7591

Bacillus firmus strain 37 produces the cyclomaltodextrin glucanotransferase (CGTase) enzyme andCGTase produces cyclodextrins (CDs) through a starch cyclization reaction. The strategy for the cloningand expression of recombinant CGTase is a potentially viable alternative for the economically viableproduction of CGTase for use in industrial processes. The present study used Bacillus subtilis WB800 asa bacterial expression host for the production of recombinant CGTase cloned from the CGTase gene of B.firmus strain 37. The CGTase gene was cloned in TOPO-TA® plasmid, which was transformed inEscherichia coli DH5α. The subcloning was carried out with pWB980 plasmid and transformation in B.subtilis WB800. The 2xYT medium was the most suitable for the production of recombinant CGTase.The enzymatic activity of the crude extract of the recombinant CGTase of B. subtilis WB800 was 1.33μmol β-CD/min/mL, or 7.4 times greater than the enzymatic activity of the crude extract of CGTaseobtained from the wild strain. Following purification, the recombinant CGTase exhibited an enzymaticactivity of 157.78 μmol β-CD/min/mL, while the activity of the CGTase from the wild strain was 9.54μmol β-CD/min/mL. When optimal CDs production conditions for the CGTase from B. firmus strain 37were used, it was observed that the catalytic properties of the CGTase enzymes were equivalent. Thestrategy for the cloning and expression of CGTase in B. subtilis WB800 was efficient, with the productionof greater quantities of CGTase than with the wild strain, offering essential data for the large-scaleproduction of the recombinant enzyme.