Cloning of the Transglutaminase Gene from Bacillus subtilis and Fusion Expression in Escherchia coli

The gene of transglutaminase was amplified from the chromosome of Bacillus subtilis DB403 by PCR and translationally fused with thioredoxin gene ( trx A) in the plasmid pET32a(+) to achieve the expression vector pET32 BTGase. The fusion protein Trx BTGase could be expressed in E. coli BL21 (DE3) after IPTG induction, with a level up to 26 percent of the total bacterial proteins. Trx BTGase was initially purified from the supernatant of the bacterial lysate by metal chelating chromatography with a purity of over 80 percent, and finally purified with Superdex 75. Enzyme activity assay demonstrated that the Trx BTGase expressed in E. coli could be functional in polymerizing other proteins, and no influence on the biological activities of BTGase to polymerize BSA (bovine serum albumin) was found in case of the thrombin cleavage of this fusion protein.;