Recombinant expression of transglutaminase from Atlantic cod in E. coli

Abstract

Transglutaminases (TGs) are a family of enzymes that catalyzes cross linking of proteins by forming covalent bonds between lysine residues and glutamine residues in different polypeptides. Cross-linking reactions are involved in blood clots, skin formation, embryogenesis and apoptosis. Clinically, they are interesting since they seem to be implicated in neurodegenerative diseases, tumours and celiac diseases. Transglutaminases also have a great potential in the food industry because of its ability to cross bind proteins that are not normally linked. A gene coding for transglutaminase from Atlantic cod was cloned into a bacterial expression vector pET151-D-TOPO. The recombinant plasmid was used to transform the bacterial expression strain BL21 and expression products were analyzed with SDS-PAGE, Immunoblotting and mass spectrometry analysis. The protein was expressed in the host in the soluble fraction, which was confirmed by SDS PAGE, imunoblotting and mass spectrometry analysis. A polyhistidine tag introduced at the amino-acid terminus of the transglutaminase protein allowed for the purification of the protein by using His-Trap columns. Large scale purification of the protein was successful after optimizing the washing and eluting conditions of the buffer. Based on these results, the bacterial expression system successfully expressed transglutaminase and possible use of the enzyme as a meat-glue in food industry is investigated.