Engineering of Escherichia coli β-galactosidase for solvent display of a functional scFv antibody fragment

AbstractProtein engineering allows the generation of hybrid polypeptides with functional domains from different origins and therefore exhibiting new biological properties. We have explored several permissive sites in Escherichia coli β-galactosidase to generate functional hybrid enzymes displaying a mouse scFv antibody fragment. When this segment was placed at the amino-terminus of the enzyme, the whole fusion protein was stable, maintained its specific activity and interacted specifically with the target antigen, a main antigenic determinant of foot-and-mouth disease virus. In addition, the antigen-targeted enzyme was enzymatically active when bound to the antigen and therefore useful as a reagent in single-step immunoassays. These results prove the flexibility of E. coli β-galactosidase as a carrier for large-sized functional domains with binding properties and prompt the further exploration of the biotechnological applicability of the scFv enzyme targeting principle for diagnosis or other biomedical applications involving antigen tagging