BETA-GALACTOSIDASE ENZYMATIC-ACTIVITY AS A MOLECULAR PROBE TO DETECT SPECIFIC ANTIBODIES

The main antigenic region of foot-and-mouth disease virus serotype C-1 , also called site A, has been inserted in zones of the beta-galactosi dase important for the stabilization of the active site, causing impor tant changes in the K-m and the specific activity of the resulting enz ymes. The peptide is displayed at the surface of the recombinant prote ins and, in all the cases, presents a good antigenicity. Among the rec ombinant proteins constructed, in proteins M278VP1 and M275SVP1 the pe ptide is inserted in a large loop of the beta-galactosidase (amino aci ds 272-288) involved in the formation of the activating interface. In these constructs, the binding of the specific antibodies directed to t he foreign peptide causes an increase of the beta-galactosidase activi ty up to about 200%. This phenomenon has been proved using monoclonal antibodies and also using polyclonal sera generated against the peptid e. Different hypothesis of the mechanism of modulation upon antibody b inding are discussed. This insertion site seems to be sensitive enough to enzymatic modulation mediated by antibody binding. We propose furt her exploring this insertion site as a tool for a rapid detection of s pecific antibodies in a quick and simple homogeneous assay based on th e colorimetric determination of beta-galactosidase activity.