Related Experiment Videos
Enzyme-complemented activatorsorbent assay (ECASA): genetic engineering for enzyme-linked immunosorbent assay-type
1Institut für Industrielle Genetik, Universität Stuttgart, Allmandring 31, Stuttgart, D-70569, Germany. kjoachim@gensun.biologie.uni-stuttgart.de
Abstract:
The sensor component of bacterial mercury resistance systems is the metalloregulatory protein MerR, which has nanomolar sensitivity and high selectivity for Hg(II). A fusion protein of MerR and the alpha-peptide part of beta-galactosidase (LacZalpha) was constructed by fusing the relevant genes. The protein exhibited both MerR functions and alpha-complementing activity to the inactive LacZDeltaM15 (M15) protein. The bifunctional character of the appropriate MerR-LacZalpha-complemented M15 protein (MerR-LacZalpha:M15 protein complex) was used to develop a Hg(II)-specific enzyme-complemented activatorsorbent assay. Hg(II) was immobilized and presented on a matrix taking advantage of the high affinity of Hg(II) to SH residues. The immobilized Hg(II) could be specifically detected down to the parts-per-billion level by quantifying the beta-galactosidase activity of the bound fusion protein complex.