Expanding the utility of beta-galactosidase complementation: piece by piece.
The ability to image and quantify multiple biomarkers in disease necessitates the development of split reporter fragment platforms. We have divided the beta-galactosidase enzyme into unique, independent polypeptides that are able to reassemble and complement enzymatic activity in bacteria and in mammalian cells. We created two sets of complementing pairs that individually have no enzymatic activity. However, when brought into close geometric proximity, the complementing pairs associated resulting in detectable enzymatic activity. We then constructed a stable ligand complex composed of reporter fragment, linker, and targeting moiety. The targeting moiety, in this case a ligand, allowed cell surface receptor targeting in vitro. Further, we were able to simultaneously visualize two cell surface receptors implicated in cancer development, epidermal growth factor receptor and transferrin receptor, using complementing pairs of the ligand-reporter fragment complex.
Broome
Ann-Marie
AM
Department of Biomedical Engineering, Case Center for Imaging Research and National Foundation for Cancer Research for Molecular Imaging, Case Western Reserve University, Cleveland, Ohio 44106, USA. ann-marie.broome@case.edu
Bhavsar
Nihir
N
Ramamurthy
Gopalakrishnan
G
Newton
Gail
G
Basilion
James P
JP
D000818:Animals; D014408:Biomarkers, Tumor / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D066246:ErbB Receptors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D048168:Escherichia coli K12 / Q000235:genetics / Q000378:metabolism; D017930:Genes, Reporter; D005816:Genetic Complementation Test; D005910:Glioma / Q000235:genetics / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011990:Receptors, Transferrin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D001616:beta-Galactosidase / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism
D014408:Biomarkers, Tumor; D010446:Peptide Fragments; D011990:Receptors, Transferrin; D066246:ErbB Receptors; D001616:beta-Galactosidase