Engineered Metal Binding Sites on Green Fluorescence Protein

Abstract

The ability to assay a variety of metals by noninvasive methods has applications in both biomedical and environmental research. Green fluorescent protein (GFP) is a protein isolated from coelenterates that exhibits spontaneous fluorescence. GFP does not require any exogenous cofactors for fluorescence, and can be easily appended to other proteins at the DNA level, producing a fluorescence-labeled target protein in vivo. Metals in close proximity to chromophores are known to quench fluorescence in a distance-dependent fashion. Potential metal binding sites on the surface of GFP have been identified and mutant proteins have been designed, created, and characterized. These metal-binding mutants of GFP exhibit fluorescence quenching at lower transition metal ion concentrations than those of the wild-type protein. These GFP mutants represent a new class of protein-based metal sensors.