A streptavidin mutant useful for directed immobilization on solid surfaces

A streptavidin mutant has been designed and produced that allows the specif ic, covalent immobilization of streptavidin on solid surfaces. This strepta vidin mutant was constructed by fusing a six-residue sequence, containing a single cysteine, to the carboxyl terminus of streptavidin. Because this mu tant has no other cysteine residues, the reactive sulfhydryl group of the c ysteine residue serves as a unique immobilization site for corrugation usin g sulfhydryl chemistry. This streptavidin mutant was efficiently immobilize d on maleimide-coated solid surfaces via, its unique immobilization site. C haracterization of the immobilized streptavidin mutant for the ability to b ind to biotinylated macromolecules and the dissociation rates of bound biot in showed that the biotin-binding properties of this mutant were minimally affected by immobilization on solid surfaces. This streptavidin could be re adily incorporated into a wide variety of solid-phase diagnostic tests and biomedical assays. This could enhance the performance of streptavidin-based solid-phase assay systems.