A streptavidin mutant useful for directed immobilization on solid surfaces.
A streptavidin mutant has been designed and produced that allows the specific, covalent immobilization of streptavidin on solid surfaces. This streptavidin mutant was constructed by fusing a six-residue sequence, containing a single cysteine, to the carboxyl terminus of streptavidin. Because this mutant has no other cysteine residues, the reactive sulfhydryl group of the cysteine residue serves as a unique immobilization site for conjugation using sulfhydryl chemistry. This streptavidin mutant was efficiently immobilized on maleimide-coated solid surfaces via its unique immobilization site. Characterization of the immobilized streptavidin mutant for the ability to bind to biotinylated macromolecules and the dissociation rates of bound biotin showed that the biotin-binding properties of this mutant were minimally affected by immobilization on solid surfaces. This streptavidin could be readily 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.
Reznik
G O
GO
Center for Advanced Biotechnology and Department of Biomedical Engineering, Boston University, Boston, Massachusetts 02215, USA.
Vajda
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S
Cantor
C R
CR
Sano
T
T
D000345:Affinity Labels / Q000737:chemistry / Q000378:metabolism; D001710:Biotin / Q000378:metabolism; D019921:Biotinylation; D003545:Cysteine / Q000737:chemistry; D004247:DNA / Q000032:analysis / Q000302:isolation & purification; D005822:Genetic Vectors; D008301:Maleimides / Q000737:chemistry; D008722:Methods; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019809:Streptavidin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013499:Surface Properties
D000345:Affinity Labels; D008301:Maleimides; D011993:Recombinant Fusion Proteins; D001710:Biotin; D004247:DNA; D019809:Streptavidin; D003545:Cysteine