Recombinant core streptavidins. A minimum-sized core streptavidin has enhanced structural stability and higher accessibility to biotinylated macromolecules.
Two recombinant core streptavidins were designed and characterized to understand the role of the terminal sequences, present in naturally truncated core streptavidins, in the properties of streptavidin. One recombinant core streptavidin, Stv-25, has an amino acid sequence very similar to natural core streptavidins. The other recombinant molecule, Stv-13, has further truncation of the terminal residues and consists essentially of only the beta-barrel structure characteristic of streptavidin. These recombinant core streptavidins are tetrameric and bind four biotins/molecule, as does natural streptavidin. The solubility characteristics of Stv-13, determined by varying the concentration of ammonium sulfate or ethanol, were almost the same as those of Stv-25 and natural core streptavidin. However, Stv-13 showed an enhanced structural stability compared with Stv-25 and natural core streptavidin. For example, Stv-13 retained greater than 80% of its biotin binding ability after incubation in 6 M guanidine hydrochloride at pH 1.5, under which conditions, Stv-25 and natural core streptavidin retained only about 20% of their biotin binding ability. In addition, Stv-13 showed higher accessibility to biotinylated DNA than natural core streptavidin. Apparently, the terminal regions, present on the surface of natural core streptavidin, spatially hinder biotinylated macromolecules from approaching the biotin binding sites.
D000595:Amino Acid Sequence; D000645:Ammonium Sulfate / Q000494:pharmacology; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D001710:Biotin; D004355:Drug Stability; D004591:Electrophoresis, Polyacrylamide Gel; D000431:Ethanol / Q000494:pharmacology; D019791:Guanidine; D006146:Guanidines / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation / Q000187:drug effects; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism; D012995:Solubility; D019809:Streptavidin; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D006146:Guanidines; D011994:Recombinant Proteins; D000431:Ethanol; D001710:Biotin; D019809:Streptavidin; D019791:Guanidine; D000645:Ammonium Sulfate