Strep Tag

strep-tags I, II and III, nano-tag and streptavidin binding peptide (SBP) tag. These tags bind directly to streptavidin without the need of biotinylation. Although both the nano-tag and SBP tag bind to streptavidin with comparable binding affinity in the nanomolar range, nano-tag is functional only at the N-terminus of the tagged proteins21. In contrast, SBP tag offers greater flexibility and can be inserted to a recombinant protein at N-terminal, internal or C-terminal position, without affecting its function. SAVSBPM18, which can bind both biotin and SBP tag with the binding affinity in the range of 10−8 M, the SAVSBPM18 affinity matrix can be applied to purify biotinylated biomolecules and SBP tagged recombinant proteins in a reusable manner. The production yield for the soluble SAVSBPM18-L-ABD(KC) was estimated to be around 300 mg/liter of cell culture. SAVSBPM18-L-ABD(KC) produced was mainly in the tetrameric state. SAVSBPM18-L-ABD(KC) readily binds to biotin. When the crude soluble fraction containing the fusion protein was loaded onto a column of biotin agarose in the presence of lactose, SAVSBPM18-L-ABD(KC) was selectively retained on the column and could be eluted by biotin. The fusion protein could be affinity purified in one step using biotin agarose L-ABD(KC) and SAVSBPM18-L-ABD(KC) could be affinity purified to a high degree of purity in one step with crude cell extracts as the starting material using Sepharose 6B-CL. Sepharose 6B-CL was selected as the matrix in this study because it is relatively uniform in size (diameter in the range of 45–145 μm with the majority around 50 μm) and it is relatively inert with negligible non-specific interactions.