The Streptavidin-Binding Peptide (SBP)-Tag is a 38-

amino acid Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha am ...

sequence that may be engineered into recombinant

proteins Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, respo ...

. Recombinant proteins containing the SBP-Tag bind to

streptavidin Streptavidin is a 66.0 (tetramer) kDa protein purified from the bacterium '' Streptomyces avidinii''. Streptavidin homo-tetramers have an extraordinarily high affinity for biotin (also known as vitamin B7 or vitamin H). With a dissociation co ...

and this property may be utilized in specific purification, detection or immobilization strategies. The sequence of the SBP tag is MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP.

Discovery

The Streptavidin-Binding Peptide was discovered within a library of seven trillion stochastically-generated peptides using the in vitro selection technique of

mRNA Display mRNA display is a display technique used for ''in vitro'' protein, and/or peptide evolution to create molecules that can bind to a desired target. The process results in translated peptides or proteins that are associated with their mRNA progenitor ...

. Selection was performed by incubating with streptavidin-agarose followed by elution with

biotin Biotin (or vitamin B7) is one of the B vitamins. It is involved in a wide range of metabolic processes, both in humans and in other organisms, primarily related to the utilization of fats, carbohydrates, and amino acids. The name ''biotin'', bor ...

. The SBP-Tag has been shown to bind streptavidin with an equilibrium

dissociation constant In chemistry, biochemistry, and pharmacology, a dissociation constant (K_D) is a specific type of equilibrium constant that measures the propensity of a larger object to separate (dissociate) reversibly into smaller components, as when a complex fa ...

of 2.5nM and is readily eluted with biotin under native conditions.

Applications

Protein purification

Because of the mild elution conditions (biotin plus wash buffer) SBP-Tagged proteins can be generated in a relatively pure state with a single purification step. There are several relatively abundant mammalian proteins that inherently associate with the

IMAC iMac is a family of all-in-one Mac desktop computers designed and built by Apple Inc. It has been the primary part of Apple's consumer desktop offerings since its debut in August 1998, and has evolved through seven distinct forms. In it ...

matrices that bind to the more commonly used

Polyhistidine-tag A polyhistidine-tag is an amino acid motif in proteins that typically consists of at least six histidine (''His'') residues, often at the N- or C-terminus of the protein. It is also known as hexa histidine-tag, 6xHis-tag, His6 tag, by the US trad ...

(His-tag). For this reason non-IMAC purification protocols, including with the SBP-Tag, are often preferred for proteins that are expressed in mammalian cells.

Protein complex purification

Complexes of interacting proteins may also be purified using the SBP-Tag because elution with biotin permits recovery under conditions in which desired complexes remain associated. For example, the

Condensin Condensins are large protein complexes that play a central role in chromosome assembly and segregation during mitosis and meiosis (Figure 1). Their subunits were originally identified as major components of mitotic chromosomes assembled in ''Xeno ...

Complex was purified by Kim ''et al.'' 010and complexes with the

TAZ Taz or TAZ may refer to: Geography *Taz (river), a river in western Siberia, Russia *Taz Estuary, the estuary of the river Taz in Russia People * Taz people, an ethnic group in Russia ** Taz language, a form of Northeastern Mandarin spoken by ...

transcriptional co-activator were purified by Zhang ''et al.''

009 009 may refer to: * OO9, gauge model railways * O09, FAA identifier for Round Valley Airport * 0O9, FAA identifier for Ward Field, see List of airports in California * British secret agent 009, see 00 Agent * BA 009, see British Airways Flight 9 * ...

The SBP-Tag has also been incorporated into several

Tandem Affinity Purification Tandem affinity purification (TAP) is an immunoprecipitation-based purification technique for studying protein–protein interactions. The goal is to extract from a cell only the protein of interest, in complex with any other proteins it interacted ...

(TAP) systems in which successive purification steps are utilized with multiple tags, for example

GFP GFP may refer to: Organisations * Gaelic Football Provence, a French Gaelic Athletic Association club * Geheime Feldpolizei, the German secret military police during the Second World War * French Group for the Study of Polymers and their Applicat ...

fusion proteins and BTK-protein complexes were purified using a TAP protocol with the SBP-Tag and the His-Tag, HDGF-protein complexes were purified using a TAP protocol with the SBP-Tag and with the

FLAG-tag FLAG-tag, or FLAG octapeptide, or FLAG epitope, is a peptide protein tag that can be added to a protein using recombinant DNA technology, having the sequence DYKDDDDK (where D= aspartic acid, Y=tyrosine, and K= lysine). It is one of the most specifi ...

and Wnt complexes were purified using a TAP protocol with the SBP-Tag and with the almodulin-Tag TAP is generally used with protein complexes and several studies report significant improvements in purity and yield when the SBP-Tag TAP systems are compared to non-SBP-Tag systems. Commercial TAP systems that use the SBP-Tag include the Interplay® Adenoviral and Mammalian TAP Systems sold by

Agilent Technologies Agilent Technologies, Inc. is an American life sciences company that provides instruments, software, services, and consumables for the entire laboratory workflow. Its global headquarters is located in Santa Clara, California. Agilent was establi ...

, similar products are sold by

Sigma-Aldrich Sigma-Aldrich (formally MilliporeSigma) is an American chemical, life science, and biotechnology company that is owned by the German chemical conglomerate Merck Group. Sigma-Aldrich was created in 1975 by the merger of Sigma Chemical Company a ...

Proteomics

Screens for biologically relevant protein-protein interactions have been performed using Tandem Affinity Purification (TAP) with the SBP-Tag and

Protein A Protein A is a 42 kDa surface protein originally found in the cell wall of the bacteria ''Staphylococcus aureus''. It is encoded by the ''spa'' gene and its regulation is controlled by DNA topology, cellular osmolarity, and a two-component system ...

, for interaction proteomics and transcription factor complexes with the SBP-Tag and

Protein G Protein G is an immunoglobulin-binding protein expressed in group C and G Streptococcal bacteria much like Protein A but with differing binding specificities. It is a ~60-kDA (65 kDA for strain G148 and 58 kDa for strain C40) cell surface prote ...

, for proteins that interact with the

Dengue Virus ''Dengue virus'' (DENV) is the cause of dengue fever. It is a mosquito-borne, single positive-stranded RNA virus of the family ''Flaviviridae''; genus ''Flavivirus''. Four serotypes of the virus have been found, a reported fifth has yet to be co ...

protein DENV-2 NS4A with the SBP-Tag and the Calmodulin Tag. and for proteins that interact with

protein phosphatase 2A Protein phosphatase 2A may refer to: * Protein phosphatase 2 Protein phosphatase 2 (PP2), also known as PP2A, is an enzyme that in humans is encoded by the ''PPP2CA'' gene. The PP2A heterotrimeric protein phosphatase is ubiquitously expressed, ...

(PP2A) with the SBP-Tag and the hemagglutinin (HA)-tag.

Imaging

The SBP-Tag will also bind to streptavidin or streptavidin reagents in solution. Applications of these engineered associations include the visualization of specific proteins within living cells, monitoring of the kinetics of the translation of individual proteins in an in vitro translation system, control of the integration of a multi-spanning membrane protein into the

endoplasmic reticulum The endoplasmic reticulum (ER) is, in essence, the transportation system of the eukaryotic cell, and has many other important functions such as protein folding. It is a type of organelle made up of two subunits – rough endoplasmic reticulum ( ...

by fusing the SBP-Tag to the N-terminal translocation sequence and then halting integration with streptavidin and restarting integration with biotin. Fluorescent streptavidin reagents (e.g. streptavidin-HRP) can be used to visualize the SBP-tag by immunoblotting of SDS-PAGE. Additionally, antibodies to the SBP-tag are available commercially.

Surface plasmon resonance

The SBP-Tag has been used to reversibly immobilize recombinant proteins onto streptavidin-functionalized surfaces thereby permitting interaction assessment such as by

surface plasmon resonance Surface plasmon resonance (SPR) is the resonant oscillation of conduction electrons at the interface between negative and positive permittivity material in a particle stimulated by incident light. SPR is the basis of many standard tools for measu ...

(SPR) techniques with re-use of the functionalized surface. SPR has also been used to compare the SBP-Tag with other streptavidin-binding peptides such as

Strep-tag The Strep-tag system is a method which allows the purification and detection of proteins by affinity chromatography. The Strep-tag II is a synthetic peptide consisting of eight amino acids (Tryptophan, Trp-Serine, Ser-Histidine, His-Proline, Pro-G ...