How does a Strep-tag work?
The Strep-tag ®II binds specifically to the engineered streptavidins, Strep-Tactin ® and Strep-Tactin ®XT, by occupying the binding pocket of the natural ligand biotin. Hence, the interaction is easily reversible by excessive addition of the competitor.
How do you purify Strep-tagged proteins?
Agarose-based matrices with the coupled Strep-Tactin ligand are the resins of choice, with a binding capacity of up to 9 mg ml(-1). For purification of lower amounts of Strep-tagged proteins, the use of Strep-Tactin magnetic beads is suitable.
What is Strep-tag amino acid sequence?
The Strep-tag II is an eight-residue minimal peptide sequence (Trp-Ser-His-Pro-Gln-Phe-Glu-Lys) that exhibits intrinsic affinity toward streptavidin and can be fused to recombinant proteins in various fashions.
Does Strep-tag bind to streptavidin?
The Strep-tag is a selected nine-amino acid peptide (AWRHPQFGG) that displays intrinsic binding affinity towards streptavidin and has been used as an affinity tag for recombinant proteins.
What is Twin Strep-tag?
Twin-Strep-tag® (TST) binds to a single Strep-Tactin® tetramer. • TST arrangement provides enhanced affinity by off-rate reduction. • TST binds reversibly and enables efficient competitive elution.
How big is a flag tag?
2. Alternatives to the Flag-tag
| Feature | Flag® Tag |
|---|---|
| Size of the tag in Da | 1012.0 Da |
| Affinity matrix | Anti Flag® antibody (approx. 150 kDa) |
| Elution conditions | Flag® peptide, low pH, or tag cleavage |
| Specificity of interaction (KD) | 100 nM (3) |
What is Tactin?
Strep•Tactin® protein, a derivative of streptavidin, has an optimized binding pocket that is specific for the Strep•Tag II peptide. It is a stable protein that has chemical properties similar to those of streptavidin, making it compatible with a broad range of detergents, chelators, salt, and redox conditions.
Does Strep II tag bind to streptavidin?
The strep-tag has the property of binding to streptavidin competitively with biotin. This behavior permits the use of very gentle conditions for the elution of a bound strep-tag fusion protein from the streptavidin affinity column, just by applying a diluted solution of biotin or one of its chemical derivatives.
What is Twin strep-tag?
What is SUMO tag?
SUMO Tag Definition Sumo tag is most frequently used as N-end fusion sequence in yeast to increase the expression and solubility of the desired recombinant protein. SUMO proteins are similar to ubiquitin in their folded structure but possess only about 20% homology to the amino acid sequence of ubiquitin.
Why are proteins flag tagged?
If there is no antibody against a given protein, adding a FLAG-tag to a protein allows the protein to be studied with an antibody against the FLAG sequence.
How do you flag a protein?
(D=Aspartic acid; K=Lysine; Y=Tyrosine). That brings the total size of the tag to 1012.9 dalton or roughly 1 kDa(1). The Flag-tag can be added either to the N-terminus or the C-terminus of a protein(1), respectively….2. Alternatives to the Flag®-tag.
| Features of the FLAG® tag: | |
|---|---|
| Specificity of interaction (KD) | 100 nM (3) |
What is Desthiobiotin?
Desthiobiotin is a modified form of biotin that displaces Strep-tag ®II and Twin-Strep-tag ® fusion proteins from the modified biotin-binding pocket of Strep-Tactin ® in a competitive manner, resulting in a mild elution. Desthiobiotin powder can be used for preparation of Strep-Tactin ® elution buffer, Buffer E.
What is an HA tag used for?
The HA (hemagglutinin) tag is derived from the human influenza virus HA protein. It is well characterized and is used extensively as a general antibody epitope tag. HA tag antibodies provide a dependable method for the detection and purification of tagged target proteins without a protein-specific antibody or probe.
What is Avitag?
Avi-tag Biotinylated Proteins are proteins that feature biotinylation at a single site contained within the Avi-tag, a unique 15 amino acid peptide. The DNA sequence coding for the Avi-tag is inserted during cloning and contained in the final expression vector construct.
Why is SUMO tag used?
Sumo tag is most frequently used as N-end fusion sequence in yeast to increase the expression and solubility of the desired recombinant protein. SUMO proteins are similar to ubiquitin in their folded structure but possess only about 20% homology to the amino acid sequence of ubiquitin.
How do I get rid of SUMO tag?
SUMO fusion proteins can be cleaved to remove the SUMO moiety using SUMO-specific proteases such as Ulp1. Here, we describe the use of vectors for the expression of recombinant proteins in E. coli as fusions to the Drosophila SUMO protein.
What is a flagged protein?
The Flag® tag, also known as the DYKDDDDK-tag, is a popular protein tag that is commonly used in affinity chromatography and protein research for over 20 years now (6,7,8,9,10,11). As its second name suggests the tag consists of an amino acid sequence DYKDDDDK. (D=Aspartic acid; K=Lysine; Y=Tyrosine).