FLAG tag PeptideVersatile fusion tag CAS# 98849-88-8 |
Quality Control & MSDS
Number of papers citing our products
Chemical structure
3D structure
Cas No. | 98849-88-8 | SDF | Download SDF |
PubChem ID | 122124 | Appearance | Powder |
Formula | C41H60N10O20 | M.Wt | 1012.97 |
Type of Compound | N/A | Storage | Desiccate at -20°C |
Solubility | H2O : 100 mg/mL (98.72 mM; Need ultrasonic) | ||
Sequence | H-Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lys-OH | ||
Chemical Name | (2S)-6-amino-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[2-[[(2S)-6-amino-2-[[(2S)-2-[[(2S)-2-amino-3-carboxypropanoyl]amino]-3-(4-hydroxyphenyl)propanoyl]amino]hexanoyl]amino]-3-carboxypropanoyl]amino]-3-carboxypropanoyl]amino]-3-carboxypropanoyl]amino]-3-carboxypropanoyl]amino]hexanoic acid | ||
SMILES | C1=CC(=CC=C1CC(C(=O)NC(CCCCN)C(=O)NC(CC(=O)O)C(=O)NC(CC(=O)O)C(=O)NC(CC(=O)O)C(=O)NC(CC(=O)O)C(=O)NC(CCCCN)C(=O)O)NC(=O)C(CC(=O)O)N)O | ||
Standard InChIKey | XZWYTXMRWQJBGX-DQCYEUCQSA-N | ||
Standard InChI | InChI=1S/C41H60N10O20/c42-11-3-1-5-22(45-36(65)24(13-19-7-9-20(52)10-8-19)47-34(63)21(44)14-29(53)54)35(64)48-26(16-31(57)58)38(67)50-28(18-33(61)62)40(69)51-27(17-32(59)60)39(68)49-25(15-30(55)56)37(66)46-23(41(70)71)6-2-4-12-43/h7-10,21-28,52H,1-6,11-18,42-44H2,(H,45,65)(H,46,66)(H,47,63)(H,48,64)(H,49,68)(H,50,67)(H,51,69)(H,53,54)(H,55,56)(H,57,58)(H,59,60)(H,61,62)(H,70,71)/t21-,22-,23-,24-,25-,26?,27-,28-/m0/s1 | ||
General tips | For obtaining a higher solubility , please warm the tube at 37 ℃ and shake it in the ultrasonic bath for a while.Stock solution can be stored below -20℃ for several months. We recommend that you prepare and use the solution on the same day. However, if the test schedule requires, the stock solutions can be prepared in advance, and the stock solution must be sealed and stored below -20℃. In general, the stock solution can be kept for several months. Before use, we recommend that you leave the vial at room temperature for at least an hour before opening it. |
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About Packaging | 1. The packaging of the product may be reversed during transportation, cause the high purity compounds to adhere to the neck or cap of the vial.Take the vail out of its packaging and shake gently until the compounds fall to the bottom of the vial. 2. For liquid products, please centrifuge at 500xg to gather the liquid to the bottom of the vial. 3. Try to avoid loss or contamination during the experiment. |
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Shipping Condition | Packaging according to customer requirements(5mg, 10mg, 20mg and more). Ship via FedEx, DHL, UPS, EMS or other couriers with RT, or blue ice upon request. |
Description | FLAG tag Peptide is a 8 amino acid peptide with an enterokinase-cleavage site used for the purification of recombinant proteins. | |||||
Targets | anti-flag M2 antibody |
FLAG tag Peptide Dilution Calculator
FLAG tag Peptide Molarity Calculator
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A fusion tag called FLAG and consisting of eight amino acids Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lys including an enterokinase-cleavage site, was specifically designed for immunoaffinity chromatography. It allows elution under non-denaturing conditions. Several antibodies against this peptide have been developed. One antibody, denoted as M1, binds the peptide in the presence of bivalent metal cations, preferably Ca+. Elution is effected by chelating agents. Another strategy is competitive elution with excess of free FLAG peptide. Antibodies M2 and M5 are applied in this procedure1.
The added marker segment is that it should not interfere with the native folding of proteins to which it is attached. Secondly, the marker peptide sequence should be water-soluble and should retain a high degree of exposure on the surface of the protein, so that it can readily interact with its ligand. It should also be suitable for a mild and inexpensive affinity purification procedure. Finally, an easy removal of the marker peptide leading to a native product is also advantageous. Due to this small size, the marker peptide can be encoded by a single synthetic oligonucleotide. It is known that aromatic amino acids are the major factors in antigen–antibody interactions.
Lys at position 3 in the marker sequence leads to a hexapeptide sequence LysAspAspAspAspLys, which ensures a maximum value on the hydrophilicity scale according to Hopp and Woods2 . Such hydrophilic sequences have been shown to express strong antigenicity and are thus likely to adopt a highly exposed conformation in the three-dimensional folding of proteins3. Another virtue of FLAG is that the longest trypsinogen prosequences are of this length. This enables the removal of the tag and the production of an authentic N-terminus of the fusion protein partner by enterokinase treatment. The FLAG peptide can be fused to either the N- or C-terminus of a given fusion protein. Nevertheless, the N-terminal fusion has several advantages. Inhibition ELISA experiments showed that the anti-Flag antibody M1 binds three to four orders of magnitude better under conditions where the a-amino group of the first amino acid is freely accessible4.
The FLAG marker peptide fusion system comprises a unique and widely useful technique for protein identification and purification. Elution of the fusion protein can be accomplished either by antibody-mediated affinity chromatography in a calcium-dependent manner, by lowering the pH, or by competitive elution with synthetic peptides1. Although highly selective, the binding capacities are low, making scale-up a costly undertaking. In addition to cost and low capacity, large-scale immunoaffinity chromatography, applied to the production of therapeutic proteins has several disadvantages: ligand leakage, instability, and need for validation of antibody production. The stability of the affinity chromatography column depends on the nature and source of the crude extracts. Furthermore, the FLAGe tag, designed to be immunogenic, must be removed from therapeutic proteins. In most cases, this can be accomplished with enterokinase. However, contaminating proteases may also produce undesired cleavages. Despite these drawbacks, the FLAG fusion is useful in research and development: FLAG proteins can be readily purified and assayed by ELISA or any other immunochemical detection method, thus expediting the raising of antisera against a desired protein and characterization studies.
References:
1. A. Einhauer, A. Jungbauer. The FLAG peptide, a versatile fusion tag for the purification of recombinant proteins. J. Biochem. Biophys. Methods 49 2001 455–465.
2. Hopp TP, Woods KR. Prediction of protein antigenic determinants from amino acid sequences. Proc Natl Acad Sci U S A 1981;78:3824–8.
3. Hopp TP. Protein surface analysis: methods for identifying antigenic determinants and other interaction sites. J Immunol Methods 1986;88:1–18.
4. Power BE, Ivancic N, Harley VR, Webster RG, Kortt AA, Irving RA, et al. High-level temperature-induced synthesis of an a
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An improved affinity tag based on the FLAG peptide for the detection and purification of recombinant antibody fragments.[Pubmed:7530459]
Biotechniques. 1994 Oct;17(4):754-61.
The commercially available monoclonal antibodies M1 and M2 were raised against and bind the FLAG sequence DYKDDDDK with high specificity. Using the calcium-dependent M1 antibody and the FLAG tag attached to the N terminus of various fragments of the antibody McPC603 expressed in Escherichia coli, we found that the M1 antibody binds with almost the same affinity to a much shorter version of this sequence (DYKD). Since most antibody light chains start with an aspartate, the addition of only three additional amino acids to the N terminus is sufficient to detect and quantify the expressed antibody fragments using standard immunological methods. Similarly, the heavy chain can be detected specifically with the sequence DYKD, which requires four additional amino acids since most heavy chains do not start with Asp. The signal sequence of both chains that is necessary for the transport of the chains to the periplasm of E. coli is processed correctly. Furthermore, we investigated the influence of the amino acid at the fifth position of the FLAG sequence on the binding affinity of the M1 antibody and found that a glutamate at this position increased the sensitivity in Western blots sixfold over the original long FLAG sequence containing an aspartate residue at this position. Together, the improved FLAG is a versatile tool for both sensitive detection and one-step purification of recombinant proteins.
The FLAG peptide, a versatile fusion tag for the purification of recombinant proteins.[Pubmed:11694294]
J Biochem Biophys Methods. 2001 Oct 30;49(1-3):455-65.
A fusion tag, called FLAG and consisting of eight amino acids (AspTyrLysAspAspAspAspLys) including an enterokinase-cleavage site, was specifically designed for immunoaffinity chromatography. It allows elution under non-denaturing conditions [Bio/Technology, 6 (1988) 1204]. Several antibodies against this peptide have been developed. One antibody, denoted as M1, binds the peptide in the presence of bivalent metal cations, preferably Ca(+). Elution is effected by chelating agents. Another strategy is competitive elution with excess of free FLAG peptide. Antibodies M2 and M5 are applied in this procedure. Examples demonstrating the versatility, practicability and limitations of this technology are given.