Protein tag

Protein tags are peptide sequences genetically grafted onto a recombinant protein. Often these tags are removable by chemical agents or by enzymatic means, such as proteolysis or intein splicing. Tags are attached to proteins for various purposes.

Affinity tags are appended to proteins so that they can be purified from their crude biological source using an affinity technique. These include chitin binding protein (CBP), maltose binding protein (MBP), Strep-tag[1] and glutathione-S-transferase (GST). The poly(His) tag is a widely used protein tag, which binds to metal matrices.

Solubilization tags are used, especially for recombinant proteins expressed in chaperone-deficient species such as E. coli, to assist in the proper folding in proteins and keep them from precipitating. These include thioredoxin (TRX) and poly(NANP). Some affinity tags have a dual role as a solubilization agent, such as MBP, and GST.

Chromatography tags are used to alter chromatographic properties of the protein to afford different resolution across a particular separation technique. Often, these consist of polyanionic amino acids, such as FLAG-tag.

Epitope tags are short peptide sequences which are chosen because high-affinity antibodies can be reliably produced in many different species. These are usually derived from viral genes, which explain their high immunoreactivity. Epitope tags include V5-tag, Myc-tag, HA-tag, Spot-tag and NE-tag. These tags are particularly useful for western blotting, immunofluorescence and immunoprecipitation experiments, although they also find use in antibody purification.

Fluorescence tags are used to give visual readout on a protein. GFP and its variants are the most commonly used fluorescence tags. More advanced applications of GFP include using it as a folding reporter (fluorescent if folded, colorless if not).

Protein tags may allow specific enzymatic modification (such as biotinylation by biotin ligase) or chemical modification (such as reaction with FlAsH-EDT2 for fluorescence imaging). Often tags are combined, in order to connect proteins to multiple other components. However, with the addition of each tag comes the risk that the native function of the protein may be abolished or compromised by interactions with the tag. Therefore, after purification, tags are sometimes removed by specific proteolysis (e.g. by TEV protease, Thrombin, Factor Xa or Enteropeptidase).

List of protein tags

(See Proteinogenic amino acid#Chemical properties for the A-Z amino-acid codes)

Peptide tags

  • AviTag, a peptide allowing biotinylation by the enzyme BirA and so the protein can be isolated by streptavidin (GLNDIFEAQKIEWHE)
  • Calmodulin-tag, a peptide bound by the protein calmodulin (KRRWKKNFIAVSAANRFKKISSSGAL)
  • polyglutamate tag, a peptide binding efficiently to anion-exchange resin such as Mono-Q (EEEEEE)
  • E-tag, a peptide recognized by an antibody (GAPVPYPDPLEPR)
  • FLAG-tag, a peptide recognized by an antibody (DYKDDDDK)[2]
  • HA-tag, a peptide from hemagglutinin recognized by an antibody (YPYDVPDYA)[3]
  • His-tag, 5-10 histidines bound by a nickel or cobalt chelate (HHHHHH)
  • Myc-tag, a peptide derived from c-myc recognized by an antibody (EQKLISEEDL)
  • NE-tag, a novel 18-amino-acid synthetic peptide (TKENPRSNQEESYDDNES) recognized by a monoclonal IgG1 antibody, which is useful in a wide spectrum of applications including Western blotting, ELISA, flow cytometry, immunocytochemistry, immunoprecipitation, and affinity purification of recombinant proteins [4]
  • S-tag, a peptide derived from Ribonuclease A (KETAAAKFERQHMDS)
  • SBP-tag, a peptide which binds to streptavidin (MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP)[5][6]
  • Softag 1, for mammalian expression (SLAELLNAGLGGS)
  • Softag 3, for prokaryotic expression (TQDPSRVG)
  • Spot-tag, a peptide recognized by a nanobody (PDRVRAVSHWSS) for immunoprecipitation, affinity purification, immunofluorescence and super resolution microscopy
  • Strep-tag, a peptide which binds to streptavidin or the modified streptavidin called streptactin (Strep-tag II: WSHPQFEK)[1]
  • TC tag, a tetracysteine tag that is recognized by FlAsH and ReAsH biarsenical compounds (CCPGCC)
  • Ty tag (EVHTNQDPLD)
  • V5 tag, a peptide recognized by an antibody (GKPIPNPLLGLDST)[7]
  • VSV-tag, a peptide recognized by an antibody (YTDIEMNRLGK)
  • Xpress tag (DLYDDDDK)

Covalent peptide tags

  • Isopeptag, a peptide which binds covalently to pilin-C protein (TDKDMTITFTNKKDAE)[8]
  • SpyTag, a peptide which binds covalently to SpyCatcher protein (AHIVMVDAYKPTK)[9]
  • SnoopTag, a peptide which binds covalently to SnoopCatcher protein (KLGDIEFIKVNK)[10]
  • SnoopTagJr, a peptide which binds covalently to DogTag, mediated by SnoopLigase (KLGSIEFIKVNK)[11]
  • DogTag, a peptide which covalently binds to SnoopTagJr, mediated by SnoopLigase (DIPATYEFTDGKHYITNEPIPPK)[12]

Protein tags

  • BCCP (Biotin Carboxyl Carrier Protein), a protein domain biotinylated by BirA enabling recognition by streptavidin
  • Glutathione-S-transferase-tag, a protein which binds to immobilized glutathione
  • Green fluorescent protein-tag, a protein which is spontaneously fluorescent and can be bound by nanobodies
  • HaloTag, a mutated bacterial haloalkane dehalogenase that covalently attaches to a reactive haloalkane substrate, this allows attachment to a wide variety of substrates.
  • Maltose binding protein-tag, a protein which binds to amylose agarose[13]
  • Nus-tag
  • Thioredoxin-tag
  • Fc-tag, derived from immunoglobulin Fc domain, allow dimerization and solubilization. Can be used for purification on Protein-A Sepharose
  • Designed Intrinsically Disordered tags containing disorder promoting amino acids (P,E,S,T,A,Q,G,..)[14]

Others

    Applications

    References

    1. 1 2 Schmidt, Thomas G.M.; Koepke, Jürgen; Frank, Ronald; Skerra, Arne (1996). "Molecular Interaction Between the Strep-tag Affinity Peptide and its Cognate Target, Streptavidin". Journal of Molecular Biology. 255 (5): 753–66. doi:10.1006/jmbi.1996.0061. PMID 8636976.
    2. ↑ Einhauer, A.; Jungbauer, A. (2001). "The FLAG™ peptide, a versatile fusion tag for the purification of recombinant proteins". Journal of Biochemical and Biophysical Methods. 49 (1–3): 455–65. doi:10.1016/S0165-022X(01)00213-5. PMID 11694294.
    3. ↑ Prakriya, Murali; Feske, Stefan; Gwack, Yousang; Srikanth, Sonal; Rao, Anjana; Hogan, Patrick G. (2006). "Orai1 is an essential pore subunit of the CRAC channel". Nature. 443 (7108): 230–3. Bibcode:2006Natur.443..230P. doi:10.1038/nature05122. PMID 16921383.
    4. ↑ Ho, Philip WL.; Tse, Zero HM.; Liu, HF.; Lu, S.; Ho, Jessica WM.; Kung, Michelle HW.; Ramsden, David B.; Ho, SL. (2013). "Assessment of cellular estrogenic activity based on estrogen receptor-mediated reduction of soluble-form catechol-O-methyltransferase (COMT) expression in an ELISA-based system". PLoS ONE. 8 (9): e74065. Bibcode:2013PLoSO...874065H. doi:10.1371/journal.pone.0074065. PMC 3765251. PMID 24040167.
    5. ↑ Keefe, Anthony D.; Wilson, David S.; Seelig, Burckhard; Szostak, Jack W. (2001). "One-Step Purification of Recombinant Proteins Using a Nanomolar-Affinity Streptavidin-Binding Peptide, the SBP-Tag". Protein Expression and Purification. 23 (3): 440–6. doi:10.1006/prep.2001.1515. PMID 11722181.
    6. ↑ Gelerter, Bruce (June 11, 2014). "PEMF For Treatment Of Corneal Disorders And Injuries".
    7. ↑ McNutt, Markey C.; Lagace, Thomas A.; Horton, Jay D. (2007). "Catalytic Activity is Not Required for Secreted PCSK9 to Reduce Low Density Lipoprotein Receptors in HepG2 Cells". Journal of Biological Chemistry. 282 (29): 20799–803. doi:10.1074/jbc.C700095200. PMID 17537735.
    8. ↑ Zakeri, Bijan; Howarth, Mark (2010). "Spontaneous Intermolecular Amide Bond Formation between Side Chains for Irreversible Peptide Targeting". Journal of the American Chemical Society. 132 (13): 4526–7. doi:10.1021/ja910795a. PMID 20235501.
    9. ↑ Zakeri, Bijan; Fierer, Jacob O.; Celik, Emrah; Chittock, Emily C.; Schwarz-Linek, Ulrich; Moy, Vincent T.; Howarth, Mark (2012). "Peptide tag forming a rapid covalent bond to a protein, through engineering a bacterial adhesin". Proceedings of the National Academy of Sciences. 109 (12): E690–7. Bibcode:2012PNAS..109E.690Z. doi:10.1073/pnas.1115485109. PMC 3311370. PMID 22366317.
    10. ↑ Veggiani, Gianluca; Nakamura, Tomohiko; Brenner, Michael; Gayet, Raphael; Yan, Jun; Robinson, Carol; Howarth, Mark (2016). "Programmable polyproteams built using twin peptide superglues". Proceedings of the National Academy of Sciences. 113 (5): 1202–7. Bibcode:2016PNAS..113.1202V. doi:10.1073/pnas.1519214113. PMID 26787909.
    11. ↑ Buldun, Can M.; Jean, Jisoo X.; Bedford, Michael R.; Howarth, Mark (14 February 2018). "SnoopLigase Catalyzes Peptide–Peptide Locking and Enables Solid-Phase Conjugate Isolation". Journal of the American Chemical Society. doi:10.1021/jacs.7b13237.
    12. ↑ Buldun, Can M.; Jean, Jisoo X.; Bedford, Michael R.; Howarth, Mark (14 February 2018). "SnoopLigase Catalyzes Peptide–Peptide Locking and Enables Solid-Phase Conjugate Isolation". Journal of the American Chemical Society. doi:10.1021/jacs.7b13237.
    13. ↑ Bedouelle, Hugues; Duplay, Pascale (Feb 1988). "Production in Escherichia coli and one-step purification of bifunctional hybrid proteins which bind maltose. Export of the Klenow polymerase into the periplasmic space". Eur J Biochem. 171 (3): 541–549. doi:10.1111/j.1432-1033.1988.tb13823.x. PMID 3278900.
    14. ↑ Minde, David P; Halff, Els F; Tans, Sander (2013). "Designing disorder: Tales of the unexpected tails". Intrinsically Disordered Proteins. 1: 5–15. doi:10.4161/idp.26790.
    This article is issued from Wikipedia. The text is licensed under Creative Commons - Attribution - Sharealike. Additional terms may apply for the media files.