Fusion Tag /Protein

Definition:
A protein or a peptide located either on the C- or N- terminal of the target protein, which facilitates one or several of the following characteristics:


Notice: The fusion protein must be continuous with the target protein -  the same open reading frame must be maintained. Stop codons between the target protein and the fusion partner must be omitted.
 
 

Frequently Used Fusion Tags

Frequently Used Fusion Proteins

Additional Reading
Some Commercially Available Plasmids containing Various Fusion Tags/Proteins 

Back to Vector Image



 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Frequently Used Fusion Tags
Name 
(Link to sequence)
Uses
Location relative to target Protein 
(N- or C-Terminal or Internal)
Further Documentation
Related Procedures
(See also Dr. Lebendiker's site)
His Tag 
(6,8, or 10 amino acids)
  • Purification - Affinity Column: His tag binds very tightly [Kd~10-13M) to immobilized divalent cations [e.g Ni+2, Cu+2, Zn+2]
  • Detection - Western Blot
N,C, I
 
  •  NOVAGEN: pET  manual (including methods of purifying many tags)   (pdf
  • NOVAGEN: Ni-NTA Hi-Bind Resins Protocols (pdf
  • QUIAGEN: Ni-NTA purification manual (pdf
  • QUIAGEN: NiNTA cell lysis under nature conditions 
  • CLONTECH: Talon Resin Protocol (pdf
  • CLONTECH: Talon Products (pdf
  • AMERSHAM BIOSCIENCES Hi Tarp Chelating (pdf
  • ACTIVE MOTIVE: Ni-Ted Profile (pdf
  • AFFILAND: Metal Purification 
  • Small scale His-Tag purification under nature conditions
  • Small scale His-Tag purification under denaturative conditions
  • Calbiochem - His Bind Purification Kits
  •  T7 Tag
    (11-16 amino acids)
    • Detection: Western Blot, Immunoprecipitation
    • Purification: Affinity Column (denaturing low pH elution needed)
    • Possibly enhanced expression levels since the T7-tag is derived from the T7 gene 10 which is the naturally most abundant phage T7 gene product.
     N, I
     
     NOVAGEN T7.Tag 
    Affinity Purification products  
    (pdf)
     S-Tag 
    (15 amino acids)
    • Detection: Western Blot, 
    • quantitative Assay RNAse S assay possible for quantitative assay of expression levels.
    • Purification: Affinity Column
     N, C, I
    S•Tag : A Multipurpose Fusion Peptide for Recombinant Proteins (pdf)
    S-Tag system (Novagen) (pdf)  
    Flag- Tag
    (8 amino acids)
    •  Purification
     N, C
     
    Sigma's Flag Protein Expression System 
     HA Tag 
    (9 mino acids)
    •  Detection
     N
     
     
     V5 Epitope
    (15 amino acids)
     
     
     
     Pel B
    •  Localization: secretion signal
    N
       
     Xpress Epitope
    (8 amino acids)
           

    Back to Fusion Tag/Protein Related Material

    Back to Vector Image

     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     

    Frequently Used Proteins
    Name 
    (Link to sequence)
    Uses
    Location relative to target Protein 
    (N- or C-Terminal or Internal)
    Further Documentation
    Related Procedures
    (See also Dr. Lebendiker's site)
     GST 
    (Glutathione S Transferase)
    (223amino acids, 26 kDa)
    • Solubility (lesser quality than Nus A or MBP) [N-term only]
    • Purification: glutathione affinity or GST antibody purification
    • Detection: Western Blot, Quantitative Assay (based on enzymatic activity)
     N, C
    Endogenous glutathione-binding proteins of insect cell lines: characterization and removal from glutathione S-transferase (GST) fusion proteins.
     
  •  CLONTECH GST products Handbook (pdf
  • AMERSHAM BIOSCIENCES-GST fusion protein Handbook (pdf) 
  •  NOVAGEN GST.Bind Kits (pdf
  • Small scale GST-fusion purification under nature conditions
  • MBP (Maltose Binding Protein)
    (40kDa)
    • Solubility (better quality than GST or Trx) [N-term only]
    • Purification: Amylose affinity purification with maltose elution
    N
    1. E-coli MBP is uncommonly effective at promoting the solubilityof polypeptides to which it is fused
    2.   Smyth DR, Mrozkiewicz MK, McGrath WJ, Listwan P, Kobe B. Crystal structures of fusion proteins with large-affinity tags. Protein Sci. 2003 Jul;12(7):1313-22. (reccommends addition of 3-6 ala-linker that stabilizes connection between target protein and MBP)
    3. Routzahn KM, Waugh DS., Differential effects of supplementary affinity tags on the solubility of MBP fusion proteins. J Struct Funct Genomics. 2002;2(2):83-92. 
  •  NEW ENGLAND BIOLAB: pMAL protein Fusion and Purification System

  •  
     Nus A
    (495 amino acids)
    •  Solubility  [N-term only]
     N
    Expression of soluble heterologous proteins via fusion with NusA protein (pdf)  
     CBP (Calmodulin Binding Peptide)
    (4kDa)
    • Purification
    • Detection
    N, C 
     

    STRATAGENE: CBP Calmodulin-Binding Peptide Affinity Tag System (pdf)

     GFP
    (220 amino acids)
    •  Detection: Immunofluoresence. Used as possible refoldind tag.
    N, C 
    Rapid protein-folding assay using GFP
     
     Thioredoxin
    (109amino acids, 11.7 kDa)
    • Solubility (lesser quality than Nus A or MBP) [N-term only]
    • Detection
    • Affinity purification with phenylarsine oxide-modified (ThioBond) resin
     N, C
     

    Using Rabbit Anti-Thioredoxin (fusion protein) Antibodies .
    (Alpha Diagnostic International, Inc)

    Mistic
    • Solubility (high level production of membrane proteins) 
    N
    Roosild TP, Greenwald J, Vega M, Castronovo S, Riek R, Choe S., NMR structure of Mistic, a membrane-integrating protein for membrane protein expressionScience. 2005 ;307(5713):1317-21 (PDF  
    Sumo
    • Solubility (high level production of membrane proteins)
    N
    1. Zuo X, Li S, Hall J, Mattern MR, Tran H, Shoo J, Tan R, Weiss SR, Butt TR. Enhanced expression and purification of membrane proteins by SUMO fusion in Escherichia coli. J Struct Funct Genomics. 2005;6(2-3):103-11 (PDF)
    2. SUMO fusion technology for enhanced protein expression and purification in prokaryotes and eukaryotes.
     
    DSBa
    • Solubility  - better formation of S-S bonds.
    N

    1. Folding mechanisms of periplasmic proteins.
    2. How proteins form disulfide bonds.

     
    DSBc
    • Solubility  - better formation of S-S bonds.
    N
    Kurokawa Y, Yanagi H, Yura T. Overproduction of bacterial protein disulfide isomerase (DsbC) and its modulator (DsbD) markedly enhances periplasmic production of human nerve growth factor in Escherichia coli. J Biol Chem. 2001 Apr 27;276(17):14393-9 (PDF)  

    Back to Fusion Tag/Protein Related Material

    Back to Vector Image
     


     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     
     

    Additional Reading

    Back to Fusion Tag/Protein Related Material

    Back to Vector Image



















































    .