Protein purification tags: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
(*** page creation *** unfinished save 3) |
(*** page creation *** finished - up to you to extend & improve ;-)) |
||
| Line 5: | Line 5: | ||
A stretch of amino acids is added to the protein and enables the recovery of the labelled protein by its unique affinity. Usually its easiest to add the tag to either end of the protein to ensure its accessibility and not to disturb the protein folding. | A stretch of amino acids is added to the protein and enables the recovery of the labelled protein by its unique affinity. Usually its easiest to add the tag to either end of the protein to ensure its accessibility and not to disturb the protein folding. | ||
=== His tag === | === His tag / IMAC === | ||
:6+ histidine (His) residues | :6+ histidine (His) residues | ||
| Line 22: | Line 22: | ||
</small> | </small> | ||
=== HA tag === | === HA tag=== | ||
:fusion with an epitope derived from the Influenza protein haemagglutinin (HA): e.g. N-YPYDVP-C | :fusion with an epitope derived from the Influenza protein haemagglutinin (HA): e.g. N-YPYDVP-C | ||
:recovery with an HA antibody | :recovery with an HA antibody | ||
| Line 51: | Line 51: | ||
:- [[wikipedia:Glutathione S-transferase|Wikipedia entry on GST]] | :- [[wikipedia:Glutathione S-transferase|Wikipedia entry on GST]] | ||
</small> | </small> | ||
== chemical tags == | |||
=== biotin - streptavidin === | === biotin - streptavidin === | ||
: | :linking amino acids, often lysines, to cofactor and vitamin biotin | ||
:purification by tight binding to streptavidin-agarose or -beads | |||
<small> | <small> | ||
:- [[wikipedia: | :- [[wikipedia:Biotin|Wikipedia entry biotin]] | ||
:- | :- [[wikipedia:Streptavidin|Wikipedia entry streptavidin]] and [[wikipedia:Avidin|avidin]] | ||
:- [http://www.iba-go.com/iba_us/prottools/prot_p_bur.html Biotin purification reagents from IBA] | |||
</small> | </small> | ||
== links == | == links == | ||
=== Wikipedia === | === Wikipedia === | ||
* [[wikipedia:Protein tag|Protein tag]] | * [[wikipedia:Protein tag|Protein tag]] | ||
* [[wikipedia:Protein_purification#Purification_of_a_tagged_protein|Purification of a tagged protein]] | * [[wikipedia:Protein_purification#Purification_of_a_tagged_protein|Purification of a tagged protein]] | ||
=== OWW === | === OWW === | ||
* example of a protocol for protein purification from bacteria | * [[Protein Expression and Purification|example of a protocol for protein purification from bacteria]] | ||
* [[Berglund:Protein Purification: Various Methods|list but no content of purification methods from Berglund lab]] | |||
=== other === | |||
* [http://www.the-scientist.com/article/display/12881/ article in ''The Scientist'' on protein purification with tags, 2002] | |||
Revision as of 16:07, 3 March 2007
Overview of tags added to protein to facilitate their purification
Amino acid tags
A stretch of amino acids is added to the protein and enables the recovery of the labelled protein by its unique affinity. Usually its easiest to add the tag to either end of the protein to ensure its accessibility and not to disturb the protein folding.
His tag / IMAC
- 6+ histidine (His) residues
- recovered by affinity to nickel or cobalt column
FLAG tag
- N-DYKDDDDK-C and others
- recovered with specific antibody
- - - more on FLAG-tag in the Wikipedia
- - protocol for purification of His-tagged protein, Kafatos lab
- - protocol for purification of His-tagged protein, Wittrup lab
HA tag
- fusion with an epitope derived from the Influenza protein haemagglutinin (HA): e.g. N-YPYDVP-C
- recovery with an HA antibody
MYC tag
- fusion with an epitope derived from the human proto-oncoprotein MYC: e.g. N-ILKKATAYIL-C, N-EQKLISEEDL-C
- recovery with an MYC antibody
- - Wikipedia entry on Myc oncogene
- - MACS-MYC vector from Miltenyi Biotec (PDF)
- - combined MYC/FLAG tag vector from Stratagene
Protein tags
Rather than adding only a few amino acids a whole protein is fused to the protein to be purified or detected. The affinity of the attached protein enables the recovery of the artificial fusion protein. As for the peptides, the protein tag is added to either end of the target protein.
GST tag
- fusion with the small glutathione-S-transferase (GST; 26 kDa)
- recovery by affinity to substrate glutathione bound to a column, e.g. glutathione sepharose
chemical tags
biotin - streptavidin
- linking amino acids, often lysines, to cofactor and vitamin biotin
- purification by tight binding to streptavidin-agarose or -beads
- - Wikipedia entry biotin
- - Wikipedia entry streptavidin and avidin
- - Biotin purification reagents from IBA
links
Wikipedia
OWW
- example of a protocol for protein purification from bacteria
- list but no content of purification methods from Berglund lab
