Protein purification tags: Difference between revisions
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'''Overview of tags added to protein to facilitate their purification''' | '''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 === | === His tag === | ||
| Line 14: | Line 10: | ||
:recovered by affinity to nickel or cobalt column | :recovered by affinity to nickel or cobalt column | ||
<small> | <small> | ||
:[[wikipedia:Polyhistidine-tag|more on the ''His tag'' in the Wikipedia]] | :- [[wikipedia:Polyhistidine-tag|more on the ''His tag'' in the Wikipedia]] | ||
</small> | </small> | ||
| Line 21: | Line 17: | ||
:recovered with specific antibody | :recovered with specific antibody | ||
<small> | <small> | ||
:[[wikipedia:FLAG-tag|- more on ''FLAG-tag'' in the Wikipedia]] | :- [[wikipedia:FLAG-tag|- more on ''FLAG-tag'' in the Wikipedia]] | ||
:- | :- [[Kafatos:His-tagged Protein Purification (denaturing)|protocol for purification of His-tagged protein, Kafatos lab]] | ||
:- [[Wittrup:His-Tag Proteins/Nickel Column Purification (suppliers, troubleshooting, etc.)|protocol for purification of His-tagged protein, Wittrup lab]] | |||
</small> | </small> | ||
=== HA tag === | === HA tag === | ||
:fusion with an epitope derived from the Influenza protein haemagglutinin (HA): e.g. N-YPYDVP-C | |||
:recovery with an HA antibody | |||
<small> | |||
:- [[wikipedia:Hemagglutinin|Wikipedia entry on haemagglutinin]] | |||
:- [http://www.clontech.com/products/detail.asp?tabno=2&catalog_id=631207&page=all Clontech: HA tag] | |||
:- [http://www.roche-applied-science.com/sis/proteinexpression/docs/manual_ha.pdf Roche HA tag vector (PDF)] | |||
</small> | |||
=== 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 | |||
<small> | |||
:- [[wikipedia:MYC|Wikipedia entry on Myc oncogene]] | |||
:- [http://www.miltenyibiotec.com/download/mm366/DS130-092-085.pdf MACS-MYC vector from Miltenyi Biotec (PDF)] | |||
:- [http://www.stratagene.com/products/displayProduct.aspx?pid=242 combined MYC/FLAG tag vector from Stratagene] | |||
</small> | |||
== 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 === | === GST tag === | ||
| Line 32: | Line 48: | ||
:recovery by affinity to substrate glutathione bound to a column, e.g. glutathione sepharose | :recovery by affinity to substrate glutathione bound to a column, e.g. glutathione sepharose | ||
<small> | <small> | ||
:- | :- [http://www6.gelifesciences.com/APTRIX/upp01077.nsf/Content/Products?OpenDocument&parentid=366157&moduleid=164393&zone=Proteomics GST cloning system of GE Healthcare] | ||
:- [[wikipedia:Glutathione S-transferase|Wikipedia entry on GST]] | |||
</small> | </small> | ||
=== biotin - streptavidin === | |||
=== | |||
: | : | ||
<small> | <small> | ||
:- | :- [[wikipedia:XX|YY]] | ||
:- | |||
</small> | </small> | ||
== chemical tags == | == chemical tags == | ||
Revision as of 15:38, 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
- 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
biotin - streptavidin
- - YY
- -
chemical tags
links
Wikipedia
OWW
- example of a protocol for protein purification from bacteria: Protein Expression and Purification
- list but no content of purification methods from Berglund lab: Berglund:Protein Purification: Various Methods
