Tandem Affinity Purification

Tandem Affinity Purification

Tandem affinity purification (TAP) is a technique for studying protein-protein interactions. It involves creating a fusion protein with a designed piece, the TAP tag, on the end.

In the original version of the technique, the protein of interest with the TAP tag first binds to beads coated with IgG, the TAP tag is then broken apart by an enzyme, and finally a different part of the TAP tag binds reversibly to beads of a different type. After the protein of interest has been washed through two affinity columns, it can be examined for binding partners.

The original TAP method involves the fusion of the TAP tag to the C-terminus of the protein under study. The TAP tag consists of calmodulin binding peptide (CBP) from the N-terminal, followed by tobacco etch virus protease (TEV protease) cleavage site and Protein A, which binds tightly to IgG. The relative order of the modules of the tag is important because Protein A needs to be at the extreme end of the fusion protein so that the entire complex can be retrieved using an IgG matrix.

Many other tag combinations have been proposed since the TAP principle was first published.

Read more about Tandem Affinity Purification:  Variant Tags, History, Process, Advantages, Disadvantages, Suitability, Applications, Other Epitope-tag Combinations

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