Difference between revisions of "Part:BBa K2586004"

Revision as of 09:21, 11 September 2018

Strep-Tag-aroE: aroE linked to a Strep-Tag

This part is the combination of <a href="https://parts.igem.org/Part:BBa_K2586003">BBa_K2586003</a> with a Strep-Tag.
The Strep-protein can be used in SPINE (Strep-protein interaction experiments). SPINE is a powerful tool to detect protein-protein interactions in vivo (1). The bait protein (aroE in this case) has to be expressed with a Strep-tag under the conditions when the potential interaction partners are presumably present in the cells. The addition of formaldehyde (FA), which easily permeates the cell wall and membrane, links the bait protein to its potential interaction partners.
The bait protein together with its potential interaction partners can then be isolated using a Strep-Tactin sepharose column by eluting with desthiobiotin- or biotin-containing buffer.
The cross-links between the bait protein and the potential interaction partner can be cleaved by heating the samples in Laemmli buffer. Finally, the co-purified interaction partner(s) can be separated by SDS PAGE, identified in mass spectrometry and Western Blotting.

Sequence and Features