what protein-protein interaction can lead to irreversible/covalent binding

Zhong Silin via methods%40net.bio.net (by stxsz1 from nottingham.ac.uk)
Fri Nov 14 19:50:48 EST 2008


Hi all
 
I want to find a method to pulldown a target protein A indirectly. (it just has to be indirect) 
 
I am thinking: to express protein A with epitope tag x first (A-x), then express a second protein B with another epitope tag y (B-y). Protein B will interact with tag x and Protein A can then be co-purified using the epitope tag y, which is fused to protein B.
 
For tag x and protein B, I only got 2 pairs of candidates: IgG and protein A/G; N- and C-terminal of the fluorescent protein. Or use the biotin ligase as protein B and fuse the recognition sequence to protein A as tag x.
 
The ideal tag x and protein B also have to be small so they wont interfere with the function of protein A. Their interaction has to be strong as well.
 
I worked on plants and I know our techniques are always a bit behind animal sciences. So you guys must have a better systems to do this. But please keep in mind that this method has to be two-step and indirect!
 
Thanks in advance
SZ 
 

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