Hey dears,
I'm getting ready to create a genetic construction where I'll tag my protein with three flags. I'm aware that I must place it in either the C or N terminal, but I'm not sure which terminal would be best given each consideration ?. Additionally,I want to know whether there is a way to confirm how this 3-flag tag affects the structure and function of my protein ?
I would be very grateful for your help
Thank you
Ayoub BOUHADADA . PhD student at Québec Trois-Rivières university
We have had FLAG on either end of our proteins. The main reason to have it on the C-terminus is if you are planning to secrete the protein and are worried that the N-terminal FLAG may interfere with secretion or expression. Since the FLAG is pretty small, it is usually safe to assume that it will not affect your protein, but if you have any structural data to show the location of the tag relative to the active or other functional sites, then you should use it. Another option is to use a cleavable tag with a protease and compare activities, or you can use a self-removing tag like the IMPACT system or the iCapTag from Protein Capture Science (disclaimer is that Protein Capture Science is my company, but we have a self-removing tag system commercially available now).
David W Wood thank you for all this advices I will try to do that.
thank you !
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