How can I select an expression tag that will not interfere with protein function?
(Thread 21630) Q1 I want to express two proteins in cultured cells and then detect the level of expression. Both proteins are uninvestigated, do not have corresponding antibodies available, and their function is unknown. Can someone suggest a tag that will be least likely to interfere with protein function? And how do I determine if the function has been preserved if I don’t know what function to assay? A1 I have had very good success with the V5 tag and occasionally also with His and Flag tags. The advantage of the His tag is that there are columns available that allow for easy purification of the soluble protein. A2 His tags are great for purification, but they are notorious for causing aggregation. I recommend the FLAG tag for a small protein. A3 Try expressing the protein with an N-terminal His tag at 25°C and also as a fusion to something like MBP or SUMO. Do these experiments in parallel and you should get at least some protein to work with fairly quickly. A4 If you want to expre
