GST tag fusion protein--- help please - (Jul/15/2013 )

Hey there,

I've been having a hard time to purify a GST tag protein. I use the pGEX4T-1 vector and so my protein is GST tagged.The fusion protein is partialy soluble.Then I purified my protein ,but there was no activity.Is there any one can help me ?

Is the protein at the approriate MW on a western? Perhaps the GST fusion is impeding enzymatic activity? If this is the case, there are several cleavable linkers you could use.

Thanks,Ahrenhase

I`ve purified the fusion protein by the Glutathione Sepharose ，the sds-page results is right .Now I am tring to cleavage the GST tag by THROMBIN.Wish me good luck!Thank you again!

Good Luck! GST often dimerizes taking your target protein with it. This can interfere with functionality. Cleaving seems like a great idea.

Thank you,Missse
I have tried several times to purify the protein,and I it`s really a hard work .Could you give me some advise about using thrombin to cleavage the FUSION？