Dear Bert You made the comment a few weeks ago not to boil helical membrane proteins for SDS-PAGE. Could i please ask, does this also apply to type I membrane proteins that only have a single a-helix, or is it just membrane proteins that are predominantly helical? Thanks Rich
On 22 February 2017 at 19:18, Bert Van-Den-Berg < bert.van-den-b...@newcastle.ac.uk> wrote: > like others I'm not clear why you care where your protein runs on > SDS-PAGE. I think the band you're seeing is in fact the tetramer, > suggesting your protein (like KcsA) is very stable. Helical membrane > proteins often migrate faster than expected (by their Mw) on SDS-PAGE. > > Also, never boil helical membrane protein samples, they will aggregate. > > > bert > > > ------------------------------ > *From:* CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of amit > gaur <cdriamitg...@gmail.com> > *Sent:* 21 February 2017 22:22 > *To:* CCP4BB@JISCMAIL.AC.UK > *Subject:* [ccp4bb] Dimer in SDS-PAGE > > Hi all, > I am trying to purify a potassium ion channel from insect cell using > baculovirus expression system. I am not seeing monomer of this protein in > SDS instead a dimer appears.So,I increased DTT in SDS buffer but no change > and dimer was intact. In size exclusion this protein appeared as a tetramer > which is common oligomerizaton of potassium channel family with GYG motif. > Can any body suggest what should I do in this case? > > Thanks and regards, > > > -- > *Dr. Amit Gaur* > > > *Post Doctoral Researcher PI: Dr. Ji-Fang Zhang Thomas Jefferson > University* > *1020, Locust Street, Suite 418* > *Philadelphia, PA 19107* > > > -- -- *Dr Richard Berry* NHMRC Career Development Fellow Department of Biochemistry and Molecular Biology Monash University Ground Floor, Building 76, ClaytonCampus Blackburn Road Clayton VIC 3800 Australia T: +61 3 9902 9239 E: richard.be...@monash.edu <name.surn...@monash.edu>
