sometimes change in ph also dissocaites the complex,so you need to screen best buffer ph for your protein complex.
atul kumar On Sun, May 23, 2010 at 8:18 PM, Maia Cherney <ch...@ualberta.ca> wrote: > -------- Original Message -------- > Subject: Is it possible for the Tris buffer to strip the Zn ions > from the Zinc Finger motif of a protein? > Date: Sun, 23 May 2010 08:45:55 -0600 > From: Maia Cherney <ch...@ualberta.ca> > To: ruheng <rh_ibp2...@hotmail.com> > > > > The complex can dissociate without loosing Zn. For example, if you dilute > it too much. There is an equilibrium between the concentrations of complex > and the individual components. > > Maia > > > > Date: Sat, 22 May 2010 11:17:41 +0800 > From: rh_ibp2...@hotmail.com > Subject: [ccp4bb] > To: CCP4BB@JISCMAIL.AC.UK > > Dear all, > > Recently, I am working on a complex which includes two protein subunits. > The interaction was based on the Zinc Finger motif of one protein. I > co-purified the complex by nickel affinity column with one protein bearing a > C terminal His tag and the other without any affinity tags. However, the > complex was disassociated when applied to size exclusion chromatography. The > buffer I use for SEC is 20mM Tris-HCl, 150mM NaCl, 1mM DTT, 5% Glycerol, pH > 7.5, whearas the buffer I use for nickel affinity column is 50mM Na2HPO4, > 10mM KH2PO4, 137mM NaCl, 2.7mM KCl, 10% Glycerol, pH7.4. So I am wondering > is it possible for the Tris buffer to strip the Zn ions from the Zinc Finger > motif of one protein that leads to the destruction of the complex? > > I will be very appreciated if anyone has some experience in such case and > would like to share with me! > > > Sincerely, > > Heng > > > Institute of Biophysics, > Chinese Academy of Sciences, > Beijing 100101, China > > ------------------------------------------------------------------------ > 搜索本应是彩色的,快来体验新一代搜索引擎-必应,精美图片每天换哦! 立即试 用！ <http://cn.bing.com/?form=CRMADS> > ------------------------------------------------------------------------ > 聊天+搜索+邮箱 想要轻松出游,手机MSN帮你搞定! 立刻下载！ <http://3g.msn.cn/> >
