 |
 |
To add to what Pat has mentioned, I work with a protein that has a number of exposed Cys residues, and it turns into a gel at 4 degrees within a week, even if I store it in buffer with reducing agents. This happens every time I store it at 4 degrees. To test if your "jelly" is due to S-S crosslinking, add a really huge excess of DTT or beta-mercaptoethanol to a small sample and see if it goes back into solution. I usually add 100mM DTT for this test. If it does turn out to be disulfide-related, you can try to add reducing agents with longer half-lives (ie TCEP instead of DTT), but unfortunately you may just have to work with the protein within a couple days of finishing the prep. Mike ----- Original Message ----- From: "Patrick Loll" <[log in to unmask]> To: [log in to unmask] Sent: Tuesday, August 30, 2011 9:06:52 AM GMT -08:00 US/Canada Pacific Subject: Re: [ccp4bb] Protein preps become a jelly Certainly not unprecedented, or even that unusual (I remember making gels from BSA and IgG solutions during grad school rotations). Gel formation usually requires crosslinking, so consider whether you might be getting adventitious disulfide bond formation. Pat On 30 Aug 2011, at 11:31 AM, aidong wrote: > Dear Buddies, > > Sorry for bothering you with an off-ccp4 question. We recently are experiencing a very strange phenomena. A couple of protein preps with reasonably high concentration (10-20mg/ml) become a jelly after storages for overnight or a couple of days at 4C. All of them have been purified by gel filtration. Some of these proteins behave like this from very first preps but some of them had been very kind to us previously. We have googled extensively in CCP4BB and www but it appears this only happens to us. It would be highly appreciated that you could exchange their experiences or provide your suggestions. > > Aidong Han, Ph.D > > Department of Biomedical Sciences > School of Life Sciences > Xiamen University > Xiamen, Fujian 361005 > China > Phone: 0592-218-8172 (O) > 0592-218-8173 (L) > Web: http://life.xmu.edu.cn/adhanlab/ -- Michael C. Thompson Graduate Student Biochemistry & Molecular Biology Division Department of Chemistry & Biochemistry University of California, Los Angeles [log in to unmask]