We had this happen with RPA14/32 heterodimer, we kept the two peaks separate, and then grew crystals from each but in different space groups and diffraction resolution. See Habel, J. E., Ohren, J. F. and Borgstahl, G. E. O. "Dynamic light scattering analysis of full-length, human RPA14/32 dimer: purification crystallization and self-association" *Acta Cryst.*D57, 254-259 (2001).
On Sun, Dec 15, 2013 at 2:33 PM, Phoebe A. Rice <pr...@uchicago.edu> wrote: > It might be that the protein is innocent and the FPLC is guilty. If > FPLC pump needs maintenance and is stuttering a bit when running at high > pressure, the salt gradient won't be as smooth as you'd like. If you have > an ionic strength detector, that trace will tell you. Otherwise, see if > other people's proteins are also looking a bit iffy under similar > circumstances. > Phoebe > ------------------------------ > *From:* CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of > Katherine Sippel [katherine.sip...@gmail.com] > *Sent:* Sunday, December 15, 2013 11:53 AM > *To:* CCP4BB@JISCMAIL.AC.UK > *Subject:* Re: [ccp4bb] A question on protein microheterogenity for > crystalization > > Hi Acoot, > > There are a lot of great suggestions here already. I've also run into this > phenomenon in couple of cases. The first was a binding protein in mixed > bound and unbound forms. The second was a case of heterogeneous > post-translational modification. In both cases I could only get crystals > from purified peaks and not pooled protein. If protein is precious I'd > second Mark's suggestion to try screening pooled protein while you scale up > your prep. > > Cheers, > Katherine > > > On Sat, Dec 14, 2013 at 6:16 AM, Acoot Brett <acootbr...@yahoo.com> wrote: > >> Dear All, >> >> When I purified my protein by ion exchange chromatography for >> crystallization, there were several peaks containing the target protein as >> analyzed by SDS-PAGE. All these peaks have the same MW as determined by gel >> filtration coupled MALLS. >> >> For crystallization purpose, can I merge the corresponding ion exchange >> chromatography peaks together? Otherwise the protein yield will be too low. >> And how to explain the heterogeneity by ion exchange chromatography in this >> situation? >> >> I am looking forward to getting a reply from you. >> >> Acoot >> > > > > -- > "Nil illegitimo carborundum"* - *Didactylos >
