Is it possible that you have a case of domain swapping that causes the trimeric assembly?
Diana ************************************************** Diana R. Tomchick Professor Departments of Biophysics and Biochemistry University of Texas Southwestern Medical Center 5323 Harry Hines Blvd. Rm. ND10.214A Dallas, TX 75390-8816 diana.tomch...@utsouthwestern.edu<mailto:diana.tomch...@utsouthwestern.edu> (214) 645-6383 (phone) (214) 645-6353 (fax) On Mar 19, 2018, at 3:07 PM, PULSARSTRIAN <bhanu.hydpri...@gmail.com<mailto:bhanu.hydpri...@gmail.com>> wrote: Dear all, Sorry for the slightly off-topic question. I am working on a non-native, de novo [4Fe-4S] protein, designed as a four-helix bundle. The in vitro reconstituted protein assembles with [4Fe-4S] (confirmed by EPR) and exists in monomer-dimer configuration (confirmed by SEC). These results have been already published. Recently we could get the [4Fe-4S] assembly directly from the E. coli (in vivo assembly). Everything is as expected (compared to reconstituted protein), except the oligomerization state. The protein assembles as trimer, in contrast to monomer-dimer configuration of the reconstituted protein. The trimeric nature of the in vivo assembled protein has been confirmed by SEC, SEC-SLS and SAXS. So, my question is, has anyone encountered such situation, where the As-purified Fe-S protein having a completely different oligomeric state compared to the in vitro reconstitution protein? Looking forward to hearing for some examples and/or references. Regards, Bhanu ________________________________ UT Southwestern Medical Center The future of medicine, today.
