It is in fact possible that different solution conditions favor different oligomeric assemblies. For example, perhaps your protein, which in one set of solution conditions prefers the monomer, prefers some other assembly under the crystallization conditions (we've seen this in our own work). Perhaps the higher (or lower) salt concentration, the presence (or absence) of the precipitation reagent, etc. pushes any oligomer equilibrium that exists for your protein.
- this all is very true; in addition, oligomeric state does depend on protein concentration, too. jsPISA at CCP4 (http://www.ccp4.ac.uk/pisa) calculates concentration profiles for probable macromolecular assemblies, which may be useful to check with. The effect of salt/precipitators/pH is not there, though. Eugene Regardless of what our final conclusion would be for this case, we became rather generally interested to find other similar cases of homomeric assemblies related only by non crystallographic translation symmetry (or as Engin Qzka pointed out "improper NCS" is the conventional terminology). So to rephrase our question we are interested to learn about additional structures of homomoeric improper ncs assemblies. An interesting case of heterologous interfaces engaged is that of the VP40 protein. A recent publication from the Saphire group (PMID 23953110<http://www.ncbi.nlm.nih.gov/pubmed/23953110>) may present an instructive situation, though I don't know the details of the NCS observed. Here an octameric assembly (PDB 4LDM, ASU=monomer) and a hexameric assembly (PDB 4LDD, ASU=dimer) of the same protein engage different interfaces on the protomers. In forming the oligomer, the protomers are aligned with alternating interfaces. Emily. I truly appreciate ANY open-minded or skeptic thought, profound or trivial that we get here! They all, definitely those made by Mark Garavito, contribute to shaping our mind around this riddle. Thanks for commenting on the skepticism, I brought it up as part of the discussion but a glitch of my own coffee time haziness might have slipped in. Perhaps I should try some o-cha instead .. :) cheers, Hay On Dec 12, 2014, at 3:05 AM, Jeremy Tame wrote: Dear Hay I suggest that you use analytical ultracentrifugation to determine the oligomeric state of the protein in solution. Mass spectrometry and light scattering are also useful, but there are so many examples of gel filtration proving erroneous it has questionable value as an analytical technique. For an example of a dimer interface predicted by PISA to be real you could look at Yoshida et al, JMB 423, 351 (2012). The protein is in fact a monomer in solution. PISA is a fantastic tool, but interfaces in crystals do not always reflect the solution state. My guess (with the information I have) is that your protein is probably a monomer too. With regard to Michael Garavito's reply requesting more information, I would like to comment that scepticism is indeed an important god in the pantheon of science, but that that minor deity open-mindedness also deserves the occasional nod. 10-fold crystal symmetry is one example, but the list of "impossible" things now become mainstream is a long one (continental drift, Earth >100,000 years old, quantum mechanics....and so on). Bayes theorem cannot help you discover the truth if you have set its prior probability to zero. But I haven't my morning o-cha yet either. good luck Jeremy On Dec 11, 2014, at 9:27 PM, Hay Dvir wrote: Dear all, We have a structure of a rather tightly packed homotrimer protein in the ASU with no apparent crystallographic or non-crystallographic rotational symmetry between monomers. Attempting to establish the biological assembly, we are very interested to hear about additional similar cases you might know of. Thanks in advance, Hay --------------------------- Hay Dvir Ph. D. Head Technion Center for Structural Biology Technion Haifa 3200003, Israel Tel: +(972)-77-887-1901<tel:%2B%28972%29-77-887-1901> Fax: +(972)-77-887-1935<tel:%2B%28972%29-77-887-1935> E-mail [email protected]<mailto:[email protected]> Website http://tcsb.technion.ac.il<http://tcsb.technion.ac.il/>
