Well, depends on what 'aggregated' really means. If it implies reasonably weak oligomerization interaction - and it might not be too strong given that the oligomers remain soluble - a chaotropic crystallization agent (on the extreme end certain high salts, consult Hofmeister for chaotropicity) may rip such soluble aggregates apart or at least get them into a conformationally reasonably well defined state. Crystals do appear/transform even from precipitates on occasion. CD will tell you about the (secondary structure) folding state, not the aggregation state, DLS/MALS would give an estimate for and distribution of the aggregation state. With light scattering, you can also do some systematic experiments exploring what might reduce the aggregate size.
I think soluble, defined secondary structure, and a lot of it, is already a good sign. BR On Tue, Feb 21, 2012 at 5:21 PM, Raji Edayathumangalam <r...@brandeis.edu> wrote: > Hi Folks, > > As crazy as it sounds, if you have crystallized and managed to solve > the structure of a protein from aggregated protein, please could you > share your experience. > > After many constructs, many many expression schemes and after the > usual rigmarole of optimization that is also often discussed on ccp4bb > (buffers, glycerol, salt concentrations, pH, detergent, additives > etc.), I now have a decently expressing truncated construct for my > protein (80 kDa) that is pure but aggregated (elutes in the void > volume from a Superdex200 column). I am tempted to make a boatload of > aggregated protein and set up some crystal trays (after perhaps > testing by CD). So I'd like to hear from folks who have been > successful in solving structures from aggregates when many many known and tested optimization methods still leave one with aggregated protein. > > Thanks. > Raji > > -- > Raji Edayathumangalam > Instructor in Neurology, Harvard Medical School Research Associate, > Brigham and Women's Hospital Visiting Research Scholar, Brandeis > University > > -- ******************************************* Jacob Pearson Keller Northwestern University Medical Scientist Training Program email: j-kell...@northwestern.edu *******************************************
