Dear Raji
Running a blue-native gel with lanes in the presence and absence of a reducing agent could prove quite informative. DLS could also return a quick result on the particle distribution in your sample. In that case I would measure samples as fractionated from the superdex200 and compare the measurements after centrifuging the same samples at 100k x g for one hour.
Best regards
Savvas
On 22 Feb 2012, at 00:21, Raji Edayathumangalam <[log in to unmask]> 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
>
>
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