I would try the following approach, some of
which you may have done already:
- Broad pH screen, by 0.5 pH units to see if you are in the pH
sweet spot.
- Finer PEG-4000 screen at the pH identified in the pH
screen--the precipitate in the well would alert me to consider
that either PEG-4000 is too high, or protein is insufficiently
pure
- Try some of your best conditions at 4 deg C instead of RT, or
some other intermediate temperature if you have the capability.
(For me, the 4 deg C often makes a big difference; I've had
minimal luck with intermediate temps.)
- Try an additive screen with 5-10% PEGS, polyols, and liquids
(don't forget DMSO, DMF, MPD, as well as EG, glycerol or
glucose, etc.), or 0.1-0.2 M salts. Whatever you have on hand,
or more thorough march through the Hampton Research additives.
If anything improves crystal form, try more (or less), or
combine additives.
- At some point, try "reversing" your protein/precipitant
ratios. You will typically have more luck in the higher
protein/lower precipitant part of the crystallization phase
diagram than the reverse. Screens are usually at the higher
precipitant/lower protein part of the crystallization phase
diagram.
Cheers.
On 8/25/2010 8:37 AM, rui wrote:
Hi, All,
I'm trying to crystallize a soluble protein and got something like
granular, they are rounded shaped and not so regular and also
don't have sharp edges. See the attached pic. The current
condition is PEG4000 and pH around 5. How can I improve this
condition? Thanks a lot.
--
Roger S. Rowlett
Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@colgate.edu
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