Dear Roger, We have had success using the non-detergent sulphobetaines (NDSBs) to improve solubility of protein samples. For a couple of projects they have proved crucial for concentrating the protein to a reasonable level for crystallization (e.g. PMID: 18765907). Be careful since there is a big variation in price dependent on which particular one you use. Use them at ~200-300 mM and you may even be able to get rid of the glycerol. You may have to incubate the protein with the NDSBs overnight before concentration to get the full effect.
Good luck Tom Walter ---- Original message ---- >Date: Wed, 25 Aug 2010 10:19:13 -0400 >From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> (on behalf of Roger Rowlett ><rrowl...@colgate.edu>) >Subject: [ccp4bb] Crystallization of low solubility proteins from >glycerol-containing solutions >To: CCP4BB@JISCMAIL.AC.UK > > Does anyone have practical experience crystallizing low > solubility proteins from solutions containing significant > (10-20%) glycerol? We can get small crystals by mixing 4:1 > ratios of protein to well solution, but the drops do not > concentrate back to the well solution volume as anticipated, > even if the well solution is brought to 10-20% glycerol as > well to balance osmolarity. Concentration of the protein > further to reduce the protein:well solution ratio may not be > practical (it crashes out) even in 20% glycerol. > Unfortunately, glycerol seems to be required to maintain > protein solubility, so that may not be practical to remove > either. > > One thought is to add additional osmolyte to the well solution > to draw down the drop volume once small crystals form, a kind > of a "macro-seeding" approach, but I am not aware of a > systematic way of doing this. Anyway, I am almost certain I am > trying to re-invent the wheel,as someone has probably done > something similar. Any suggestions would be appreciated. > > Cheers, > -- > > ---------------------------------------------------------- > > 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
