Hi Sajid, with my very (!) limited experience I can recommend trying to co-crystallise with your cryo-buffer at very low concentration. It helped me (once!) to get diffraction going from about 2.7 to 1.5 ... oh, and it got rid of some nasty twinning problem as well. I guess the thinking is that if some cryo-buffer molecule goes into crystal contacts it will destroy (to some extent) crystals that haven't seen the buffer molecules before (and increase mosaicity etc). But if they have seen the cryo molecule the crystal contacts are already satisfied and adding more (to avoid ice) doesn't destroy the contacts.
At least that is my simplistic view ... Unfortunately I missed some of the tutorials/talks Elspeth Garman gave at this years RapiData workshop: but I gather there is a huge potential of improving freezing (or is it cryo-cooling?). Just look at some of Elspeth's papers and other papers that cite them. Cheers Clemens On Tue, Jun 03, 2008 at 01:45:42PM -0400, Christian Biertuempfel wrote: > Hi Sajid, > Just a simple test for your problem: Incubate your crystal longer in > your cryo/stabilization solution. This helps sometimes to lower > mosaicity. Of course, you can also try co-crystallization with glycerol > (2%, 5% or 10%). > > Good Luck, > christian > > > sajid akthar wrote: > >Dear All > > > >My protein size is ~30kD and crystallizes with > >19%Peg3350, 0.2M Nacl, and 0.1M Na Cacodylate buffer. > > > >Please refer the attached crystal image with this. The > >crystal looks like good enough for home source. These > >crystals appears in 4-5 days at room temp. > > > >Sometimes I'm getting crystals like this, but very few > >in 24 well tray. Most of the time, I found the drop > >contains needles. If I reduce the precipitant little > >bit, I wont find any change in the drop even after > >long time. Changing pH (or temp)of the buffer does not > >help me any better. The crystal appears only around > >5.5pH. > > > >The problem is mosaicity. This crystal diffracted in > >home source upto 3.2A and the mosaicity is 2.5degree. > >Almost all the good crystal like this having same > >mosaicity. > > > >Good cryo condition so far that I found was > >10%Glycerol with mother liquor. Other conditions > >weekens the diffraction quality or increase mosaicity. > > > >In many crystal I could see some crack in the middle > >of the crystal, it looks like twin crystal. Or the > >crystal appears with some sattelite crystals. > > > >Can anyone suggest me some good way to overcome these > >problems. > > > >Thankz > > > >Sajid > > > > > > > > From Chandigarh to Chennai - find friends all over India. Go to > > http://in.promos.yahoo.com/groups/citygroups/ > > > > > >------------------------------------------------------------------------ > > > > > _______________________________________________________________________ > > Dr. Christian Biertümpfel > Laboratory of Molecular Biology > > NIDDK/National Institutes of Health phone: +1 301 402 4647 > 9000 Rockville Pike, Bldg. 5, Rm. B1-03 fax: +1 301 496 0201 > Bethesda, MD 20892-0580 > USA > _______________________________________________________________________ > -- *************************************************************** * Clemens Vonrhein, Ph.D. vonrhein AT GlobalPhasing DOT com * * Global Phasing Ltd. * Sheraton House, Castle Park * Cambridge CB3 0AX, UK *-------------------------------------------------------------- * BUSTER Development Group (http://www.globalphasing.com) ***************************************************************
