Tiancen, How critical is it that the crystals remain in the sulfate salts? I ask because we recently had success with an RNA crystal that was very intractable to heavy atom soaks because of very high lithium sulfate growth conditions, and the fact that cryo-protection was best done by increasing the lithium sulfate to near saturation. At that high sulfate concentration, many of our favorite heavy atoms (mostly divalent and trivalent cations since the crystals were RNA) are insoluble, and the high Li+ concentration competed off the heavy atoms.
What I did was move the crystals, in steps, from the 2M lithium sulfate into high concentration of lithium Acetate. What this did was increase the solubility of the heavy atom cations, which still maintaining high ionic strength and offering cryoprotection. Ultimately, what worked was 3M lithium acetate + 100 mM cobalt hexammine or iridium hexammine (yes, 100 mM trivalent!). We got a nice derivative, and in fact the diffraction limit of the crystal increased by almost 0.5 Angstroms! This was with an RNA crystal, so your problems may be different, but perhaps something analogous might work, or at least you can get some ideas from our experience. In short, if solubility of the heavy atoms is an issue, you may be able to move the crystals into an alternate high salt condition that will give better solubility, and then crank the heavy atom concentration up to compete with the salt. Good luck, Jeff _______________________________________ Jeffrey S. Kieft, Ph.D. Assistant Professor Dept. of Biochemistry and Molecular Genetics University of Colorado School of Medicine http://www.uchsc.edu/sm/bbgn/kieftj.htm http://www.evolutionarygenomics.com/CERT/CERT.html _____________________________________ For mail: UCHSC at Fitzsimons Mail Stop 8101, PO Box 6511 Aurora, CO 80045 For courier/packages: South Building RC-1, Room 9110 12801 East 17th Ave. Aurora, CO 80010 phone: 303-724-3257 fax: 303-724-3215 email: [EMAIL PROTECTED] "Open your eyes. You have only to see things clearly, to understand." -Leonardo da Vinci -----Original Message----- From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Tiancen Hu Sent: Thursday, April 19, 2007 10:27 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] recommendation for heavy atoms used in lithium sulfate and ammonium sulfate Dear all, Could any one recommend some heavy atoms used for crystals grown in 0.1M tri-sodium citrate pH 5.6, 1M Lithium sulfate and 0.5M ammonium sulfate? I read from Hampton user guide of heavy atom kit that "high salt concentrations are not the ideal medium for heavy atom reactions with macromolecules", so is there any type of heavy atom we should avoid using? We do not have any experience in preparing heavy atom derivative, so any suggestions, experience or references will be greatly appreciated. Thanks in advance! Tiancen Hu Shanghai Institute of Materia Medica
