Given that your tartrate crystals diffract so much better than your non-tartrate crystals, and tartrate is found in the active site, you might think that tartrate binding in the active site is related to the good diffraction of your crystals. Soaking out a bound tartrate is going to be hard, when you have 1.1 M of it around.
You might like to try a gentle cross linking of the tartrate crystals, then slowly transferring them to another solution (Perhaps malonate? Test to see first if Malonate binds the protein). Once you are out of 1.1 M tartrate you might have a better chance of displacing the tartrate in the active site with your ligand. Janet From: CCP4 bulletin board [mailto:[email protected]] On Behalf Of Nemanja Vuksanovic Sent: Sunday, 22 April 2018 12:39 PM To: [email protected] Subject: [ccp4bb] Removing tartrate ions from the active site Dear All, I've recently crystallized a protein under a condition containing 1.1 M ammonium tartrate. (I was able to get a few other hits but only these crystals diffract below 2.5 A). Upon refinement, it appears that the electron density in the active site shows a mixture of tartrate and the native ligand. This occurs even after overnight soaks with 50 mM of the ligand ( the Km is about 1 mM). Would you recommend dehydrating crystals with PEG when soaking them with ligand or some different method to minimize the presence of tartrate? Thanks in advance, Nemanja -- Graduate Student Department of Chemistry and Biochemistry University of Wisconsin-Milwaukee
