Dear Dhaval, To check whether your compound precipitates, you could set up crystallization drops with everything (buffer, precipitant, compound), but without the protein. If the precipitates still form, it’s your compound. Remember, getting cocrystals with your ligand of interest might be as difficult as getting the protein crystals themselves! -Did you try soaking your compound into apo-crystals? It is not the best method, but if it works it is fast and you do not have to find (de novo) cocrystallization conditions. -Many crystals tolerate 10-15% DMSO. If your compound precipitates, you could try adding some DMSO or ethanol. The latter may be more friendly for your protein. -You could try to prepare a diluted complex and concentrate the complex. -You may have to try a de novo crystallization screen, since your complex crystallizes under completely different conditions as the apo-protein -By searching the internet, you can find many more tips.
Best, Herman Von: CCP4 bulletin board [mailto:[email protected]] Im Auftrag von dhaval patel Gesendet: Samstag, 17. Dezember 2016 09:01 An: [email protected] Betreff: [ccp4bb] Need suggestion Dear CCP4 users, I am trying to co-crystal my protein of interest with a ligand and within 2days of crystallization setup, I guess there is a formation of aggregrates or something like that (see attachment picture). My experiment details are as follow: Protein - 40kDa LIgand- 1728 Da Buffer condition - 25% PEG 1500 and its variation with no salts Protein used for crystallization - 5mg/ml (130uM) in 20mM Tris pH 7.5 Ligand used for crystallization - 250 uM, 500 uM and 1mM dissolved in water. Crystallization method - microbatch under oil I am observing the same kind of pattern in almost all wells in the plates. Any help is appreciated -- Dhaval Patel PhD Student, Bioinformatics & Structural Biology Indian Institute of Advanced Research +91-9925450504
