Hi Frank, The previous suggestions are great. In my case, I had soft crystals (bend from 180° to ~130-140°) in the loop using the LCP but they still diffracted. There are a lot of "heroic stories" on how some people solved a structure, you should just try as many ways as possible (do not only rely for a long time on only one condition or construct), and remember "methods" are just methods (i.e., there are hundreds of LCP robots in the world..). Protein engineering, homologs (maybe hyperthermophiles).. stabilization.. are great ways. Recently I had tested various crystals grown by vapor diffusion for a protein purified in different ways/detergents, so obtaining crystals is very common, but it's just the start. I also had crystals using bicelles but saw huge spots to 2 A (no patterns.. rather looked like detergent/lipid). Good luck
toufic el arnaout ________________________________________ From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of crystalboy [yyb...@gmail.com] Sent: Wednesday, October 23, 2013 11:22 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] membrane protein optimization Hi CCP4BB Forks, In recently I got a membrane protein crystal in the quite normal membrane protein crystallization conditions as other persons reported, like PEG400 16-19%, pH 6.0-7.5, with 50 mM MgCl2 (in my case) by using sitting drop method. These crystals are around 50-100 uM. They look like trapezoid crystal. My problem is all of my crystals have not diffraction in home source X-ray and just poor diffraction at Synchrotron (lower than 20 A). My crystals like to appear on the surface of the drop. Look like my crystals are quite light. I had tried to use a needle to touch them. Unlike other protein crystal, my crystal looks like quite "soft". When I touch it, it didn't crack, but was bend or mashed. I had tried to do additive screen and detergent screen. It seems they are not useful. Do anyone have good ideas to optimization these crystals? Thanks for your suggestions. Frank
