Rhys,
We have solved several structures in our lab using bicelles (all beta-barrels). It is generally the case that we want to minimize excess detergent for crystallization and it is the same for bicelles. At least for our cases, we have not done anything extra to remove excess detergent and so far it seems to work ok for us and we have been able to reproduce the crystals nicely from prep to prep. But, my *guess* is that the detergents probably do have some effect on the bicelles and have the potential to either prevent or even promote crystallization depending on the protein target, etc. To start, I would suggest just doing your prep as normal and then go into bicelles to see if you get any crystals in broad screening. If not, then you could re-address these 'effect of detergent and detergent concentration' issues later. Not sure how familiar you are with bicelles, but I suggest testing your sample buffer prior to using your protein sample, just to make sure the bicelles are stable in your buffer. If the buffer is ok, then i would test a small sample of your protein, say 8 uL of protein + 2 uL bicelles, to make sure the protein sample is stable in the bicelles as well. You don't want to add bicelles to all your protein sample just to find that everything precipitates out. Clear to opaque in color has produced nice crystals for me, but solid white milky color is probably not good. Hope some of this helps, good luck! Cheers, Nick -------------------------------- [ Nicholas Noinaj ] the Buchanan Lab Laboratory of Molecular Biology LMB-NIDDK, NIH 50 South Drive, Room 4505 Bethesda, MD 20892-8030 1-301-594-9230 (lab) 1-859-893-4789 (cell) [email protected] [ the Buchanan Lab ] http://www-mslmb.niddk.nih.gov/buchanan/ -----Original Message----- From: RHYS GRINTER [mailto:[email protected]] Sent: Friday, December 13, 2013 4:54 AM To: [email protected] Subject: [ccp4bb] Bicelle Crystallisation Hi All, I'm thinking of embarking on some crystallisation of a membrane protein in bicelles in the new year. In the methods I've read you simply take your protein in detergent and add the bicelle mxiture (Chapso+DMPC) to your protein allow the solution to equilibrate and set up your screens. What I was wondering is will the original concentration of detergent in your protein sample, effect the likelihood of crystallization (i.e. by changing the structure of the bicelle)? If so do people generally seek to minimize the detergent concentration of the sample before setting up the bicelles. Cheers, Rhys
