Sure it's not the peptide that crystallizes ? You could try adding glycerol, exchanging the PEG, lowering the PEG playing with the pH or changing to a different buffer system. You might have done all these already, then you could also try a larger drop size. You could also mix your protein with your reservoir and spin it down, then setup the tray but not adding additional reservoir to it. How about temperature ? Is your protein more soluble at 4˚C or 37˚C ?
Just some thought before the sun rises, Jürgen - Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry & Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-3655 http://web.mac.com/bosch_lab/ On Oct 10, 2010, at 2:15 AM, Dilip Badgujar wrote: > hi guys > > I am trying to crystallize protein complexed with peptide(14mer). > In initial screening i got microcrystals in following condition > - 0.2M MgCL2 > 0.1M Tris pH 8.5 > 28 % PEG 4000 > Ptotein concentration - 12.5mg/ml and peptide conc.-5mg/ml > Complex ratio- (Protein-Peptide)(1:1.5) > Incubation temp.-22 degree > Method - Sitting drop vapor diffusion method > > within four hours of setting trials I can see too many microcrystals. > Though i tried reduction of protein and precipitant conc. as well as using > mineral oil to decrease evaporation rate but still it is not working. > > I am waiting for your valuable suggestions. >
