RAMC 2001 – Pharmacia Corporation Company Confidential! Exploring two variables in vapor diffusion crystallization: plate types and crystallization scans Jennifer Pawlitz, Stephanie Shieh, Jennifer Sharamitaro, Eric Sturman, Mairi Lough, Anna Stevens Discovery Research Pharmacia Corporation Chesterfield Village, St. Louis, Missouri VDX Hampton Research Hanging Drop 750uL Reservoir Volume Q-Plate Hampton Research Sitting Drop 1mL Reservoir Volume Q-Plate II Hampton Research Sitting Drop 750uL Reservoir Volume Cryschem Hampton Research Sitting Drop 500uL Reservoir Volume CombiClover Emerald BioStructures Sitting Drop 750uL Reservoir Volume Description of Plate Types PrecipitantsSaltsBuffers Hampton Research Crystal Screen I 48 Conditions (used solutions 1-48) Pegs(P400, 1500, 4K, 8K), isopropanol, MPD, salts (acetates, citrates, formates, phosphates, sulfates, tartrate) acetates, chlorides, citrates, formates, phosphates, sulfates, tartrate, none Imidazole, NaAcetate, NaCacodylate, NaCitrate, NaHEPES, Tris, none pH Hampton Research Crystal Screen II 48 Conditions (used solutions 1-48) Pegs, Monomethyl Ether Pegs, Jeffamine, organics, non-volatile organics, salts chlorides, phosphates, sulfates, tartrate, none Bicine, HEPES, MES, NaAcetate, NaCitrate, Tris, none pH Emerald Biostructures Wizard I 48 Conditions Pegs, organics, non-volatile organics, salts NaCl, MgCl 2, ZnAcetate, CaAcetate, Li 2 SO 4 (all at 0.2M), none CHES, HEPES, imidazole, CAPS, citrate, MES, acetate, cacodylate, Na/K phosphate, phosphate-citrate pH Emerald Biostructures Wizard II 48 Conditions Pegs, organics, non-volatile organics, salts NaCl, MgCl 2, ZnAcetate, CaAcetate, Li 2 SO 4 (all at 0.2M), none CHES, HEPES, imidazole, CAPS, citrate, MES, acetate, cacodylate, Na/K phosphate, phosphate-citrate pH Pharmacia 2Poly72 72 Conditions Pegs (200, 400, 1450, 2K, 3K, 3350, 4K, 6K, 8K, 10K, 20K) Monomethyl Ether Pegs(550, 750, 2K, 5K) NH 4 Phosphate, NaCitrate, Li 2 SO 4, NaBr, CaCl 2, MgCl 2, NaCl, KCl, LiCl, none NaAcetate, MES ADA, PIPES, HEPES, EPPS, TAPS, CHES pH Pharmacia Peg72 72 Conditions Pegs (P4K, P8K), Monomethyl Ether Peg 550 CaCl 2, NH 4 Phosphate, NH 4 Sulfate, NaCitrate, Li 2 SO 4 MgCl 2, NaCl, NaBr, NaCl, KCl, LiCl NaAcetate, MES ADA, PIPES, HEPES, EPPS, TAPS, CHES pH Description of Crystallization Scans Conclusions Plate Type Experiment –Combiclover plate gave the most crystal hits over all proteins –14 conditions gave hits with the Combiclover plate and no others –3 conditions gave hits with Q-plate or Q- plate II and no others –2 conditions gave hits with all trays except VDX and Combiclover –For only 5 conditions (across all proteins) did all 5 trays give hits Crystallization Scan Experiment –Hampton Research Crystal Screen I had the best overall crystallization rate of success –Overall, there was a ~5-15% chance of crystallizing a protein from any one of the scans –Only 11 conditions (from a total of 336) yielded crystals of 2 or more of the 4 proteins tested