Exploring two variables in vapor diffusion crystallization plate

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 Description of Plate Types VDX • Hampton Research • Hanging Drop • 750 u. L Reservoir Volume Q-Plate • Hampton Research • Sitting Drop • 1 m. L Reservoir Volume Q-Plate II • Hampton Research • Sitting Drop • 750 u. L Reservoir Volume Cryschem • Hampton Research • Sitting Drop • 500 u. L Reservoir Volume Combi. Clover • Emerald Bio. Structures • Sitting Drop • 750 u. L Reservoir Volume Description of Crystallization Scans Precipitants Salts Buffers Hampton Research Crystal Screen I 48 Conditions (used solutions 1 -48) Pegs(P 400, 1500, 4 K, 8 K), isopropanol, MPD, salts (acetates, citrates, formates, phosphates, sulfates, tartrate) acetates, chlorides, citrates, formates, phosphates, sulfates, tartrate, none Imidazole, Na. Acetate, Na. Cacodylate, Na. Citrate, Na. HEPES, Tris, none p. H 4. 6 -8. 5 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, Na. Acetate, Na. Citrate, Tris, none p. H 4. 6 -9. 0 Emerald Biostructures Wizard I 48 Conditions Pegs, organics, nonvolatile organics, salts Na. Cl, Mg. Cl 2, Zn. Acetate, Ca. Acetate, Li 2 SO 4 (all at 0. 2 M), none CHES, HEPES, imidazole, CAPS, citrate, MES, acetate, cacodylate, Na/K phosphate, phosphate-citrate p. H 4. 2 -10. 5 Emerald Biostructures Wizard II 48 Conditions Pegs, organics, nonvolatile organics, salts Na. Cl, Mg. Cl 2, Zn. Acetate, Ca. Acetate, Li 2 SO 4 (all at 0. 2 M), none CHES, HEPES, imidazole, CAPS, citrate, MES, acetate, cacodylate, Na/K phosphate, phosphate-citrate p. H 4. 2 -10. 5 Pharmacia 2 Poly 72 72 Conditions Pegs (200, 400, 1450, 2 K, 3350, 4 K, 6 K, 8 K, 10 K, 20 K) Monomethyl Ether Pegs(550, 750, 2 K, 5 K) NH 4 Phosphate, Na. Citrate, Li 2 SO 4, Na. Br , Ca. Cl 2 , Mg. Cl 2, Na. Cl, KCl, Li. Cl, none Na. Acetate, MES ADA, PIPES, HEPES, EPPS, TAPS, CHES p. H 4. 5 -9. 0 Pegs (P 4 K, P 8 K), Monomethyl Ether Peg 550 Ca. Cl 2, NH 4 Phosphate, NH 4 Sulfate, Na. Citrate, Li 2 SO 4 Mg. Cl 2, Na. Cl, Na. Br, Na. Cl, KCl, Li. Cl Na. Acetate, MES ADA, PIPES, HEPES, EPPS, TAPS, CHES p. H 4. 5 -9. 0 Pharmacia Peg 72 72 Conditions RAMC 2001 – Pharmacia Corporation Company Confidential! 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 Qplate 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