Use of High Throughput Screening to Determine Lead Crystallization Conditions and as a Tool for Basic Research
Sample Database BIG BROTHER Reader Tables RAID1 Failover fileserver 2 TB RAID5 snapshots HT Infrastructure:Computer Hardware and Software BACKUP Tape Optical Disc Offsite Data to Primary fileserver 1 TB RAID5 subsystem
New Old How can we improve our output? Improved image quality = Improved lead detection
Images courtesy of Greiner Bio-One PS Glass LBR Outcome Verification Identify the outcome as crystalline Identify the composition
Co-Crystallants • The SGPP co-crystallants were designed to promote 2 types of macromolecular interactions: • intra-molecular interactions (freezers) and • inter-molecular interactions (glues) • The library uses “privileged structural motifs”, fused-ring aromatics (triazoliums or guanidiniums) “decorated with bromines”*. *Hovey, B., Verlinde, C. L. M. J., Merritt, E. A. & Hol, W. G. J. (1999). Structure-based discovery of a pore-binding ligand: Towards assembly inhibitors for cholera and related AB5 toxins. J. Mol. Biol. 285, 1169-1178.
01 04 02 10 09 05 11 12 13 Co-Crystallants Used in This Study
Co-Crystallant Experiments • Crystallization in HTS laboratory • Compared the crystallization behavior of: • native protein • protein-DMSO • protein-DMSO-10mM Co-Crystallant • DLS • Look for evidence of aggregation in the presence of the co-crystallants.
DLS Results • Sample aggregation occurred in the presence of the co-crystallants.
Acknowledgements • Robert J. Collins, Nancy A. Fehrman, Stacey M. Gulde, Angela M. Lauricella, Carrie A. Mancuso, Christina K. Veatch and George T. DeTitta • Jizhen Li, Brian Carroll, Erkang Fan, Mark Robien and Wim G.J. Hol
Acknowledgements • Work supported in part by the John R. Oishei Foundation, the Cummings Foundation, the Western New York Foundation, NASA NAG8-1594, NASA NAG8-1839, NASA NCC8-232, NIH RR016924 NIH P50 GM-62413 and NIH P50 GM-64655. • Special thanks to Greiner Bio-One