Screening for ligands using a generic and high-throughput light-scattering-based assay.
Rapid identification of small molecules that interact with protein targets using a generic screening method greatly facilitates the development of therapeutic agents. The authors describe a novel method for performing homogeneous biophysical assays in a high-throughput format. The use of light scattering as a method to evaluate protein stability during thermal denaturation in a 384-well format yields a robust assay with a low frequency of false positives. This novel method leads to the identification of interacting small molecules without the addition of extraneous fluorescent probes. The analysis and interpretation of data is rapid, with sensitivity for protein stability comparable to differential scanning calorimetry. The authors propose potential uses in drug discovery, structural genomics, and functional genomics as a method to evaluate small-molecule interactions, identify natural cofactors that stabilize target proteins, and identify natural substrates and products for previously uncharacterized protein targets.
Senisterra
Guillermo A
GA
Structural Genomics Consortium, University of Toronto, Toronto, Ontario, Canada. senisterra@utoronto.ca
Markin
Eugene
E
Yamazaki
Ken
K
Hui
Raymond
R
Vedadi
Masoud
M
Awrey
Donald E
DE
D001665:Binding Sites; D002152:Calorimetry, Differential Scanning / Q000379:methods*; D002950:Citrate (si)-Synthase / Q000378:metabolism; D020650:Combinatorial Chemistry Techniques; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D007202:Indicators and Reagents; D008024:Ligands; D062907:Oxaloacetic Acid / Q000378:metabolism; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D013696:Temperature
D007202:Indicators and Reagents; D008024:Ligands; D011506:Proteins; D062907:Oxaloacetic Acid; D002950:Citrate (si)-Synthase