“Localization of small-ligand binding to pharmacological targets using cryo-electron microscopy”

Friday August 28, 2015

“Localization of small-ligand binding to pharmacological targets using cryo-electron microscopy”

NIH National Cancer Institute, Mario Borgnia

Advances in cryo-electron microscopy are driving the determination of small protein complexes at near-atomic resolution with unprecedented rapidity. For example, computer driven image acquisition together with streamlined image processing pipelines offer the opportunity to localize the binding of ligands with low molecular weight to their target proteins in a relatively automated way. Moreover, improvements in classification algorithms enable the reconstruction of sub-nanometer resolution density maps of distinct conformations from a mixture in single droplet of protein solution.

To test the practicality of this approach, we combined automated procedures and advanced with direct electron detectors to localize ADP (427 Da) and other small ligands on glutamate dehydrogenase (GDH), a clinically significant 365 kDa enzyme that is a relevant pharmaceutical target for cancer, Parkinson’s, and diabetes.

Images from single specimens collected in a single session provided enough information to localize nucleotides in a complex at ~3.5 Å resolution. The validity of the results was confirmed by comparison with the crystallographic coordinates of GDH in complex with a variety of ligands. The methods we present provide a streamlined path to rapidly solve the structure of macromolecular complexes and to image the binding target of drug molecules at near atomic resolution.