Breaking Cryo-EM Resolution Barriers to Facilitate Drug Discovery
Using cryo-EM we were able to capture images of glutamate dehydrogenase (GDH), an enzyme found in cells, at a resolution of 1.8 angstroms, a level of detail at which the structure of the central parts of the enzyme could be visualized in atomic detail. We also imaged two small proteins in this study, isocitrate dehydrogenase (IDH1) and lactate dehydrogenase (LDH), that are active targets for cancer drug development.
– Near-atomic-resolution structure of isocitrate dehydrogenase, a < 100 kDa enzyme.
– 1.8 Å resolution cryo-EM map of 334 kDa glutamate dehydrogenase.
– 2.8 Å resolution cryo-EM map of lactate dehydrogenase, a 145 kDa protein.
– Localization of small-molecule inhibitors bound to metabolic protein complexes.
Recent advances in single-particle cryoelecton microscopy (cryo-EM) are enabling generation of numerous near-atomic resolution structures for well-ordered protein complexes with sizes ≥ ∼200 kDa. Whether cryo-EM methods are equally useful for high-resolution structural analysis of smaller, dynamic protein complexes such as those involved in cellular metabolism remains an important question. Here, we present 3.8 Å resolution cryo-EM structures of the cancer target isocitrate dehydrogenase (93 kDa) and identify the nature of conformational changes induced by binding of the allosteric small-molecule inhibitor ML309. We also report 2.8-Å- and 1.8-Å-resolution structures of lactate dehydrogenase (145 kDa) and glutamate dehydrogenase (334 kDa), respectively. With these results, two perceived barriers in single-particle cryo-EM are overcome: (1) crossing 2 Å resolution and (2) obtaining structures of proteins with sizes < 100 kDa, demonstrating that cryo-EM can be used to investigate a broad spectrum of drug-target interactions and dynamic conformational states.