Drug Discovery

Figure 8

Atomic-resolution model derived from cryo-EM

structure in the presence of

bound inhibitor. Left: Cryo-EM structure of the C5-Cobra

Venom Factor (CVF) complex at 3.35 Å resolution.

Complement C5 alpha chain in green, beta chain in orange, cobra venom factor in grey, inhibitor in blue. Right:

Electron microscopy density map (blue mesh) for bound inhibitor with binding site residues shown as green

sticks. Images of PDB-entry 6i2x created with Pymol by Hans Raaijmakers

team was able to ascertain the precise interaction between insulin and the insulin receptor, opening up exciting new avenues in drug discovery (Figure 5)6.

Viruses and antibodies Cryo-EM reveals structure of viral epitope, enabling development of new antibody screen against Polyomaviruses Human polyomaviruses (BK and JC) establish per- sistent infection in kidneys during childhood with minimal clinical manifestations. The virus reacti- vates under immunodeficient conditions, causing nephropathy and haemorrhagic cystitis. Novartis developed a new high-throughput,

functional antibody screen to examine the response to polyomavirus with isolated monoclonal anti- bodies that neutralise BK and JC virus subtypes. The objective of this research was to identify a

complex binding site of the virus capsid protein, which was not possible with crystallography due to the complex’s quaternary structure. Cryo-EM reveals the quaternary nature of viral

epitope and unravels potent modality for inhibiting polyomavirus infection in kidney transplant recip- ients and other immunocompromised patients (Figure 6)7.

Small molecules How cryo-EM illuminates stalling ribosome by small molecules and opens new therapeutic strategy? One of the new strategies for therapeutic develop- ment is using small molecules that inhibit protein synthesis by selectively stalling the ribosome. Structures of human Ribosome Nascent Chain


(RNC) complexes, stalled by drug-like molecule PF846, selectively blocks the translation of a small number of proteins by an unknown mechanism. The objective in this research by Pfizer was to

determine the structures of RNC complexes which could not be solved by X-ray crystallography due to their size, complexity and conformational flexi- bility, establishing structural foundation for devel- oping therapeutic small molecules inhibitors. “The requirement of X-ray crystallography is

that you need to obtain a diffracting crystal to get atomic resolution. That has been a big hurdle if you’re working on challenging targets. Almost every target we’re working on now is very chal- lenging,” said Dr Seungil Han, Associate Research Fellow at Pfizer. High-resolution cryo-EM structures reveal how

small molecule PF846 selectively stalls the transla- tion of the human ribosome by binding in the ribo- some exit tunnel and altering the path of the nascent polypeptide chain. “The cryo-EM method is well-suited for provid-

ing critical structural information to influence the discovery of biologics, vaccines and gene therapies. However, the most surprising aspect has been our ability to provide binding site and binding pose information for small molecules on large proteins or protein complexes,” said Dr Han (Figure 7)8.

Cryo-EM-enabled development of new small- molecule inhibitor for therapy against Paroxysmal Nocturnal Hemoglobinuria (PNH) Paroxysmal Nocturnal Hemoglobinuria (PNH) is a rare, acquired and life-threatening disease leading

Drug Discovery World Winter 2019/20

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