High-resolution cryo-EM proteasome structures in drug development

Morris, E. P. and da Fonseca, P. C.A. (2017) High-resolution cryo-EM proteasome structures in drug development. Acta Crystallographica. Section D: Structural Biology, 73(6), pp. 522-533. (doi: 10.1107/S2059798317007021) (PMID:28580914) (PMCID:PMC5458494)

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Abstract

With the recent advances in biological structural electron microscopy (EM), protein structures can now be obtained by cryo-EM and single-particle analysis at resolutions that used to be achievable only by crystallographic or NMR methods. We have explored their application to study protein–ligand inter­actions using the human 20S proteasome, a well established target for cancer therapy that is also being investigated as a target for an increasing range of other medical conditions. The map of a ligand-bound human 20S proteasome served as a proof of principle that cryo-EM is emerging as a realistic approach for more general structural studies of protein–ligand interactions, with the potential benefits of extending such studies to complexes that are unfavourable to other methods and allowing structure determination under conditions that are closer to physiological, preserving ligand specificity towards closely related binding sites. Subsequently, the cryo-EM structure of the Plasmodium falciparum 20S proteasome, with a new prototype specific inhibitor bound, revealed the molecular basis for the ligand specificity towards the parasite complex, which provides a framework to guide the development of highly needed new-generation antimalarials. Here, the cryo-EM analysis of the ligand-bound human and P. falciparum 20S proteasomes is reviewed, and a complete description of the methods used for structure determination is provided, including the strategy to overcome the bias orientation of the human 20S proteasome on electron-microscope grids and details of the icr3d software used for three-dimensional reconstruction.

Item Type:Articles
Additional Information:Proceedings of the CCP-EM Spring Symposium 2017, Didcot, UK, 26-26 Apr 2017.
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:da Fonseca, Professor Paula
Authors: Morris, E. P., and da Fonseca, P. C.A.
College/School:College of Medical Veterinary and Life Sciences > School of Molecular Biosciences
Journal Name:Acta Crystallographica. Section D: Structural Biology
Publisher:International Union of Crystallography
ISSN:2059-7983
ISSN (Online):2059-7983
Published Online:31 May 2017
Copyright Holders:Copyright © 2017 The Authors
First Published:First published in Acta Crystallographica. Section D: Structural Biology 73(6): 522-533
Publisher Policy:Reproduced in accordance with the publisher copyright policy

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