Rawson, S., Bisson, C. orcid.org/0000-0002-9430-6822, Hurdiss, D.L. orcid.org/0000-0003-3834-5808 et al. (6 more authors) (2018) Elucidating the structural basis for differing enzyme inhibitor potency by cryo-EM. Proceedings of the National Academy of Sciences, 115 (8). pp. 1795-1800. ISSN 0027-8424
Abstract
Histidine biosynthesis is an essential process in plants and microorganisms, making it an attractive target for the development of herbicides and antibacterial agents. Imidazoleglycerol-phosphate dehydratase (IGPD), a key enzyme within this pathway, has been biochemically characterized in bothSaccharomyces cerevisiae(Sc_IGPD) andArabidopsis thaliana(At_IGPD). The plant enzyme, having been the focus of in-depth structural analysis as part of an inhibitor development program, has revealed details about the reaction mechanism of IGPD, whereas the yeast enzyme has proven intractable to crystallography studies. The structure-activity relationship of potent triazole-phosphonate inhibitors of IGPD has been determined in both homologs, revealing that the lead inhibitor (C348) is an order of magnitude more potent againstSc_IGPD thanAt_IGPD; however, the molecular basis of this difference has not been established. Here we have used single-particle electron microscopy (EM) to study structural differences between theAtandSc_IGPD homologs, which could influence the difference in inhibitor potency. The resulting EM maps at ∼3 Å are sufficient to de novo build the protein structure and identify the inhibitor binding site, which has been validated against the crystal structure of theAt_IGPD/C348 complex. The structure ofSc_IGPD reveals that a 24-amino acid insertion forms an extended loop region on the enzyme surface that lies adjacent to the active site, forming interactions with the substrate/inhibitor binding loop that may influence inhibitor potency. Overall, this study provides insights into the IGPD family and demonstrates the power of using an EM approach to study inhibitor binding.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2018 the Author(s). Published by PNAS. This open access article is distributed under Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (https://creativecommons.org/licenses/by-nc-nd/4.0/). |
Keywords: | IGPD; electron microscopy; enzyme inhibition; histidine biosynthesis; structure-based drug design |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > School of Biosciences (Sheffield) > Department of Molecular Biology and Biotechnology (Sheffield) |
Funding Information: | Funder Grant number SYNGENTA AG PO 8700157282 BIOTECHNOLOGY AND BIOLOGICAL SCIENCES RESEARCH COUNCIL (BBSRC) BB/I003703/1 BIOTECHNOLOGY AND BIOLOGICAL SCIENCES RESEARCH COUNCIL (BBSRC) Vacation Studentship BIOTECHNOLOGY AND BIOLOGICAL SCIENCES RESEARCH COUNCIL (BBSRC) BB/D524975/1 |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 27 Feb 2018 16:25 |
Last Modified: | 18 Apr 2024 16:01 |
Status: | Published |
Publisher: | National Academy of Sciences |
Refereed: | Yes |
Identification Number: | 10.1073/pnas.1708839115 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:127740 |