Muench, SP, Rawson, S, Eyraud, V et al. (7 more authors) (2014) PA1b inhibitor binding to subunits c and e of the vacuolar ATPase reveals Its insecticidal mechanism. Journal of Biological Chemistry, 289 (23). 16399 - 16408. ISSN 0021-9258
Abstract
The vacuolar ATPase (V-ATPase) is a 1MDa transmembrane proton pump that operates via a rotary mechanism fuelled by ATP. Essential for eukaryotic cell homeostasis, it plays central roles in bone remodeling and tumor invasiveness, making it a key therapeutic target. Its importance in arthropod physiology also makes it a promising pesticide target. The major challenge in designing lead compounds against the V-ATPase is its ubiquitous nature, such that any therapeutic must be capable of targeting particular isoforms. Here, we have characterized the binding site on the V-ATPase of pea albumin 1b (PA1b), a small cystine knot protein that shows exquisitely selective inhibition of insect V-ATPases. Electron microscopy shows that PA1b binding occurs across a range of equivalent sites on the c ring of the membrane domain. In the presence of Mg-�ATP, PA1b localizes to a single site, distant from subunit a, which is predicted to be the interface for other inhibitors. Photoaffinity labeling studies show radiolabeling of subunits c and e. In addition, weevil resistance to PA1b is correlated with bafilomycin resistance, caused by mutation of subunit c. The data indicate a binding site to which both subunits c and e contribute and inhibition that involves locking the c ring rotor to a static subunit e and not subunit a. This has implications for understanding the V-ATPase mechanism and that of inhibitors with therapeutic or pesticidal potential. It also provides the first evidence for the position of subunit e within the complex
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | (c) 2014, The Authors. This is an Open Access article distributed in accordance with the Creative Commons Attribution (CC BY 3.0) licence, which permits others to distribute, remix, adapt, build upon this work, and license their derivative works on different terms, provided the original work is properly cited. |
Keywords: | ATP; ATPase; Bafilomycin; Bafilomycin resistance; Binding; Binding site; Bone; C ring; C-ring; Cell; Complex; Complexes; Domain; Electron microscopy; Electron-mocroscopy; Ha; Inhibition; Inhibitor; Inhibitor binding; Inhibitors; Isoform; Lisoforms; Mechanism; Membrane; Membrane domain; Microscopy; Mutation; Photoaffinity labeling; Protein; Proton; Proton pump; Pump; Resitance; Rotor; Site; Sites; Subunit; Subunit A; Subunit C; Subunit E; Subunit-C; Subunit-E; Subunits; Target; Tumor; V ATPase; V ATPases; V-ATPase; V-ATPases; Vacuolar; Vacuolar ATPase; Vacuolar-ATPase |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 26 Mar 2015 18:22 |
Last Modified: | 18 Aug 2015 19:45 |
Published Version: | http://dx.doi.org/10.1074/jbc.M113.541250 |
Status: | Published |
Publisher: | American Society for Biochemistry and Molecular Biology |
Identification Number: | 10.1074/jbc.M113.541250 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:83668 |