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Title: Studies of Toxoplasma gondii and Plasmodium falciparum enoyl acyl carrier protein reductase and implications for the development of antiparasitic agents

Abstract

The crystal structures of T. gondii and P. falciparum ENR in complex with NAD{sup +} and triclosan and of T. gondii ENR in an apo form have been solved to 2.6, 2.2 and 2.8 Å, respectively. Recent studies have demonstrated that submicromolar concentrations of the biocide triclosan arrest the growth of the apicomplexan parasites Plasmodium falciparum and Toxoplasma gondii and inhibit the activity of the apicomplexan enoyl acyl carrier protein reductase (ENR). The crystal structures of T. gondii and P. falciparum ENR in complex with NAD{sup +} and triclosan and of T. gondii ENR in an apo form have been solved to 2.6, 2.2 and 2.8 Å, respectively. The structures of T. gondii ENR have revealed that, as in its bacterial and plant homologues, a loop region which flanks the active site becomes ordered upon inhibitor binding, resulting in the slow tight binding of triclosan. In addition, the T. gondii ENR–triclosan complex reveals the folding of a hydrophilic insert common to the apicomplexan family that flanks the substrate-binding domain and is disordered in all other reported apicomplexan ENR structures. Structural comparison of the apicomplexan ENR structures with their bacterial and plant counterparts has revealed that although the active sites ofmore » the parasite enzymes are broadly similar to those of their bacterial counterparts, there are a number of important differences within the drug-binding pocket that reduce the packing interactions formed with several inhibitors in the apicomplexan ENR enzymes. Together with other significant structural differences, this provides a possible explanation of the lower affinity of the parasite ENR enzyme family for aminopyridine-based inhibitors, suggesting that an effective antiparasitic agent may well be distinct from equivalent antimicrobials.« less

Authors:
 [1];  [2];  [3];  [1];  [3];  [4];  [3];  [1]
  1. The Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, University of Sheffield, Firth Court, Western Bank, Sheffield S10 2TN (United Kingdom)
  2. Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, MD 21205 (United States)
  3. Department of Ophthalmology and Visual Sciences, Paediatrics (Infectious Diseases) and Pathology and the Committees on Molecular Medicine, Genetics, Immunology and The College, The University of Chicago, Chicago, IL 60637 (United States)
  4. Department of Immunology, University of Strathclyde, Glasgow G4 0NR, Scotland (United Kingdom)
Publication Date:
OSTI Identifier:
22348014
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section D: Biological Crystallography; Journal Volume: 63; Journal Issue: Pt 3; Other Information: PMCID: PMC2483495; PMID: 17327670; PUBLISHER-ID: en5210; OAI: oai:pubmedcentral.nih.gov:2483495; Copyright (c) International Union of Crystallography 2007; This is an open-access article distributed under the terms described at http://journals.iucr.org/services/termsofuse.html.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
Denmark
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; AFFINITY; CARRIERS; CRYSTAL STRUCTURE; CRYSTALS; INTERACTIONS; PLASMA; STOWING; SUBSTRATES

Citation Formats

Muench, Stephen P., Prigge, Sean T., McLeod, Rima, Rafferty, John B., Kirisits, Michael J., Roberts, Craig W., Mui, Ernest J., and Rice, David W., E-mail: d.rice@sheffield.ac.uk. Studies of Toxoplasma gondii and Plasmodium falciparum enoyl acyl carrier protein reductase and implications for the development of antiparasitic agents. Denmark: N. p., 2007. Web. doi:10.1107/S0907444906053625.
Muench, Stephen P., Prigge, Sean T., McLeod, Rima, Rafferty, John B., Kirisits, Michael J., Roberts, Craig W., Mui, Ernest J., & Rice, David W., E-mail: d.rice@sheffield.ac.uk. Studies of Toxoplasma gondii and Plasmodium falciparum enoyl acyl carrier protein reductase and implications for the development of antiparasitic agents. Denmark. doi:10.1107/S0907444906053625.
Muench, Stephen P., Prigge, Sean T., McLeod, Rima, Rafferty, John B., Kirisits, Michael J., Roberts, Craig W., Mui, Ernest J., and Rice, David W., E-mail: d.rice@sheffield.ac.uk. Thu . "Studies of Toxoplasma gondii and Plasmodium falciparum enoyl acyl carrier protein reductase and implications for the development of antiparasitic agents". Denmark. doi:10.1107/S0907444906053625.
@article{osti_22348014,
title = {Studies of Toxoplasma gondii and Plasmodium falciparum enoyl acyl carrier protein reductase and implications for the development of antiparasitic agents},
author = {Muench, Stephen P. and Prigge, Sean T. and McLeod, Rima and Rafferty, John B. and Kirisits, Michael J. and Roberts, Craig W. and Mui, Ernest J. and Rice, David W., E-mail: d.rice@sheffield.ac.uk},
abstractNote = {The crystal structures of T. gondii and P. falciparum ENR in complex with NAD{sup +} and triclosan and of T. gondii ENR in an apo form have been solved to 2.6, 2.2 and 2.8 Å, respectively. Recent studies have demonstrated that submicromolar concentrations of the biocide triclosan arrest the growth of the apicomplexan parasites Plasmodium falciparum and Toxoplasma gondii and inhibit the activity of the apicomplexan enoyl acyl carrier protein reductase (ENR). The crystal structures of T. gondii and P. falciparum ENR in complex with NAD{sup +} and triclosan and of T. gondii ENR in an apo form have been solved to 2.6, 2.2 and 2.8 Å, respectively. The structures of T. gondii ENR have revealed that, as in its bacterial and plant homologues, a loop region which flanks the active site becomes ordered upon inhibitor binding, resulting in the slow tight binding of triclosan. In addition, the T. gondii ENR–triclosan complex reveals the folding of a hydrophilic insert common to the apicomplexan family that flanks the substrate-binding domain and is disordered in all other reported apicomplexan ENR structures. Structural comparison of the apicomplexan ENR structures with their bacterial and plant counterparts has revealed that although the active sites of the parasite enzymes are broadly similar to those of their bacterial counterparts, there are a number of important differences within the drug-binding pocket that reduce the packing interactions formed with several inhibitors in the apicomplexan ENR enzymes. Together with other significant structural differences, this provides a possible explanation of the lower affinity of the parasite ENR enzyme family for aminopyridine-based inhibitors, suggesting that an effective antiparasitic agent may well be distinct from equivalent antimicrobials.},
doi = {10.1107/S0907444906053625},
journal = {Acta Crystallographica. Section D: Biological Crystallography},
number = Pt 3,
volume = 63,
place = {Denmark},
year = {Thu Mar 01 00:00:00 EST 2007},
month = {Thu Mar 01 00:00:00 EST 2007}
}
  • Acyl carrier protein (ACP) plays a central role in fatty acid biosynthesis by transferring the acyl groups from one enzyme to another for the completion of the fatty acid synthesis cycle. Holo-ACP is the obligatory substrate for the synthesis of acyl-ACPs which act as the carrier and donor for various metabolic reactions. Despite its interactions with numerous proteins in the cell, its mode of interaction is poorly understood. Here, we report the over-expression of PfACP in minimal medium solely in its holo form and in high yield. Expression in minimal media provides a means to isotopically label PfACP for highmore » resolution multi-nuclear and multi-dimensional NMR studies. Indeed, the proton-nitrogen correlated NMR spectrum exhibits very high chemical shift dispersion and resolution. We also show that holo-PfACP thus expressed is amenable to acylation reactions using Escherichia coli acyl-ACP synthetase as well as by standard chemical methods.« less
  • Enoyl-acyl carrier protein reductase (FabI) catalyzes the last rate-limiting step in the elongation cycle of the fatty-acid biosynthesis pathway and has been validated as a potential antimicrobial drug target in Francisella tularensis. The development of new antibiotic therapies is important both to combat potential drug-resistant bioweapons and to address the broader societal problem of increasing antibiotic resistance among many pathogenic bacteria. The crystal structure of FabI from F. tularensis (FtuFabI) in complex with the inhibitor triclosan and the cofactor NAD{sup +} has been solved to a resolution of 2.1 {angstrom}. Triclosan is known to effectively inhibit FabI from different organisms.more » Precise characterization of the mode of triclosan binding is required to develop highly specific inhibitors. Comparison of our structure with the previously determined FtuFabI structure (PDB code 2jjy) which is bound to only NAD{sup +} reveals the conformation of the substrate-binding loop, electron density for which was missing in the earlier structure, and demonstrates a shift in the conformation of the NAD{sup +} cofactor. This shift in the position of the phosphate groups allows more room in the active site for substrate or inhibitor to bind and be better accommodated. This information will be crucial for virtual screening studies to identify novel scaffolds for development into new active inhibitors.« less
  • Enoyl-acyl carrier protein (ACP) reductases are responsible for bacterial type II fatty-acid biosynthesis and are attractive targets for developing novel antibiotics. The S. pneumoniae enoyl-ACP reductase (FabK) was crystallized and selenomethionine MAD data were collected to 2 Å resolution. The enoyl-acyl carrier protein (ACP) reductase from Streptococcus pneumoniae (FabK; EC 1.3.1.9) is responsible for catalyzing the final step in each elongation cycle of fatty-acid biosynthesis. Selenomethionine-substituted FabK was purified and crystallized by the hanging-drop vapour-diffusion method at 277 K. The crystal belongs to space group P2{sub 1}, with unit-cell parameters a = 50.26, b = 126.70, c = 53.63 Å,more » β = 112.46°. Diffraction data were collected to 2.00 Å resolution using synchrotron beamline BL32B2 at SPring-8. Two molecules were estimated to be present in the asymmetric unit, with a solvent content of 45.1%.« less