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PreprintCited by 1Enzyme Structure and FunctionProtein Structure and DynamicsComputational Drug Discovery Methods

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Structure and catalytic mechanism of methylisocitrate lyase, a potential drug target against Coxiella burnetii

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Stuart, William S., Jenkins, Christopher H., Ireland, Philip M., Isupov, Michail N., Norville, Isobel H., Harmer, Nicholas J.·2025·Journal of Biological Chemistry
DOI 10.1101/2025.01.06.631558PMC PMC12145553

Preprint — published as 10.1016/j.jbc.2025.108517 · Paper page

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Abstract

We present a comprehensive investigation into the catalytic mechanism of methylisocitrate lyase, a potential drug target candidate against the zoonotic pathogen Coxiella burnetii , the causative agent of Q fever and a federal select agent. Current treatment regimens are prolonged, often with incomplete clearance of the pathogen. We utilised a structure-based bioinformatics pipeline to identify methylisocitrate lyase as a candidate therapeutic target against C. burnetii from a list of essential genes. Wild-type C. burnetii methylisocitrate lyase has a k cat of 32,000 s -1 (compared to 105 s -1 for Salmonella enterica ) and isocitrate inhibits with a K I of 6 mM. We have determined the previously uncharacterised substrate-bound structure of this enzyme family, alongside product and inhibitor-bound structures. These structures of wild-type enzyme reveal that in the active state the catalytic C118 is positioned 2.98 Å from O5 of methylisocitrate and Arg152 moves towards the substrate relative to the inhibitor bound structure. Analysis of structure-based mutants reveals that Arg152 and Glu110 are both essential for catalysis. We suggest that Arg152 acts as the catalytic base that initiates the methylisocitrate lyase reaction. These results deepen our understanding of the catalytic mechanism of methylisocitrate lyase and could aid the development of new therapeutics against C. burnetii .

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Metadata

Authors
Stuart, W. S., Jenkins, C. H., Ireland, P. M., Isupov, M. N., Norville, I. H., & Harmer, N. J.
Affiliations
  • University of Exeter (GB)
  • Defence Science and Technology Laboratory (GB)
Corresponding author
Harmer, Nicholas J.
ORCID
Venue
Journal of Biological Chemistry
Publisher
openRxiv
Published
2025-01-06
Volume
301
Issue
6
Pages
108517-108517
ISSN
0021-9258 (ISSN-L)
DOI
10.1101/2025.01.06.631558
PMC
PMC12145553
OpenAlex
W4409499652
Cited by
1
References
1
Citations by year
2026: 1
FWCI
0.21
Type
journal article
Language
en
Open access
Licence
creativecommons.org/licenses/by/4.0/ (vor)
Locations
  • Journal of Biological Chemistry (publishedVersion)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
  • PubMed Central (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • Defence Science and Technology Laboratory
  • Biotechnology and Biological Sciences Research Council (BB/M009122/1)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Enzyme Structure and Function
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Drug Discovery › Computational Drug Discovery Methods
Keywords
Coxiella burnetii, Q fever, catalytic mechanism, therapeutic target, essential genes, enzyme kinetics
MeSH terms
Bacterial Proteins, Catalysis, Enzyme Inhibitors, Coxiella burnetii, Crystallography, X-Ray, Catalytic Domain

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