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Cited by 9Enzyme Structure and FunctionNeutrophil, Myeloperoxidase and Oxidative MechanismsRedox biology and oxidative stress

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X-ray structure and enzymatic study of a bacterial NADPH oxidase highlight the activation mechanism of eukaryotic NOX

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Petit-Hartlein, Isabelle, Vermot, Annelise, Thepaut, Michel, Humm, Anne-Sophie, Dupeux, Florine, Dupuy, Jerome, Chaptal, Vincent, Marquez, Jose Antonio, Smith, Susan ME, Fieschi, Franck·2024·eLife
DOI 10.7554/elife.93759.3

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Abstract

NADPH oxidases (NOX) are transmembrane proteins, widely spread in eukaryotes and prokaryotes, that produce reactive oxygen species (ROS). Eukaryotes use the ROS products for innate immune defense and signaling in critical (patho)physiological processes. Despite the recent structures of human NOX isoforms, the activation of electron transfer remains incompletely understood. SpNOX, a homolog from Streptococcus pneumoniae , can serves as a robust model for exploring electron transfers in the NOX family thanks to its constitutive activity. Crystal structures of SpNOX full-length and dehydrogenase (DH) domain constructs are revealed here. The isolated DH domain acts as a flavin reductase, and both constructs use either NADPH or NADH as substrate. Our findings suggest that hydride transfer from NAD(P)H to FAD is the rate-limiting step in electron transfer. We identify significance of F397 in nicotinamide access to flavin isoalloxazine and confirm flavin binding contributions from both DH and Transmembrane (TM) domains. Comparison with related enzymes suggests that distal access to heme may influence the final electron acceptor, while the relative position of DH and TM does not necessarily correlate with activity, contrary to previous suggestions. It rather suggests requirement of an internal rearrangement, within the DH domain, to switch from a resting to an active state. Thus, SpNOX appears to be a good model of active NOX2, which allows us to propose an explanation for NOX2’s requirement for activation.

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Metadata

Authors
Petit-Hartlein, I., Vermot, A., Thepaut, M., Humm, A.-S., Dupeux, F., Dupuy, J., Chaptal, V., Marquez, J. A., Smith, S. M., & Fieschi, F.
Affiliations
  • Univ. Grenoble Alpes, CNRS, CEA, Institut de Biologie Structurale
  • Centre National de la Recherche Scientifique (FR)
  • European Molecular Biology Laboratory (DE)
  • Lyon 1 Université (FR)
  • European Molecular Biology Laboratory
  • Kennesaw State University (US)
ORCID
Venue
eLife
Publisher
eLife Sciences Publications, Ltd
Published
2024-04-19
Volume
13
ISSN
2050-084X (ISSN-L)
DOI
10.7554/elife.93759.3
OpenAlex
W4394951969
Cited by
9
References
2
Citations by year
2026: 5 · 2025: 2 · 2024: 2
FWCI
0.88
Type
journal article
Language
en
Open access
Licence
Locations
eLife (publishedVersion, cc-by)
Funders
  • European Commission (iNEXT Discovery (grant agreement ID: 871037), Fragment Screen (grant agreement ID: 101094131))
  • Agence Nationale de la Recherche (FRISBI (ANR-10- INSB-0502), GRAL (ANR-10-LABX-49-01), Bandit (ANR17-CE11-0013))
  • Université Grenoble Alpes (Emergence program, Invited professorship)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Enzyme Structure and Function
  • Life Sciences › Immunology and Microbiology › Immunology › Neutrophil, Myeloperoxidase and Oxidative Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Redox biology and oxidative stress
Keywords
NADPH oxidase, Streptococcus pneumoniae, flavin reductase, electron transfer, hydride transfer, FAD, flavin binding, reactive oxygen species, crystal structure, enzyme activation mechanism

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