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TitleStructural and mechanistic insights into Streptococcus pneumoniae NADPH oxidase.
Journal, issue, pagesNat Struct Mol Biol, Vol. 31, Issue 11, Page 1769-1777, Year 2024
Publish dateJul 22, 2024
AuthorsVictor R A Dubach / Pablo San Segundo-Acosta / Bonnie J Murphy /
PubMed AbstractNicotinamide adenine dinucleotide phosphate (NADPH) oxidases (NOXs) have a major role in the physiology of eukaryotic cells by mediating reactive oxygen species production. Evolutionarily distant ...Nicotinamide adenine dinucleotide phosphate (NADPH) oxidases (NOXs) have a major role in the physiology of eukaryotic cells by mediating reactive oxygen species production. Evolutionarily distant proteins with the NOX catalytic core have been found in bacteria, including Streptococcus pneumoniae NOX (SpNOX), which is proposed as a model for studying NOXs because of its high activity and stability in detergent micelles. We present here cryo-electron microscopy structures of substrate-free and nicotinamide adenine dinucleotide (NADH)-bound SpNOX and of NADPH-bound wild-type and F397A SpNOX under turnover conditions. These high-resolution structures provide insights into the electron-transfer pathway and reveal a hydride-transfer mechanism regulated by the displacement of F397. We conducted structure-guided mutagenesis and biochemical analyses that explain the absence of substrate specificity toward NADPH and suggest the mechanism behind constitutive activity. Our study presents the structural basis underlying SpNOX enzymatic activity and sheds light on its potential in vivo function.
External linksNat Struct Mol Biol / PubMed:39039317 / PubMed Central
MethodsEM (single particle)
Resolution2.2 - 2.64 Å
Structure data

EMDB-18644, PDB-8qt6:
Cryo-EM structure of Streptococcus pneumoniae NADPH oxidase
Method: EM (single particle) / Resolution: 2.29 Å

EMDB-18645, PDB-8qt7:
Cryo-EM structure of Streptococcus pneumoniae NADPH oxidase in complex with NADPH
Method: EM (single particle) / Resolution: 2.2 Å

EMDB-18646, PDB-8qt9:
Cryo-EM structure of stably reduced Streptococcus pneumoniae NADPH oxidase in complex with NADH
Method: EM (single particle) / Resolution: 2.36 Å

EMDB-18647, PDB-8qta:
Cryo-EM structure of Streptococcus pneumoniae NADPH oxidase F397A mutant in complex with NADPH
Method: EM (single particle) / Resolution: 2.64 Å

Chemicals

ChemComp-FAD:
FLAVIN-ADENINE DINUCLEOTIDE / FAD*YM

ChemComp-HEM:
PROTOPORPHYRIN IX CONTAINING FE

ChemComp-HOH:
WATER

ChemComp-NDP:
NADPH DIHYDRO-NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE

ChemComp-NAI:
1,4-DIHYDRONICOTINAMIDE ADENINE DINUCLEOTIDE

Source
  • streptococcus pneumoniae (bacteria)
KeywordsMEMBRANE PROTEIN / NADPH oxidase / ROS producing / flavoprotein / heme protein

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