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In Vivo Hydroxyl Radical Protein Footprinting for the Study of Protein Interactions in Caenorhabditis elegans
Published on: April 1, 2020
In vitro oxidative inactivation of human presequence protease (hPreP)
Pedro Filipe Teixeira1, Catarina Moreira Pinho, Rui M Branca
1Arrhenius Laboratories for Natural Sciences, Department of Biochemistry and Biophysics, Stockholm University, SE-106 91 Stockholm, Sweden. pedro@dbb.su.se
Free Radical Biology & Medicine
|October 9, 2012
Summary
Hydrogen peroxide (H2O2) inhibits human presequence protease (hPreP) activity, impacting mitochondrial function. Methionine 206 may protect hPreP from oxidation, and methionine sulfoxide reductase A (MsrA) can restore its function.
Area of Science:
- Mitochondrial biology
- Enzymology
- Neuroscience
Background:
- Presequence protease (PreP) degrades mitochondrial peptides, including amyloid-beta.
- Reduced PreP activity is linked to Alzheimer's disease (AD) and increased mitochondrial reactive oxygen species (ROS).
Purpose of the Study:
- To investigate the effects of hydrogen peroxide (H2O2) on recombinant human PreP (hPreP) activity.
- To explore the role of methionine 206 (M206) and methionine sulfoxide reductase A (MsrA) in hPreP redox regulation.
Main Methods:
- In vitro assays using recombinant human PreP (hPreP).
- Exposure to varying concentrations of hydrogen peroxide (H2O2).
- Analysis of protein oxidation (carbonylation) and stability, and enzyme activity restoration.
Main Results:
- H2O2 inhibited hPreP activity in a dose-dependent manner, causing oxidation and reduced stability.
- Mutation of M206 to leucine increased hPreP sensitivity to oxidation, suggesting M206 acts as an internal antioxidant.
- Methionine sulfoxide reductase A (MsrA) restored the activity of oxidized hPreP, indicating hPreP is an MsrA substrate.
Conclusions:
- hPreP activity is redox-controlled in the mitochondrial matrix.
- Conserved M206 residue likely protects hPreP from oxidative damage.
- MsrA can repair oxidative damage to hPreP, maintaining mitochondrial function.
