Global Protein Oxidation Profiling Suggests Efficient Mitochondrial Proteome Homeostasis During Aging

Area of Science:

• Gerontology
• Biochemistry
• Molecular Biology

Background:

• The free radical theory of aging posits that reactive oxygen species (ROS) cause age-related protein oxidation.
• Mitochondria are a major source of cellular ROS, making mitochondrial proteins key targets for oxidative damage during aging.

Purpose of the Study:

• To investigate ROS-induced oxidative damage in the mitochondrial proteome of the aging model Podospora anserina.
• To quantify oxidative modifications and compare oxidized versus non-oxidized protein species.

Main Methods:

• Developed a gel-free sample preparation method with reducing and iron-chelating agents to minimize artificial oxidation.
• Employed long gradient nLC-ESI-MS/MS for sensitive peptide separation and iTRAQ quantification.
• Established a statistical workflow for comprehensive analysis of oxidative modifications.

Main Results:

• Quantified 2341 proteins, with 746 showing both unmodified and oxidatively modified species.
• Identified 22 distinct oxidative amino acid modifications, with methionine oxidation being most prevalent, followed by carbonylation, N-formylkynurenine, and pyrrolidinone formation.
• Observed a positive correlation between protein amount changes and oxidative damage for most proteins, and a decrease in protein amounts at late age.

Conclusions:

• Podospora anserina effectively counteracts ROS-induced protein damage during aging, provided protein synthesis is functional.
• Findings suggest a protein damage homeostasis mechanism rather than a massive increase in protein oxidation with age.
• The relationship between damaged and undamaged protein species remains relatively constant even at late stages of aging.