Unit 07: Stress and Redox Regulation Study Notes

Unit 07: Stress and Redox Regulation

1. Mechanisms of Redox Regulation and Signal Transduction by ROS

  • Redox Regulation Mechanisms

    • Nrf2/Keap1 Pathway

      • Involves the transcription factor Nrf2 (Nuclear Factor, Erythroid 2 Like 2) and the sensor protein Keap1 (Kelch Like ECH Associated Protein 1).

    • Peroxiredoxins

      • A family of peroxidases that reduce and inactivate hydrogen peroxide and organic peroxides.

    • Thioredoxin System

      • Consists of thioredoxin and thioredoxin reductase, and plays a critical role in redox homeostasis.

2. Redox State of Cells

  • Definition of Redox State

    • Refers to the overall balance of oxidized and reduced species in the cellular environment.

  • Redox Compartmentalization

    • Different cellular compartments (e.g., mitochondria, cytoplasm, extracellular space) have varying redox potentials and functions.

3. ROS Toxicity vs. ROS Signaling

  • ROS Toxicity

    • Certain ROS (e.g., hydroxyl radical, hypochlorous acid, peroxynitrite) can be harmful.

  • ROS Signaling

    • Molecules like hydrogen peroxide and nitric oxide act as essential physiological regulators of signaling pathways.

Key Concept: NOX Enzymes

  • NOX2: Involved in ROS production in phagocytes like neutrophils, vital for antimicrobial functions.

  • NOX1: Activated in intestinal epithelial cells by bacteria-released signals (e.g., fMLP), causing localized ROS production that affects cell functions such as proliferation and differentiation.

4. Redox-mediated Signaling

  • Specific redox regulation by ROS is executed by covalent modifications of cysteine residues in target proteins.

  • Oxidation of thiol (-SH) groups leads to reversible modifications of enzymatic activity.

Cysteine Biochemistry

  1. Formation of Disulfide Bonds

    • Two thiol groups can be oxidized to form covalent disulfide bonds (–S–S–).

  2. Oxidation Forms

    • Sulfenic Acid (RSOH): Relatively unstable and can further oxidize to sulfinic (RSO2H) and sulfonic forms (RSO3H).

  3. Glutathionylation (RSSG)

    • A post-translational modification where glutathione binds to proteins, forming disulfide bonds.

5. Redox-sensitive Signaling Molecules and Processes

  • ROS impact various cellular processes (proliferation, differentiation, survival) by interacting with redox-sensitive molecules.

  • Potential Damages: DNA damage, iron homeostasis issues, influences on anti-inflammatory responses.

  • Key Players in Redox Regulation:

    • Nrf2 and Keap1

    • Enzymes like peroxiredoxins and thioredoxin.

6. ROS-mediated Nrf2/Keap1 Signaling

  • Adaptive Signaling Pathway:

    • Under oxidative stress, modifications of Keap1 by ROS release and stabilize Nrf2.

    • Nrf2 then translocates to the nucleus and binds with Maf to activate antioxidant genes.

  • Cytoprotective Genes Upregulated by Nrf2:

    • Superoxide dismutase, catalase, glutathione peroxidase, which inactivate harmful ROS.

    • Enzymes that control thiol/disulfide exchanges and ROS regulation are also upregulated.

7. Peroxiredoxins (Prxs)

  • Definition:

    • A family of enzymes that reduce hydrogen peroxide, essential in cellular antioxidant defense.

    • Typically small proteins (22-30 kDa) found across all biological kingdoms.

Functionality of Peroxiredoxins

  1. Structure and Mechanism

    • Contain critical cysteine residues:

      • Peroxidatic Cysteine (Cys47): oxidized to sulfenic acid (Cys–SOH).

      • Resolving Cysteine (Cys170): forms disulfide bond for Prx functionality.

  2. Oxidation and Regeneration

    • Reaction mechanism allows for reversible oxidation.

    • Thioredoxin serves as a reductant to regenerate oxidized Prxs.

8. Thioredoxin System

  • Role:

    • Regulates protein thiol/disulfide balance through disulfide reductase activity.

  • Components:

    • Thioredoxin (Trx): Small reductase enzyme that catalyzes reductions via thiol-disulfide exchange.

    • Thioredoxin Reductase (TrxR): Selenoenzyme that reduces oxidized Trx using NADPH.

9. Redox State and Redox Potential

  • Definition and Calculation:

    • Redox state characterized quantitatively by redox potential calculated with the Nernst equation:
      E = E^0 - \frac{2.3RT}{zF}\log\left(\frac{[red]}{[ox]}\right)

  • Example of Redox Couples:

    • NAD+/NADH, GSSG/2GSH, cysteine/cystine.

  • GSH/GSSG Example:

    • Ratio is indicative of reducing conditions; typical intracellular levels are:

      • GSH: ~3-10 mM;

      • GSH/GSSG: >100/1.

      • Extracellular (plasma): GSH: ~2-10 µM; GSH/GSSG: ~5/1.

10. Plasma Redox Potential and Aging

  • Redox dynamics relate to aging; measured redox states can give insights into health conditions.

  • Health Metrics:

    • Suggested oxidative stress thresholds:

      • Cys/CySS: -62 mV;

      • GSH/GSSG: -119 mV.

  • Correlation:

    • Increased age correlates with rising oxidative events and changes in redox states.

11. Health Implications of Redox Regulation

  • Dietary Considerations:

    • Antioxidants in fruits and berries can positively influence redox states.

  • The maintenance of reduced microenvironment is essential for health and may influence aging processes.