Hypochlorous acid modifies calcium release channel function from skeletal muscle sarcoplasmic reticulum

Area of Science:

• Biochemistry
• Molecular Biology
• Physiology

Background:

• Hydrogen peroxide (H2O2) at millimolar concentrations releases Ca(2+) from sarcoplasmic reticulum (SR) vesicles.
• Hypochlorous acid (HOCl) is a potent oxidant, suggesting potential effects on protein function.
• The SR Ca(2+) release channel is crucial for muscle contraction and calcium homeostasis.

Purpose of the Study:

• To investigate the effects of HOCl on the SR Ca(2+) release channel mechanism.
• To determine the concentration-dependent effects of HOCl on Ca(2+) release and ryanodine binding.
• To elucidate the interaction of HOCl with critical thiol groups in the SR Ca(2+) release channel.

Main Methods:

• Assessing Ca(2+) release from actively loaded SR vesicles treated with varying HOCl concentrations.
• Measuring [(3)H]ryanodine binding to SR Ca(2+) release channels.
• Conducting single-channel reconstitution experiments with SR vesicles and HOCl.
• Utilizing a fluorogenic probe to detect modifications of thiol groups on SR vesicles.

Main Results:

• HOCl activates the SR Ca(2+) release channel and induces rapid Ca(2+) release in a concentration-dependent manner.
• HOCl enhances high-affinity [(3)H]ryanodine binding at 10-100 microM but inhibits it at higher concentrations.
• HOCl directly activates and subsequently inhibits Ca(2+) channel activity in single-channel recordings.
• HOCl treatment modifies thiol groups essential for SR Ca(2+) channel function.

Conclusions:

• Micromolar concentrations of HOCl can significantly alter SR Ca(2+) handling.
• HOCl acts as a modulator of the SR Ca(2+) release channel, impacting Ca(2+) flux.
• The findings highlight HOCl's role in modifying critical thiol residues within the SR Ca(2+) release mechanism.