Mia40 is a facile oxidant of unfolded reduced proteins but shows minimal isomerase activity

Area of Science:

• Mitochondrial biology
• Protein folding
• Biochemistry

Background:

• Mia40 is crucial for oxidative protein folding in the mitochondrial intermembrane space (IMS).
• It mediates electron transfer from client proteins to Erv1 (lfALR).
• Previous understanding suggested specific sequence contexts or IMS signals were required for efficient disulfide bond formation.

Purpose of the Study:

• To investigate the substrate specificity of the human Mia40/lfALR system.
• To determine if specific amino acid sequences or IMS targeting signals are essential for disulfide bond introduction.
• To assess the Mia40/lfALR system's interaction with non-cognate substrates.

Main Methods:

• Testing reduced pancreatic ribonuclease A, avian lysozyme, and riboflavin binding protein as substrates.
• Assessing the impact of rRNase oxidation on Mia40/lfALR system turnover.
• Comparing Mia40's protein disulfide isomerase activity to standard PDI.
• Evaluating Mia40's amphipathic peptide binding using an insulin reductase assay.

Main Results:

• Non-cognate substrates like rRNase, lysozyme, and riboflavin binding protein are efficiently oxidized by Mia40/lfALR.
• These substrates lack previously assumed sequence features for IMS disulfide bond formation.
• Mia40 is not an effective protein disulfide isomerase compared to PDI.
• Mia40 demonstrates avid binding to amphipathic peptides.

Conclusions:

• The Mia40/lfALR system exhibits broad sequence specificity for oxidative folding.
• Potential substrates may be protected from adventitious oxidation via kinetic sequestration within the IMS.
• This broad specificity expands the known roles of Mia40 in mitochondrial protein maturation.