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

• Published in: Archives of biochemistry and biophysics Vol. 579; pp. 1 - 7
• Main Authors: Hudson, Devin A., Thorpe, Colin
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.08.2015
• Subjects: Amino Acid Sequence; Augmenter of liver regeneration; Binding Sites; Computer Simulation; Cytochrome Reductases - chemistry; Cytochrome Reductases - ultrastructure; Disulfide exchange; Enzyme Activation; Humans; Isomerases - chemistry; Isomerases - ultrastructure; Mia40; Mitochondrial intermembrane space targeting sequence; Mitochondrial Membrane Transport Proteins - chemistry; Mitochondrial Membrane Transport Proteins - ultrastructure; Models, Chemical; Models, Molecular; Molecular Sequence Data; Oxidants - chemistry; Oxidation-Reduction; Oxidative protein folding; Oxidoreductases Acting on Sulfur Group Donors; Protein Binding; Protein Conformation; Protein disulfide isomerase; Protein Folding; Structure-Activity Relationship; Mitochondrial intermembrane space targeting sequence; Protein disulfide isomerase; Disulfide exchange; Augmenter of liver regeneration; Oxidative protein folding; Mia40
• ISSN: 0003-9861; 1096-0384
• DOI: 10.1016/j.abb.2015.05.005

[Display omitted] •Mia40 is a facile oxidant of a range of non-cognate unfolded reduced proteins.•This broad specificity impacts consideration of protein trafficking within the IMS.•Mia40 shows minimal disulfide isomerase activity with scrambled proteins.•The need for additional disulfide isomerase activity in the IMS is considered. Mia40 participates in oxidative protein folding within the mitochondrial intermembrane space (IMS) by mediating the transfer of reducing equivalents from client proteins to FAD-linked oxidoreductases of the Erv1 family (lfALR in mammals). Here we investigate the specificity of the human Mia40/lfALR system towards non-cognate unfolded protein substrates to assess whether the efficient introduction of disulfides requires a particular amino acid sequence context or the presence of an IMS targeting signal. Reduced pancreatic ribonuclease A (rRNase), avian lysozyme, and riboflavin binding protein are all competent substrates of the Mia40/lfALR system, although they lack those sequence features previously thought to direct disulfide bond formation in cognate IMS substrates. The oxidation of rRNase by Mia40 does not limit overall turnover of unfolded substrate by the Mia40/lfALR system. Mia40 is an ineffective protein disulfide isomerase when its ability to restore enzymatic activity from scrambled RNase is compared to that of protein disulfide isomerase. Mia40’s ability to bind amphipathic peptides is evident by avid binding to the isolated B-chain during the insulin reductase assay. In aggregate these data suggest that the Mia40/lfALR system has a broad sequence specificity and that potential substrates may be protected from adventitious oxidation by kinetic sequestration within the mitochondrial IMS.