Manipulation of oxidative protein folding and PDI redox state in mammalian cells.
In the endoplasmic reticulum (ER), disulfide bonds are simultaneously formed in nascent proteins and removed from incorrectly folded or assembled molecules. In this compartment, the redox state must be, therefore, precisely regulated. Here we show that both human Ero1-Lalpha and Ero1-Lbeta (hEROs) facilitate disulfide bond formation in immunoglobulin subunits by selectively oxidizing PDI. Disulfide bond formation is controlled by hEROs, which stand at a crucial point of an electron-flow starting from nascent secretory proteins and passing through PDI. The redox state of ERp57, another ER-resident oxidoreductase, is not affected by over-expression of Ero1-Lalpha, suggesting that parallel and specific pathways control oxidative protein folding in the ER. Mutants in the Ero1-Lalpha CXXCXXC motif act as dominant negatives by limiting immunoglobulin oxidation. PDI-dependent oxidative folding in living cells can thus be manipulated by using hERO variants.
Mezghrani
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A
Department of Molecular Pathology and Medicine, DiBiT HSR, Milano, Italy.
Fassio
A
A
Benham
A
A
Simmen
T
T
Braakman
I
I
Sitia
R
R
D020816:Amino Acid Motifs; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation; D010084:Oxidation-Reduction; D010088:Oxidoreductases; D050862:Oxidoreductases Acting on Sulfur Group Donors; D010100:Oxygen / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D019704:Protein Disulfide-Isomerases / Q000737:chemistry* / Q000378:metabolism*; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D013997:Time Factors; D014162:Transfection
D004220:Disulfides; D007074:Immunoglobulin G; D008562:Membrane Glycoproteins; C405327:ERO1A protein, human; D010088:Oxidoreductases; D050862:Oxidoreductases Acting on Sulfur Group Donors; C485767:ERO1B protein, human; D019704:Protein Disulfide-Isomerases; D003545:Cysteine; D010100:Oxygen