title: Ero1-α and PDIs constitute a hierarchical electron transfer network of endoplasmic reticulum oxidoreductases.
abstract: Ero1-α and endoplasmic reticulum (ER) oxidoreductases of the protein disulfide isomerase (PDI) family promote the efficient introduction of disulfide bonds into nascent polypeptides in the ER. However, the hierarchy of electron transfer among these oxidoreductases is poorly understood. In this paper, Ero1-α-associated oxidoreductases were identified by proteomic analysis and further confirmed by surface plasmon resonance. Ero1-α and PDI were found to constitute a regulatory hub, whereby PDI induced conformational flexibility in an Ero1-α shuttle cysteine (Cys99) facilitated intramolecular electron transfer to the active site. In isolation, Ero1-α also oxidized ERp46, ERp57, and P5; however, kinetic measurements and redox equilibrium analysis revealed that PDI preferentially oxidized other oxidoreductases. PDI accepted electrons from the other oxidoreductases via its a' domain, bypassing the a domain, which serves as the electron acceptor from reduced glutathione. These observations provide an integrated picture of the hierarchy of cooperative redox interactions among ER oxidoreductases in mammalian cells.
authors.affiliation: Molecular Profiling Research Center for Drug Discovery, National Institute of Advanced Industrial Science and Technology, Koto-ku, Tokyo 135-0064, Japan.
authors.forename: Kazutaka
authors.initials: K
authors.lastname: Araki
authors.forename: Shun-ichiro
authors.initials: S
authors.lastname: Iemura
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Kamiya
authors.forename: David
authors.initials: D
authors.lastname: Ron
authors.forename: Koichi
authors.initials: K
authors.lastname: Kato
authors.forename: Tohru
authors.initials: T
authors.lastname: Natsume
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Nagata
mesh_terms: D020134:Catalytic Domain; D004579:Electron Transport; D004583:Electrons; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D005978:Glutathione / Q000378:metabolism; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D009682:Magnetic Resonance Spectroscopy; D008562:Membrane Glycoproteins / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism*; D010101:Oxygen Consumption; D019704:Protein Disulfide-Isomerases / Q000378:metabolism*; D040901:Proteomics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D020349:Surface Plasmon Resonance
chemical_list: D008562:Membrane Glycoproteins; C405327:ERO1A protein, human; D010088:Oxidoreductases; D019704:Protein Disulfide-Isomerases; D005978:Glutathione
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