Abstract
Ero1p is a key enzyme in the disulfide bond formation pathway in eukaryotic cells in both aerobic and anaerobic environments. It was previously demonstrated that Ero1p can transfer electrons from thiol substrates to molecular oxygen. However, the fate of electrons under anaerobic conditions and the final fate of electrons under aerobic conditions remained obscure. To address these fundamental issues in the Ero1p mechanism, we studied the transfer of electrons from recombinant yeast Ero1p to various electron acceptors. Under aerobic conditions, reduction of molecular oxygen by Ero1p yielded stoichiometric hydrogen peroxide. Remarkably, we found that reduced Ero1p can transfer electrons to a variety of small and macromolecular electron acceptors in addition to molecular oxygen. In particular, Ero1p can catalyze reduction of exogenous FAD in solution. Free FAD is not required for the catalysis of dithiol oxidation by Ero1p, but it is sufficient to drive disulfide bond formation under anaerobic conditions. These findings provide insight into mechanisms for regenerating oxidized Ero1p and maintaining disulfide bond formation under anaerobic conditions in the endoplasmic reticulum.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Binding Sites
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Catalysis
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Disulfides / metabolism*
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Electron Transport
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Endoplasmic Reticulum / enzymology*
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Flavins / pharmacology
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Glycoproteins / chemistry
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Glycoproteins / genetics
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Glycoproteins / metabolism*
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Hydrogen Peroxide / pharmacology
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Models, Molecular
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Oxidation-Reduction / drug effects
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Oxidoreductases Acting on Sulfur Group Donors / chemistry
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Oxidoreductases Acting on Sulfur Group Donors / genetics
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Oxidoreductases Acting on Sulfur Group Donors / metabolism*
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Protein Structure, Tertiary
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Saccharomyces cerevisiae / cytology
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Saccharomyces cerevisiae / enzymology
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism*
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Saccharomyces cerevisiae Proteins / chemistry
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism*
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Spectrum Analysis
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Substrate Specificity
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Sulfhydryl Compounds / metabolism
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Titrimetry
Substances
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Disulfides
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Flavins
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Glycoproteins
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Saccharomyces cerevisiae Proteins
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Sulfhydryl Compounds
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Hydrogen Peroxide
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Oxidoreductases Acting on Sulfur Group Donors
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ERO1 protein, S cerevisiae