Glutathione directly reduces an oxidoreductase in the endoplasmic reticulum of mammalian cells.
The formation of disulfide bonds is an essential step in the folding of many glycoproteins and secretory proteins. Non-native disulfide bonds are often formed between incorrect cysteine residues, and thus the cell has dedicated a family of oxidoreductases that are thought to isomerize non-native bonds. For an oxidoreductase to be capable of performing isomerization or reduction reactions, it must be maintained in a reduced state. Here we show that most of the oxidoreductases are predominantly reduced in vivo. Following oxidative stress the oxidoreductases are quickly reduced, demonstrating that a robust reductive pathway is in place in mammalian cells. Using ERp57 as a model we show that the reductive pathway is cytosol-dependent and that the component responsible for the reduction of the oxidoreductases is the low molecular mass thiol glutathione. In addition, ERp57 is not reduced following oxidative stress when inhibitors of glutathione synthesis or glutathione reduction are added to cells. Glutathione directly reduces ERp57 at physiological concentrations in vitro, and biotinylated glutathione forms a mixed disulfide with ERp57 in microsomes. Our results demonstrate that glutathione plays a direct role in the isomerization of disulfide bonds by maintaining the mammalian oxidoreductases in a reduced state.
Jessop
Catherine E
CE
Faculty of Life Sciences, The Michael Smith Building, University of Manchester, Manchester M13 9PT, United Kingdom.
Bulleid
Neil J
NJ
D000818:Animals; D019921:Biotinylation; D003600:Cytosol / Q000378:metabolism; D004220:Disulfides; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D005354:Fibrosarcoma / Q000378:metabolism; D005978:Glutathione / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D006023:Glycoproteins / Q000737:chemistry; D006360:Heat-Shock Proteins / Q000502:physiology; D006801:Humans; D007535:Isomerases / Q000502:physiology; D008861:Microsomes / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010088:Oxidoreductases / Q000378:metabolism*; D010100:Oxygen / Q000737:chemistry; D019704:Protein Disulfide-Isomerases; D013438:Sulfhydryl Compounds; D013997:Time Factors
D004220:Disulfides; D006023:Glycoproteins; D006360:Heat-Shock Proteins; D013438:Sulfhydryl Compounds; D010088:Oxidoreductases; D007535:Isomerases; D019704:Protein Disulfide-Isomerases; C085868:PDIA3 protein, human; D005978:Glutathione; D010100:Oxygen