Trends in Cell Biology
ReviewPathways for protein disulphide bond formation
Section snippets
A pathway for protein disulphide bond formation in the ER
A genetic dissection of oxidative protein folding in yeast began with the isolation of an essential and conserved gene, ERO1 (ER oxidation), encoding a novel ER membrane protein required for protein oxidation in the ER14, 15 (Table 1). A temperature-sensitive allele of ERO1 was identified in a screen for mutants defective in the export from the ER of secretory proteins containing disulphide bonds14. Mutations in ERO1 were also isolated in a screen for S. cerevisiae strains with diminished
The role of glutathione in oxidative protein folding in the ER
The thiol–disulphide redox status of intralumenal glutathione has long been the focus of considerations of how relatively oxidizing conditions are established within the ER12. Glutathione is the major small-molecule redox buffer in the ER, and the ratio of the concentration of GSH to GSSG in the ER (1:1 to 3:1) is similar to that found in redox buffers affording optimal rates for oxidative refolding in vitro12. From these, observations, it was natural to suppose that GSSG serves as the primary
Similarities in eukaryotic and prokaryotic disulphide-bond-forming pathways
The pathway for protein disulphide bond formation in the bacterial periplasm provides a useful analogy for the protein oxidation system in eukaryotes. Two enzymes drive disulphide bond formation in periplasmic proteins: the thioredoxin-like thiol–disulphide oxidoreductase DsbA and the cytoplasmic membrane protein DsbB29, 30 (Table 1). The active-site cysteines of DsbA form a disulphide bond that is transferred directly to periplasmic proteins31, after which DsbA is efficiently re-oxidized by
The family of PDI homologues
Several oxidoreductases homologous to PDI are found in the ER of both yeast and mammalian cells (Table 2). These PDI homologues have been implicated in diverse processes including not only oxidative protein folding but also the assembly of multiprotein complexes and the recognition of misfolded proteins in the ER39. Erp57 and Erp72 are mammalian homologues of PDI induced under conditions of ER stress39 (Table 2). Yeast homologues of PDI expressed in the ER lumen are Mpd1p, Mpd2p, Eug1p and Eps1p
Models for disulphide bond isomerization in eukaryotes
The similarities observed thus far between eukaryotic and prokaryotic disulphide-bond-forming systems suggest that commonalities might also be found between disulphide bond isomerization pathways in eukaryotes and the DsbC–DsbD system in prokaryotes. Eukaryotic secretory proteins typically contain more disulphide bonds than their prokaryotic counterparts, indicating that there might be an even greater need for disulphide bond reducing or reshuffling functions in the ER than in the periplasm.
Where do the oxidizing equivalents come from?
Although Ero1p appears to be the key conduit for introducing oxidizing equivalents into the ER lumen, it remains unclear how Ero1p itself is re-oxidized. Because Ero1p engages in thiol–disulphide exchange with Pdi1p, the identification of one or more redox-active cysteine pairs is anticipated in Ero1p. These active-site cysteines might be found amongst the seven conserved cysteine residues of Ero1p, three of which appear in the sequence Cys-x-x-Cys-x-x-Cys near the C-terminus of Ero1p14, 15.
Summary and future prospects
The work reviewed here places Ero1p and Pdi1p in an enzymatic pathway for protein disulphide bond formation in the ER16 and demonstrates that intralumenal GSH competes with protein thiols for oxidizing equivalents derived from Ero1p25. These studies provide a solid framework for further genetic and biochemical analysis of oxidative protein folding in eukaryotes. It will be of great interest to see whether analogies between eukaryotic and prokaryotic systems continue to hold as more details of
Acknowledgements
We are grateful to Peter Chivers and Fredrik Åslund for comments on this manuscript. We apologize to those authors whose work we could not cite directly owing to space limitations. Work in the laboratory of the authors was supported by grants from the National Institute of General Medical Sciences (to C.A.K.), an NIH predoctoral traineeship (to A.R.F.) and an NIH National Research Service Award (to J.W.C.).
References (56)
Acceleration of reactivation of reduced bovine pancreatic ribonuclease by a microsomal system from rat liver
J. Biol. Chem.
(1963)- et al.
The essential function of yeast protein disulfide isomerase does not reside in its isomerase activity
Cell
(1993) - et al.
The ERO1 gene of yeast is required for oxidation of protein dithiols in the endoplasmic reticulum
Mol. Cell
(1998) Ero1p: a novel and ubiquitous protein with an essential role in oxidative protein folding in the endoplasmic reticulum
Mol. Cell
(1998)- et al.
Ero1p oxidizes protein disulfide isomerase in a pathway for disulfide bond formation in the endoplasmic reticulum
Mol. Cell
(1999) The essential function of protein-disulfide isomerase is to unscramble non-native disulfide bonds
J. Biol. Chem.
(1995)Isolation and characterization of a yeast gene, MPD1, the overexpression of which suppresses inviability caused by protein disulfide isomerase depletion
FEBS Lett.
(1995)Overproduction of Mpd2p suppresses the lethality of protein disulfide isomerase depletion in a CXXC sequence dependent manner
Biochem. Biophys. Res. Commun.
(1997)Preferential transport of glutathione versus glutathione disulfide in rat liver microsomal vesicles
J. Biol. Chem.
(1999)Kinetic analysis of the mechanism and specificity of protein-disulfide isomerase using fluorescence-quenched peptides
J. Biol. Chem.
(1998)
Electron avenue: pathways of disulfide bond formation and isomerization
Cell
Identification of a protein required for disulfide bond formation in vivo
Cell
Oxidative protein folding is driven by the electron transport system
Cell
DsbA–DsbB interaction through their active site cysteines. Evidence from an odd cysteine mutant of DsbA
J. Biol. Chem.
Human protein disulfide isomerase functionally complements a dsbA mutation and enhances the yield of pectate lyase C in Escherichia coli
J. Biol. Chem.
Enhanced catalysis of ribonuclease B folding by the interaction of calnexin or calreticulin with ERp57
J. Biol. Chem.
The unfolded protein response: an intracellular signalling pathway with many surprising features
Trends Cell Biol.
ERp72, an abundant luminal endoplasmic reticulum protein, contains three copies of the active site sequences of protein disulfide isomerase
J. Biol. Chem.
A set of endoplasmic reticulum proteins possessing properties of molecular chaperones includes Ca(2+)-binding proteins and members of the thioredoxin superfamily
J. Biol. Chem.
Two resident ER-proteins, CaBP1 and CaBP2, with thioredoxin domains, are substrates for thioredoxin reductase: comparison with protein disulfide isomerase
FEBS Lett.
Molecular cloning of the cDNA encoding a novel protein disulfide isomerase-related protein (PDIR)
FEBS Lett.
Principles that govern the folding of protein chains
Science
Catalysis of the oxidative folding of ribonuclease A by protein disulfide isomerase: dependence of the rate on the composition of the redox buffer
Biochemistry
Formation of three-dimensional structure in proteins. I. Rapid nonenzymic reactivation of reduced lysozyme
Biochemistry
Catalysis of the oxidative folding of ribonuclease A by protein disulfide isomerase: pre-steady-state kinetics and the utilization of the oxidizing equivalents of the isomerase
Biochemistry
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