Ero1p Oxidizes Protein Disulfide Isomerase in a Pathway for Disulfide Bond Formation in the Endoplasmic Reticulum

ArticleinMolecular Cell 4(4):469-77 · November 1999with12 Reads
DOI: 10.1016/S1097-2765(00)80198-7 · Source: PubMed
Native protein disulfide bond formation in the endoplasmic reticulum (ER) requires protein disulfide isomerase (PDI) and Ero1p. Here we show that oxidizing equivalents flow from Ero1p to substrate proteins via PDI. PDI is predominantly oxidized in wild-type cells but is reduced in an ero1-1 mutant. Direct dithiol-disulfide exchange between PDI and Ero1p is indicated by the capture of PDI-Ero1p mixed disulfides. Mixed disulfides can also be detected between PDI and the ER precursor of carboxypeptidase Y (CPY). Further, PDI1 is required for the net formation of disulfide bonds in newly synthesized CPY, indicating that PDI functions as an oxidase in vivo. Together, these results define a pathway for protein disulfide bond formation in the ER. The PDI homolog Mpd2p is also oxidized by Ero1p.
    • "The ER is a consumer of glutathione, which is imported from the cytosol and utilized in disulfide bond formation during protein folding (Chakravarthi et al., 2006). PDI, which transfers oxidative equivalents to newly synthesized proteins, is one of the central proteins involved in oxidative protein folding (Frand and Kaiser, 1999). Thus, we evaluated whether formation of high-molecular weight complexes was enhanced in CCl 4 -treated samples by analyzing complexes formed by PDI on non-reducing gels. "
    [Show abstract] [Hide abstract] ABSTRACT: Endoplasmic reticulum (ER) stress and associated protein aggregation are closely associated with human diseases, including alterations in hepatic lipid metabolism. Inhibition of ER stress can have a significant effect on the prevention of hepatic dyslipidemia. Here, we studied the role of 4-phenylbutyric acid (4-PBA), a chemical chaperone, on ER stress-induced hepatic lipid accumulation. We studied ER stress induction following CCl4 exposure and delineated mechanisms of the CCl4-induced ER stress response in liver tissue from mice. CCl4 affected the formation of disulfide bonds through excessive hyper-oxidation of protein disulfide isomerase (PDI). Increased complex formation between PDI and its client proteins persisted in CCl4-exposed samples. Conversely, 4-PBA inhibited ER stress via secretion of apolipoprotein B and prevention of hepatic lipid accumulation. We also studied the mechanism-based pharmacokinetic and pharmacodynamic profiles and identified the ER stress-related proteins GRP78 and CHOP, along with plasma apolipoprotein B and triglyceride levels, as novel biomarkers of ER stress-induced hepatic lipid accumulation. ER stress and its clinical relevance for therapeutic approaches were well correlated with the activity of the ER stress regulator 4-PBA, which may be a promising drug candidate for the treatment of hepatic lipid accumulation, such as hepatic steatosis.
    Full-text · Article · Mar 2016
    • "The bEro1 expression also markedly increased in the posterior silk grand of 5th instar larvae as well as bPdi. Increasing evidence suggests that ERO1 directly oxidizes PDI, which participates in disulfide bond formation for newly synthesized proteins [3,6,26]. Therefore, cooperative activity and expression of bPDI and bERO1 play important roles in the formation of disulfide linkages among fibroin heavy and light chains in B. mori. "
    [Show abstract] [Hide abstract] ABSTRACT: We isolated a complementary DNA (cDNA) clone encoding endoplasmic reticulum oxidoreductin 1 (bERO1, a specific oxidant of protein disulfide isomerase (PDI)) from Bombyx mori. This protein has a putative open reading frame (ORF) of 489 amino acids and a predicted size of 57.4 kDa. Although bERO1 protein shares less than 57% amino acid sequence homology with other reported ERO1s, it contains two conserved redox active motifs, a Cys-X-X-X-X-Cys motif of N-terminal and Cys-X-X-Cys-X-X-Cys motif of C-terminal. Both motifs are typically present in ERO1 protein family members. The bEro1 mRNA expression was highest in posterior silk gland on the sixth day of the 5th instar larvae. Expression of bEro1 mRNA also markedly increased during endoplasmic reticulum (ER) stress induced by stimulation with antimycin, calcium ionophore A23187, dithiothreitol, H₂O₂, monencin, and tunicamycin. In addition, expression levels of bEro1 exactly coincided with that of bPdi. This is the first result suggesting that bERO1 plays an essential role in ER quality control through the combined activities of bERO1 and bPDI as a catalyst of protein folding in the ER and sustaining cellular redox homeostasis.
    Full-text · Article · Nov 2015
    • "2.9. Determination of redox state of TrPDI2 QM9414 mycelium was collected after Avicel induction for the indicated time and subjected to protein extraction and redox state analysis of TrPDI2 with 4-acetamido-4 -maleimidylstilbene- 2,2 –disulfonic acid (AMS, Molecular Probes) as the thiol-active reagent using western blotting, as previously described [7]. "
    [Show abstract] [Hide abstract] ABSTRACT: The majority of the cysteine residues in the secreted proteins form disulfide bonds via protein disulfide isomerase (PDI)-mediated catalysis, stabilizing the enzyme activity. The role of PDI in cellulase production is speculative, as well as the possibility of PDI as a target for improving enzyme production efficiency of Trichoderma reesei, a widely used producer of enzyme for the production of lignocellulose-based biofuels and biochemicals. Here, we report that a PDI homolog, TrPDI2 in T. reesei exhibited a 36.94% and an 11.81% similarity to Aspergillus niger TIGA and T. reesei PDI1, respectively. The capability of TrPDI2 to recover the activity of reduced and denatured RNase by promoting refolding verified its protein disulfide isomerase activity. The overexpression of Trpdi2 increased the secretion and the activity of CBH1 at the early stage of cellulase induction. In addition, both the expression level and redox state of TrPDI2 responded to cellulase induction in T. reesei, providing sustainable oxidative power to ensure cellobiohydrolase maturation and production. The results suggest that TrPDI2 may contribute to cellobiohydrolase secretion by enhancing the capability of disulfide bond formation, which is essential for protein folding and maturation. Copyright © 2015 Elsevier Inc. All rights reserved.
    Full-text · Article · May 2015
    Guokun WangGuokun WangPin LvPin LvRonglin HeRonglin He+1more author...[...]
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