Dataset Information

Molecular bases of cyclic and specific disulfide interchange between human ERO1alpha protein and protein-disulfide isomerase (PDI).

ABSTRACT: In the endoplasmic reticulum (ER) of human cells, ERO1? and protein-disulfide isomerase (PDI) constitute one of the major electron flow pathways that catalyze oxidative folding of secretory proteins. Specific and limited PDI oxidation by ERO1? is essential to avoid ER hyperoxidation. To investigate how ERO1? oxidizes PDI selectively among more than 20 ER-resident PDI family member proteins, we performed docking simulations and systematic biochemical analyses. Our findings reveal that a protruding ?-hairpin of ERO1? specifically interacts with the hydrophobic pocket present in the redox-inactive PDI b'-domain through the stacks between their aromatic residues, leading to preferred oxidation of the C-terminal PDI a'-domain. ERO1? associated preferentially with reduced PDI, explaining the stepwise disulfide shuttle mechanism, first from ERO1? to PDI and then from oxidized PDI to an unfolded polypeptide bound to its hydrophobic pocket. The interaction of ERO1? with ERp44, another PDI family member protein, was also analyzed. Notably, ERO1?-dependent PDI oxidation was inhibited by a hyperactive ERp44 mutant that lacks the C-terminal tail concealing the substrate-binding hydrophobic regions. The potential ability of ERp44 to inhibit ERO1? activity may suggest its physiological role in ER redox and protein homeostasis.

SUBMITTER: Masui S

PROVIDER: S-EPMC3091233 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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Molecular bases of cyclic and specific disulfide interchange between human ERO1alpha protein and protein-disulfide isomerase (PDI).

Masui Shoji S Vavassori Stefano S Fagioli Claudio C Sitia Roberto R Inaba Kenji K

The Journal of biological chemistry 20110311 18

In the endoplasmic reticulum (ER) of human cells, ERO1α and protein-disulfide isomerase (PDI) constitute one of the major electron flow pathways that catalyze oxidative folding of secretory proteins. Specific and limited PDI oxidation by ERO1α is essential to avoid ER hyperoxidation. To investigate how ERO1α oxidizes PDI selectively among more than 20 ER-resident PDI family member proteins, we performed docking simulations and systematic biochemical analyses. Our findings reveal that a protrudin ...[more]

PMID: 21398518

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Project description:Hypoxia-inducible factor-1alpha (HIF-1alpha), a transcription factor, plays a critical role in adaption to hypoxia, which is a major feature of diseases, including cancer. Protein disulfide isomerase (PDI) is up-regulated in numerous cancers and leads to cancer progression. PDI, a member of the TRX superfamily, regulates the transcriptional activities of several transcription factors. To investigate the mechanisms by which PDI affects the function of HIF-1alpha, the overexpression or knockdown of PDI was performed. The overexpression of PDI decreased HIF-1alpha expression in the human hepatocarcinoma cell line, Hep3B, whereas the knockdown of endogenous PDI increased its expression. NH4Cl inhibited the decrease in HIF-1alpha expression by PDI overexpression, suggesting that HIF-1alpha was degraded by the lysosomal pathway. HIF-1alpha is transferred to lysosomal membranes by heat shock cognate 70 kDa protein (HSC70). The knockdown of HSC70 abolished the decrease, and PDI facilitated the interaction between HIF-1alpha and HSC70. HIF-1alpha directly interacted with PDI. PDI exists not only in the endoplasmic reticulum (ER), but also in the cytosol. Hypoxia increased cytosolic PDI. We also investigated changes in the redox state of HIF-1alpha using PEG-maleimide, which binds to thiols synthesized from disulfide bonds by reduction. An up-shift in the HIF-1alpha band by the overexpression of PDI was detected, suggesting that PDI formed disulfide bond in HIF-1alpha. HIF-1alpha oxidized by PDI was not degraded in HSC70-knockdown cells, indicating that the formation of disulfide bond in HIF-1alpha was important for decreases in HIF-1alpha expression. To the best of our knowledge, this is the first study to show the regulation of the expression and redox state of HIF-1alpha by PDI. We also demonstrated that PDI formed disulfide bonds in HIF-1alpha 1-245 aa and decreased its expression. In conclusion, the present results showed that PDI is a novel factor regulating HIF-1alpha through lysosome-dependent degradation by changes in its redox state.

Dataset Information

Molecular bases of cyclic and specific disulfide interchange between human ERO1alpha protein and protein-disulfide isomerase (PDI).

Publications

Molecular bases of cyclic and specific disulfide interchange between human ERO1alpha protein and protein-disulfide isomerase (PDI).

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