Crystal Structure of the ERp44-Peroxiredoxin 4 Complex Reveals the Molecular Mechanisms of Thiol-Mediated Protein Retention

Yang, Kun; Li, De Feng; Xie, Wang; Liang, Jinsong; Sitia, Roberto; Wang, Chih Chen; Wang, Xi

doi:10.1016/j.str.2016.08.002

Cited by 42 publications

(37 citation statements)

References 46 publications

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“…During the evaluation process of this paper, Yang et al reported the crystal structure of the ERp44-Prx4 complex (32), which overall supports our proposed model of ERp44 binding to the client proteins. The observation that ERp44 prefers the oxidized form of Prx4 to its reduced form (32) can be explained by the different electrostatic surface properties between these two redox states.…”

Section: Discussionsupporting

confidence: 80%

“…The crystal structure of the ERp44-Prx4 complex (32) reveals that a Cys29 (ERp44)-Cys208 (Prx4) intermolecular disulfide bond and a number of hydrogen bonds form at the interface between the ERp44 a domain and the Prx4 homodimer. In this context, the flexible C-terminal region of Prx4 assumes a β-strand, which forms hydrogen bonds with the β4-strand of ERp44 (32). Taken together, these observations suggest the following general mechanism of ERp44-client complex formation.…”

Section: Discussionmentioning

confidence: 99%

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Structural basis of pH-dependent client binding by ERp44, a key regulator of protein secretion at the ER–Golgi interface

Watanabe

Harayama

Kanemura

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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ERp44 retrieves some endoplasmic reticulum (ER)-resident enzymes and immature oligomers of secretory proteins from the Golgi. Association of ERp44 with its clients is regulated by pH-dependent mechanisms, but the molecular details are not fully understood. Here we report high-resolution crystal structures of human ERp44 at neutral and weakly acidic pH. These structures reveal key regions in the C-terminal tail (C tail) missing in the original crystal structure, including a regulatory histidine-rich region and a subsequent extended loop. The former region forms a short α-helix (α16), generating a histidine-clustered site (His cluster). At low pH, the three Trx-like domains of ERp44 ("a," "b," and "b'") undergo significant rearrangements, likely induced by protonation of His157 located at the interface between the a and b domains. The α16-helix is partially unwound and the extended loop is disordered in weakly acidic conditions, probably due to electrostatic repulsion between the protonated histidines in the His cluster. Molecular dynamics simulations indicated that helix unwinding enhances the flexibility of the C tail, disrupting its normal hydrogen-bonding pattern. The observed pH-dependent conformational changes significantly enlarge the positively charged regions around the client-binding site of ERp44 at low pH. Mutational analyses showed that ERp44 forms mixed disulfides with specific cysteines residing on negatively charged loop regions of Ero1α. We propose that the protonation states of the essential histidines regulate the ERp44-client interaction by altering the C-tail dynamics and surface electrostatic potential of ERp44.

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Section: Discussionsupporting

confidence: 80%

Section: Discussionmentioning

confidence: 99%

Structural basis of pH-dependent client binding by ERp44, a key regulator of protein secretion at the ER–Golgi interface

Watanabe

Harayama

Kanemura

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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“…The mutation of Ero1α S145E was created using the Fast Mutagenesis System (TransGen). The expression of pcDNA3.1‐Ero1α‐myc, pcDNA3.1‐Ero1α‐HA, and/or pcDNA3.1‐HA‐ERp44 in mammalian cells was as previously described (Wang et al , ; Yang et al , ). pcDNA3.1‐Ero1α‐HA‐KDEL was generated by insertion of a KDEL motif between HA and the termination codon.…”

Section: Methodsmentioning

confidence: 99%

Secretory kinase Fam20C tunes endoplasmic reticulum redox state via phosphorylation of Ero1α

et al. 2018

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Family with sequence similarity 20C (Fam20C), the physiological Golgi casein kinase, phosphorylates numerous secreted proteins that are involved in a wide variety of biological processes. However, the role of Fam20C in regulating proteins in the endoplasmic reticulum (ER) lumen is largely unknown. Here, we report that Fam20C interacts with various luminal proteins and that its depletion results in a more reduced ER lumen. We further show that ER oxidoreductin 1α (Ero1α), the pivotal sulfhydryl oxidase that catalyzes disulfide formation in the ER, is phosphorylated by Fam20C in the Golgi apparatus and retrograde-transported to the ER mediated by ERp44. The phosphorylation of Ser145 greatly enhances Ero1α oxidase activity and is critical for maintaining ER redox homeostasis and promoting oxidative protein folding. Notably, phosphorylation of Ero1α is induced under hypoxia, reductive stress, and secretion-demanding conditions such as mammalian lactation. Collectively, our findings open a door to uncover how oxidative protein folding is regulated by phosphorylation in the secretory pathway.

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“…The complex structure of the Am Prx homolog and Am Trx homolog has been reported, which allowed us to build a structural model of the Am Prx‐ Am Trx complex. The structure of Am Trx modeled by the SWISS‐MODEL server (https://www.swissmodel.expasy.org/) was docked to a C terminus‐modified Am Prx dimer according to the interaction in human Prx4‐Erp44 complex (PDB entry http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5HQP) . In this model, the α4‐β6 loop did not contact Am Trx, but it was located ~ 20 Å away from the Am Trx binding site (Fig.…”

Section: Resultsmentioning

confidence: 99%

Crystal structure of Akkermansia muciniphila peroxiredoxin reveals a novel regulatory mechanism of typical 2‐Cys Prxs by a distinct loop

Wang

et al. 2020

FEBS Letters

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Edited by Stuart FergusonPeroxiredoxins (Prxs) are thiol-specific antioxidant proteins commonly found in organisms that protect cells from the damage of reactive oxygen species produced by metabolism and that participate in cell signaling. The Prx from the bacterium Akkermansia muciniphila (AmPrx) is a typical 2-Cys Prx characterized by two conserved cysteines: Cys49 and Cys183. Here, we verified the peroxidase activity of AmPrx and determined its crystal structure in reduced form, which is a doughnut-shaped decamer composed of five dimers. Particularly, a distinct loop between the a4 helix and b6 strand is involved in the decameric interaction. Deletion of this loop destroys the decameric structure and significantly decreases the peroxidase activity of AmPrx. Our findings reveal a novel regulatory mechanism of typical 2-Cys Prx, in which the a4-b6 loop affects the assembly of Prx and, therefore, regulates its peroxidase activity.

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Crystal Structure of the ERp44-Peroxiredoxin 4 Complex Reveals the Molecular Mechanisms of Thiol-Mediated Protein Retention

Cited by 42 publications

References 46 publications

Structural basis of pH-dependent client binding by ERp44, a key regulator of protein secretion at the ER–Golgi interface

Structural basis of pH-dependent client binding by ERp44, a key regulator of protein secretion at the ER–Golgi interface

Secretory kinase Fam20C tunes endoplasmic reticulum redox state via phosphorylation of Ero1α

Crystal structure of Akkermansia muciniphila peroxiredoxin reveals a novel regulatory mechanism of typical 2‐Cys Prxs by a distinct loop

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