Redox-dependent domain rearrangement of protein disulfide isomerase coupled with exposure of its substrate-binding hydrophobic surface

Olivier Serve, Yukiko Kamiya, Aya Maeno, Michiko Nakano, Chiho Murakami, Hiroaki Sasakawa, Yoshiki Yamaguchi, Takushi Harada, Eiji Kurimoto, Maho Yagi-Utsumi, Takeshi Iguchi, Kenji Inaba, Jun Kikuchi, Osamu Asami, Tsutomu Kajino, Toshihiko Oka, Masayoshi Nakasako, Koichi Kato

Research output: Contribution to journalArticle

47 Citations (Scopus)

Abstract

Protein disulfide isomerase (PDI) is a major protein in the endoplasmic reticulum, operating as an essential folding catalyst and molecular chaperone for disulfide-containing proteins by catalyzing the formation, rearrangement, and breakage of their disulfide bridges. This enzyme has a modular structure with four thioredoxin-like domains, a, b, b′, and a′, along with a C-terminal extension. The homologous a and a′ domains contain one cysteine pair in their active site directly involved in thiol-disulfide exchange reactions, while the b′ domain putatively provides a primary binding site for unstructured regions of the substrate polypeptides. Here, we report a redox-dependent intramolecular rearrangement of the b′ and a′ domains of PDI from Humicola insolens, a thermophilic fungus, elucidated by combined use of nuclear magnetic resonance (NMR) and small-angle X-ray scattering (SAXS) methods. Our NMR data showed that the substrates bound to a hydrophobic surface spanning these two domains, which became more exposed to the solvent upon oxidation of the active site of the a′ domain. The hydrogen-deuterium exchange and relaxation data indicated that the redox state of the a′ domain influences the dynamic properties of the b′ domain. Moreover, the SAXS profiles revealed that oxidation of the a′ active site causes segregation of the two domains. On the basis of these data, we propose a mechanistic model of PDI action; the a′ domain transfers its own disulfide bond into the unfolded protein accommodated on the hydrophobic surface of the substrate-binding region, which consequently changes into a "closed" form releasing the oxidized substrate.

Original languageEnglish
Pages (from-to)361-374
Number of pages14
JournalJournal of Molecular Biology
Volume396
Issue number2
DOIs
Publication statusPublished - 2010 Feb

Fingerprint

Protein Disulfide-Isomerases
Disulfides
Oxidation-Reduction
Catalytic Domain
Magnetic Resonance Spectroscopy
X-Rays
Protein Unfolding
Thioredoxins
Molecular Chaperones
Deuterium
Sulfhydryl Compounds
Endoplasmic Reticulum
Cysteine
Hydrogen
Proteins
Fungi
Binding Sites
Peptides
Enzymes

Keywords

  • Domain rearrangement
  • Molecular chaperone
  • NMR
  • Protein disulfide isomerase
  • SAXS

ASJC Scopus subject areas

  • Molecular Biology

Cite this

Redox-dependent domain rearrangement of protein disulfide isomerase coupled with exposure of its substrate-binding hydrophobic surface. / Serve, Olivier; Kamiya, Yukiko; Maeno, Aya; Nakano, Michiko; Murakami, Chiho; Sasakawa, Hiroaki; Yamaguchi, Yoshiki; Harada, Takushi; Kurimoto, Eiji; Yagi-Utsumi, Maho; Iguchi, Takeshi; Inaba, Kenji; Kikuchi, Jun; Asami, Osamu; Kajino, Tsutomu; Oka, Toshihiko; Nakasako, Masayoshi; Kato, Koichi.

In: Journal of Molecular Biology, Vol. 396, No. 2, 02.2010, p. 361-374.

Research output: Contribution to journalArticle

Serve, O, Kamiya, Y, Maeno, A, Nakano, M, Murakami, C, Sasakawa, H, Yamaguchi, Y, Harada, T, Kurimoto, E, Yagi-Utsumi, M, Iguchi, T, Inaba, K, Kikuchi, J, Asami, O, Kajino, T, Oka, T, Nakasako, M & Kato, K 2010, 'Redox-dependent domain rearrangement of protein disulfide isomerase coupled with exposure of its substrate-binding hydrophobic surface', Journal of Molecular Biology, vol. 396, no. 2, pp. 361-374. https://doi.org/10.1016/j.jmb.2009.11.049
Serve, Olivier ; Kamiya, Yukiko ; Maeno, Aya ; Nakano, Michiko ; Murakami, Chiho ; Sasakawa, Hiroaki ; Yamaguchi, Yoshiki ; Harada, Takushi ; Kurimoto, Eiji ; Yagi-Utsumi, Maho ; Iguchi, Takeshi ; Inaba, Kenji ; Kikuchi, Jun ; Asami, Osamu ; Kajino, Tsutomu ; Oka, Toshihiko ; Nakasako, Masayoshi ; Kato, Koichi. / Redox-dependent domain rearrangement of protein disulfide isomerase coupled with exposure of its substrate-binding hydrophobic surface. In: Journal of Molecular Biology. 2010 ; Vol. 396, No. 2. pp. 361-374.
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AU - Nakano, Michiko

AU - Murakami, Chiho

AU - Sasakawa, Hiroaki

AU - Yamaguchi, Yoshiki

AU - Harada, Takushi

AU - Kurimoto, Eiji

AU - Yagi-Utsumi, Maho

AU - Iguchi, Takeshi

AU - Inaba, Kenji

AU - Kikuchi, Jun

AU - Asami, Osamu

AU - Kajino, Tsutomu

AU - Oka, Toshihiko

AU - Nakasako, Masayoshi

AU - Kato, Koichi

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N2 - Protein disulfide isomerase (PDI) is a major protein in the endoplasmic reticulum, operating as an essential folding catalyst and molecular chaperone for disulfide-containing proteins by catalyzing the formation, rearrangement, and breakage of their disulfide bridges. This enzyme has a modular structure with four thioredoxin-like domains, a, b, b′, and a′, along with a C-terminal extension. The homologous a and a′ domains contain one cysteine pair in their active site directly involved in thiol-disulfide exchange reactions, while the b′ domain putatively provides a primary binding site for unstructured regions of the substrate polypeptides. Here, we report a redox-dependent intramolecular rearrangement of the b′ and a′ domains of PDI from Humicola insolens, a thermophilic fungus, elucidated by combined use of nuclear magnetic resonance (NMR) and small-angle X-ray scattering (SAXS) methods. Our NMR data showed that the substrates bound to a hydrophobic surface spanning these two domains, which became more exposed to the solvent upon oxidation of the active site of the a′ domain. The hydrogen-deuterium exchange and relaxation data indicated that the redox state of the a′ domain influences the dynamic properties of the b′ domain. Moreover, the SAXS profiles revealed that oxidation of the a′ active site causes segregation of the two domains. On the basis of these data, we propose a mechanistic model of PDI action; the a′ domain transfers its own disulfide bond into the unfolded protein accommodated on the hydrophobic surface of the substrate-binding region, which consequently changes into a "closed" form releasing the oxidized substrate.

AB - Protein disulfide isomerase (PDI) is a major protein in the endoplasmic reticulum, operating as an essential folding catalyst and molecular chaperone for disulfide-containing proteins by catalyzing the formation, rearrangement, and breakage of their disulfide bridges. This enzyme has a modular structure with four thioredoxin-like domains, a, b, b′, and a′, along with a C-terminal extension. The homologous a and a′ domains contain one cysteine pair in their active site directly involved in thiol-disulfide exchange reactions, while the b′ domain putatively provides a primary binding site for unstructured regions of the substrate polypeptides. Here, we report a redox-dependent intramolecular rearrangement of the b′ and a′ domains of PDI from Humicola insolens, a thermophilic fungus, elucidated by combined use of nuclear magnetic resonance (NMR) and small-angle X-ray scattering (SAXS) methods. Our NMR data showed that the substrates bound to a hydrophobic surface spanning these two domains, which became more exposed to the solvent upon oxidation of the active site of the a′ domain. The hydrogen-deuterium exchange and relaxation data indicated that the redox state of the a′ domain influences the dynamic properties of the b′ domain. Moreover, the SAXS profiles revealed that oxidation of the a′ active site causes segregation of the two domains. On the basis of these data, we propose a mechanistic model of PDI action; the a′ domain transfers its own disulfide bond into the unfolded protein accommodated on the hydrophobic surface of the substrate-binding region, which consequently changes into a "closed" form releasing the oxidized substrate.

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KW - Molecular chaperone

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KW - SAXS

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