Immobilized oxidoreductase as an additive for refolding inclusion bodies: Application to antibody fragments

Kouhei Tsumoto; Mitsuo Umetsu; Hidenari Yamada; Takahiko Ito; Satoru Misawa; Izumi Kumagai

doi:10.1093/protein/gzg064

Immobilized oxidoreductase as an additive for refolding inclusion bodies: Application to antibody fragments

Kouhei Tsumoto, Mitsuo Umetsu, Hidenari Yamada, Takahiko Ito, Satoru Misawa, Izumi Kumagai

ENG - Biomolecular Engineering

Research output: Contribution to journal › Article › peer-review

21 Citations (Scopus)

Abstract

Three foldases - the apical domain of GroEL (mini-chaperone) and two oxidoreductases (DsbA and DsbC) from Escherichia coli - were studied in refolding a protein with immunoglobulin fold (immunoglobulin-folded protein) that had been produced as inclusion bodies in E.coli. The foldases promoted the refolding of single-chain antibody fragments from denaturant-solubilized and reduced inclusion bodies in vitro, and also effectively functioned as alternatives for labilizing agent and oxidizing reagent in the stepwise dialysis system. Immobilization of the oxidoreductases enhanced refolding and recovery of functional single-chain antibody in the dialysis system, suggesting that immobilized oxidoreductases can be used as an effective additive for refolding immunoglobulin-folded proteins in vitro.

Original language	English
Pages (from-to)	535-541
Number of pages	7
Journal	Protein Engineering
Volume	16
Issue number	7
DOIs	https://doi.org/10.1093/protein/gzg064
Publication status	Published - 2003 Jul 1

Keywords

Foldase
Immobilization
Immunoglobulin fold
Oxidoreductase
Refolding

ASJC Scopus subject areas

Biochemistry
Molecular Biology

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10.1093/protein/gzg064

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title = "Immobilized oxidoreductase as an additive for refolding inclusion bodies: Application to antibody fragments",

abstract = "Three foldases - the apical domain of GroEL (mini-chaperone) and two oxidoreductases (DsbA and DsbC) from Escherichia coli - were studied in refolding a protein with immunoglobulin fold (immunoglobulin-folded protein) that had been produced as inclusion bodies in E.coli. The foldases promoted the refolding of single-chain antibody fragments from denaturant-solubilized and reduced inclusion bodies in vitro, and also effectively functioned as alternatives for labilizing agent and oxidizing reagent in the stepwise dialysis system. Immobilization of the oxidoreductases enhanced refolding and recovery of functional single-chain antibody in the dialysis system, suggesting that immobilized oxidoreductases can be used as an effective additive for refolding immunoglobulin-folded proteins in vitro.",

keywords = "Foldase, Immobilization, Immunoglobulin fold, Oxidoreductase, Refolding",

author = "Kouhei Tsumoto and Mitsuo Umetsu and Hidenari Yamada and Takahiko Ito and Satoru Misawa and Izumi Kumagai",

note = "Funding Information: This work was supported by the Proposal-Based R&D Promotion Program (I.K.) and the Industrial Technology Research Grant Program in 2000 (K.T.) from the New Energy and Industrial Technology Development Organization (NEDO) of Japan, and by Grant-in-Aids for General Research (I.K.) and for Younger Scientists (K.T.) from the Japan Society for the Promotion of Science. This research was also partially supported by the Ministry of Education, Science, Sports and Culture, a Grant-in-Aid for the COE project, Giant Molecules and Complex Systems, 2002.",

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doi = "10.1093/protein/gzg064",

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AU - Tsumoto, Kouhei

AU - Umetsu, Mitsuo

AU - Yamada, Hidenari

AU - Ito, Takahiko

AU - Misawa, Satoru

AU - Kumagai, Izumi

N1 - Funding Information: This work was supported by the Proposal-Based R&D Promotion Program (I.K.) and the Industrial Technology Research Grant Program in 2000 (K.T.) from the New Energy and Industrial Technology Development Organization (NEDO) of Japan, and by Grant-in-Aids for General Research (I.K.) and for Younger Scientists (K.T.) from the Japan Society for the Promotion of Science. This research was also partially supported by the Ministry of Education, Science, Sports and Culture, a Grant-in-Aid for the COE project, Giant Molecules and Complex Systems, 2002.

PY - 2003/7/1

Y1 - 2003/7/1

N2 - Three foldases - the apical domain of GroEL (mini-chaperone) and two oxidoreductases (DsbA and DsbC) from Escherichia coli - were studied in refolding a protein with immunoglobulin fold (immunoglobulin-folded protein) that had been produced as inclusion bodies in E.coli. The foldases promoted the refolding of single-chain antibody fragments from denaturant-solubilized and reduced inclusion bodies in vitro, and also effectively functioned as alternatives for labilizing agent and oxidizing reagent in the stepwise dialysis system. Immobilization of the oxidoreductases enhanced refolding and recovery of functional single-chain antibody in the dialysis system, suggesting that immobilized oxidoreductases can be used as an effective additive for refolding immunoglobulin-folded proteins in vitro.

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KW - Immunoglobulin fold

KW - Oxidoreductase

KW - Refolding

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Immobilized oxidoreductase as an additive for refolding inclusion bodies: Application to antibody fragments

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Keywords

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