title: Protein refolding is improved by adding nonionic polyethylene glycol monooleyl ethers with various polyethylene glycol lengths.
abstract: Protein refolding from bacterial inclusion bodies is a crucial step for the production of recombinant proteins, but the refolding step often results in significantly lower yields due to aggregation. To prevent aggregation, chemical additives are often used. However, the ability of additives to effectively increase refolding yields are protein dependent, and therefore, it is important to understand the manner in which the substructures of additives confer suitable properties on protein refolding. We focused attention on nonionic detergents, the polyethylene glycol monooleyl ether (PGME) series, and systematically studied the influence of two to 90 polyethylene glycol (PEG) lengths of PGMEs on the refolding of pig muscle lactate dehydrogenase (LDH), hen egg white lysozyme, and yeast α-glucosidase. PGMEs with longer PEG lengths such as PGME20, 50, and 90 suppressed aggregation, and increased refolding yields. Notably, PGME20 increased the LDH yield to 56.7% from 2.5% without additives. According to the refolding kinetic analysis of LDH, compared with PGME50 and 90, the refolding rate constant in PGME20 solutions remained relatively high at a broad range of concentrations because of its weaker steric hindrance of intramolecular interactions involved in folding, leading to a preference for refolding over aggregation. These findings should provide basic guidelines to identify appropriate PEG-based nonionic detergents for protein refolding.
authors.affiliation: Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, Tokyo, Japan.
authors.forename: Etsushi
authors.initials: E
authors.lastname: Yamamoto
authors.affiliation: Research Center for Advanced Science and Technology (RCAST), The University of Tokyo, Tokyo, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Yamaguchi
authors.affiliation: Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, Tokyo, Japan.|Department of Bioengineering, School of Engineering, The University of Tokyo, Tokyo, Japan.
authors.forename: Teruyuki
authors.initials: T
authors.lastname: Nagamune
mesh_terms: D000818:Animals; D004987:Ethers / Q000737:chemistry; D005656:Fungal Proteins; D002479:Inclusion Bodies / Q000737:chemistry; D007770:L-Lactate Dehydrogenase; D009113:Muramidase; D011092:Polyethylene Glycols / Q000737:chemistry*; D058849:Protein Refolding; D011994:Recombinant Proteins / Q000737:chemistry* / Q000378:metabolism*; D013552:Swine; D000520:alpha-Glucosidases
chemical_list: D004987:Ethers; D005656:Fungal Proteins; D011994:Recombinant Proteins; D011092:Polyethylene Glycols; D007770:L-Lactate Dehydrogenase; C421864:hen egg lysozyme; D009113:Muramidase; D000520:alpha-Glucosidases
keywords: Aggregation inhibitors; Nonionic detergents; PEG-lipid; Protein refolding; Refolding additives