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Masaki Okumura1,2, Masatoshi Saiki2,3, Hiroshi Yamaguchi1 and Yuji Hidaka2 1 School of Science and Technology, Kwansei Gakuin University, Hyogo, Japan 2 Graduate School of Science and Engineering, Kinki University, Osaka, Japan 3 Department of Applied Chemistry, Faculty of Engineering, Tokyo University of Science, Yamaguchi, Japan Keywords arginine; disulfide; folding; glutathione; uroguanylin Correspondence Y. Hidaka, Graduate School of Science and Engineering, Kinki University, 3-4-1 Kowakae, Higashi-Osaka, Osaka 577-8502, Japan Fax: +81 6 6723 2721 Tel: +81 6 6721 2332 E-mail: yuji@life.kindai.ac.jp (Received 20 October 2010, revised 18 January 2011, accepted 28 January 2011) doi:10.1111/j.1742-4658.2011.08039.x Protein folding occurs simultaneously with disulfide bond formation. In general,. | IFEBS Journal Acceleration of disulfide-coupled protein folding using glutathione derivatives Masaki Okumura1 2 Masatoshi Saiki2 3 Hiroshi Yamaguchi1 and Yuji Hidaka2 1 Schoolof Science and Technology Kwansei Gakuin University Hyogo Japan 2 Graduate Schoolof Science and Engineering Kinki University Osaka Japan 3 Department of Applied Chemistry Faculty of Engineering Tokyo University of Science Yamaguchi Japan Keywords arginine disulfide folding glutathione uroguanylin Correspondence Y. Hidaka Graduate Schoolof Science and Engineering Kinki University 3-4-1 Kowakae Higashi-Osaka Osaka 577-8502 Japan Fax 81 6 6723 2721 Tel 81 6 6721 2332 E-mail yuji@life.kindai.ac.jp Received 20 October 2010 revised 18 January 2011 accepted 28 January 2011 doi 10.1111 j.1742-4658.2011.08039.x Protein folding occurs simultaneously with disulfide bond formation. In general the in vitro folding proteins containing disulfide bond s is carried out in the presence of redox reagents such as glutathione to permit native disulfide pairing to occur. It is well known that the formation of a disulfide bond and the correct tertiary structure of a target protein are strongly affected by the redox reagent used. However little is known concerning the role of each amino acid residue of the redox reagent such as glutathione. Therefore we prepared glutathione derivatives - glutamyl-cysteinyl-argi-nine ECR and arginyl-cysteinyl-glycine RCG - and examined their ability to facilitate protein folding using lysozyme and prouroguanylin as model proteins. When the reduced and oxidized forms of RCG were used folding recovery was greater than that for a typical glutathione redox system. This was particularly true when high protein concentrations were employed whereas folding recovery using ECR was similar to that of the glutathione redox system. Kinetic analyses of the oxidative folding of prou-roguanylin revealed that the folding velocity KRCG 3.69 X 10-3 s-1 using reduced RCG oxidized RCG was approximately .
