Effective renaturation of reduced lysozyme by gentle removal of urea |
| |
Authors: | Maeda Yoshitake; Koga Hiroshi; Yamada Hidenori; Ueda Tadashi; Imoto Taiji |
| |
Affiliation: | 1Graduate School of Pharmaceutical Sciences, Kyushu University 62 Maidashi, Higashi-ku, Fukuoka 812
2Department of Bioengineering Science, Faculty of Engineering Tsushima Naka, Okayama 700, Japan |
| |
Abstract: | To increase the folding yield of concentrated reduced lysozyme,we developed a renaturation method by means of dialysis fromconcentrated urea with redox agents. After lysozyme was incubatedin the reducing buffer (8 M urea solution) with oxidized glutathione,renaturation of reduced lysozyme was started by dialysis againstthe dialyzing buffer containing 8 M urea with redox agents.The urea concentration of the dialyzing bottle was graduallydiluted with dialyzing buffer without urea at a flow rate of0.1 ml/min by high pressure pump. Using this systematic dialysis,a concentration as high as 5 mg/ml of reduced lysozyme couldbe renaturated in 80% yield, while the folding yield was <5%even at a concentration of 1 mg/ml using a conventional rapiddilution method Goldberg et al. (1991) Biochemistry, 30, 27902797].Therefore, it was concluded that gentle removal of urea fromdenatured proteins, dissolved in concentrated urea solution,by means of dialysis should be useful to renature denaturedproteins effectively. |
| |
Keywords: | aggregation/ dialysis/ effective renaturation/ lysozyme/ urea |
本文献已被 Oxford 等数据库收录! |
|