title: Correct protein folding in glycerol.
abstract: Water is the natural medium for protein folding, which is also used in all in vitro studies. In the present work, we posed, and answered affirmatively, a question of whether it is possible to fold correctly a typical protein in a nonaqueous solvent. To this end, unfolded and reduced hen egg-white lysozyme was refolded and reoxidized in glycerol containing varying amounts of water. The unfolded/reduced enzyme was found to regain spontaneously substantial catalytic activity even in the nearly anhydrous solvent; for example, the refolding yield in 99% glycerol was still some one-third of that in pure water, and one-half of that was regained even in 99.8% glycerol. The less than full recovery of the enzymatic activity in glycerol is, as in water, because of competing protein aggregation during the refolding. Lysozyme reoxidation in glycerol was successfully mediated by two dissimilar oxidizing systems, and the refolding yield was markedly affected by the pH of the last aqueous solution before the transfer into glycerol. No recovery of the lysozyme activity was observed when the refolding/reoxidation reaction was carried out in the denaturing solvent dimethyl sulfoxide. This study paves the way for a systematic investigation of the solvent effect on protein folding and demonstrates that water is not a unique milieu for this process.
authors.affiliation: Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: R V
authors.initials: RV
authors.lastname: Rariy
authors.forename: A M
authors.initials: AM
authors.lastname: Klibanov
mesh_terms: D000818:Animals; D002645:Chickens; D004121:Dimethyl Sulfoxide; D005990:Glycerol; D066298:In Vitro Techniques; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism; D010084:Oxidation-Reduction; D011489:Protein Denaturation; D017510:Protein Folding; D012997:Solvents; D014867:Water
chemical_list: D012997:Solvents; D014867:Water; D009113:Muramidase; D005990:Glycerol; D004121:Dimethyl Sulfoxide