Oxidative refolding of lysozyme in trifluoroethanol (TFE) and ethylene glycol: interfering role of preexisting α-helical structure and intermolecular hydrophobic interactions

Abstract

The oxidative refolding of equilibrium intermediates of lysozyme stabilized in trifluoroethanol (TFE) and ethylene glycol was monitored. Equilibrium intermediates of disulfide reduced lysozyme in TFE are known to contain considerable amounts of α-helical structure and resemble the early intermediate in the oxidative refolding of lysozyme. We find that the intermediates in TFE do not proceed to folding; they form aggregates. However, interestingly, intermediates in ethylene glycol refold to the native state with improved folding yield. Secondary structure of these intermediates was monitored by far-UV circular dichroism. Our results indicate that formation of α-helical structure prior to oxidative refolding does not help the process in the case of lysozyme. Interfering with intermolecular hydrophobic interactions in the unfolded state is more productive.