Refolding of denatured/reduced lysozyme by aromatic thiols in the absence of small molecule disulfide

Abstract

To increase the folding rate and gain insight into the folding process, the lysozyme was selected as a model protein and refolded by the aromatic thiols (ArSH) alone and by redox buffers glutathione/glutathione disulfide (GSH/GSSG). The optimum folding conditions, including the concentration of aromatic thiols, and the composition and pH of buffer solutions, were investigated. Attempts were first made to understand how aromatic thiols alone affect the folding performance of disulfide-containing proteins. The results showed that redox buffers of both aliphatic thiols and aromatic thiols can enhance the folding rate and yield of denatured reduced proteins effectively. However, the folding rates by aromatic thiols were up to 2–15 times greater than those with GSH/GSSG under various pH values (6.0–9.0). More importantly, the research demonstrated that the aromatic thiols alone can also increase the rate of proteins folding significantly at wide pH values, which has never been previously reported. It is expected that the research will provide a novel method for the folding of disulfide-containing proteins.