During initial production and isolation of proteins, many factors such as overproduction, solvent interactions, mechanical interference, or others may result in the denaturation of proteins.
Understanding protein folding involves understanding both the energy landscape of the protein system, and refolding techniques have been significantly improve as time-resolved techniques, including neutron scattering, have been developed and perfected by researchers around the globe. The techniques involve observation of protein dynamics in order to assess the critical point of refolding, information which can lead to the development of refolding solutions (Bu et al. 2001). Neutron scattering and similar inventive techniques, such as stopped-flow florescence.
In order to scale up for commercial use, protein refolding techniques must be scale invariant, compatible for a large range of proteins, simple to automated, and overall economical. Methods that rely on denaturant dilution and column-based methodology generally will meet these criteria (Middelberg 2002). The technology of refolding has grown exponentially in the past decade, and new methods must be carefully designed to facilitate the automated and rapid determination of the conditions that must be met for refolding in order to be commercially viable. It, however, remains to be seen if researchers can translate new technologies—and possibly even the discovery of a new protein state—into technology that will improve efficiency in bimolecular research industries.
Before use, proteins are generally solubilised before use in high concentrations of quanidinium chloride (GdmCl) and urea (De Bernardez 1998 and Schwarz et al. 1998). Either of these two solvents may cause certain proteins to denature, and refolding involves diluting to a low concentration zone.