Refolding yields

Merli, S., A. Corti, and G. Cassani (1995). Production of soluble tumor necrosis factor receptor type I in Escherichia coli optimization of the refolding yields by a microtiter dilution assay. Anal Biochem 230(1) 85-91. 

Refolding yields can be improved by utilizing defensive reversible denaturation". This technique has been shown to be effective in restoring the activity of denatured interferon preparations (103). It was observed that interferons denatur by a variety of techniques can be partially refolded by incubation in low concentrations of urea. When SDS was added to the urea-interferon solutions prior to renaturation, full activity of the interferons could be restored. Stewart et al. (103) interpreted the stabilizing effect of SDS as "defensively" denaturing the protein solution. 

An Engineering Approach to Achieving High-Protein Refolding Yields... 

Refolding yield optimimization. Equations (3) through (7) describe the kinetics of hCAB refolding under a given set of environmental conditions. However, when diafiltration is employed to remove the denaturant, the denaturant concentration is a function of time. For a constant volume, constant filtration rate diafiltration system, the equation describing the rate of denaturant agent removal is ... 

Cost of the Bench-scale Process. The cost of materials for the treatment of 1 liter of partially purified rSLPI, containing approximately 1 gram of the protein of interest was 20.65. At a refold yield of 50%, this represented a cost of refolding of 41.29 per active gram of rSLPI produced. Note that this cost calculation does not include the upstream cost of preparing the partially purified rSLPI. 

Once solubility problems have been overcome, rapid optimization of refold yield can be achieved, provided an accurate assay for the correctly folded species has been developed. The complexity of the reaction is increased because many of the variables interact with each other. We have demonstrated the effectiveness of addressing the interaction of chemical-physical conditions by a statistical experimental design approach. This allows one to optimize a host of variables without performing experiments involving every single combination. 

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