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- W4317475595 abstract "Inclusion bodies (IB) are dense insoluble aggregates of mostly misfolded polypeptides that usually result from recombinant protein overexpression. IB formation has been observed in protein expression systems such as E. coli, yeast, and higher eukaryotes. To recover soluble recombinant proteins in their native state, IB are commonly first solubilized with a high concentration of denaturant. This is followed by concurrent denaturant removal or reduction and a transition into a refolding-favorable chemical environment to facilitate the refolding of solubilized protein to its native state. Due to the high concentration of denaturant used, conventional refolding approaches can result in dilute products and are buffer inefficient. To circumvent the limitations of conventional refolding approaches, a temperature-based refolding approach which combines a low concentration of denaturant (0.5 M guanidine hydrochloride, GdnHCl) with a high temperature (95 °C) during solubilization was proposed. In this chapter, we describe a temperature-based refolding approach for the recovery of core streptavidin (cSAV) from IB. Through the temperature-based approach, intensification was achieved through the elimination of a concentration step which would be required by a dilution approach and through a reduction in buffer volumes required for dilution or denaturant removal. High-temperature treatment during solubilization may have also resulted in the denaturation and aggregation of undesired host-cell proteins, which could then be removed through a centrifugation step resulting in refolded cSAV of high purity without the need for column purification. Refolded cSAV was characterized by biotin-binding assay and SDS-PAGE, while purity was determined by RP-HPLC." @default.
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- W4317475595 date "2023-01-01" @default.
- W4317475595 modified "2023-10-18" @default.
- W4317475595 title "Intensification of Inclusion Body Processing via Temperature-Based Refolding" @default.
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- W4317475595 doi "https://doi.org/10.1007/978-1-0716-2930-7_13" @default.
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