Features of the solubilization of interferon beta-1B from inclusion bodies

Abstract

A new solubilization method of recombinant interferon beta-1b (IFNβ-1b) from the inclusion bodies was developed. This method allows to extract the target protein selectively in the solutions of different alcohols, such as ethanol, propanol and isopropanol. It was shown that the more effective IFNβ-1b solubilization was achieved in the 55% propanol solution. This method allowed to extract the target protein from inclusion bodies around 85-90%, and significantly reduced Escherichia coli content in the solubilizate, in comparison with standard methods.

Solubilization of Human Interferon β-1b Inclusion Body Proteins by Organic Solvents

“…value higher than twelve results in partial degradation of the protein. 21 In fact, they confirmed the previous results which have shown that the dissolution of IBs at high pH's can lead to irreversible protein denaturation. 22 On the other hand, protein dissolution can be both detergent and pH-dependent which indicates the presence of ionic and hydrophilic interactions in IBs.…”

“…There is an estimation that organic solvents such as alcohols with less polarity have more effect on solubilizing IB proteins; therefore, 2-butanol was added to the reactions as a constant content. 21 As Figure 1 indicates, the acidic environment has a higher impact on solubilizing the secondary nativelike proteins from inclusion bodies and have yielded more than other buffer solutions (Figure 1; sample no. 6).…”

“…22 On the other hand, protein dissolution can be both detergent and pH-dependent which indicates the presence of ionic and hydrophilic interactions in IBs. 21 Moreover, according to findings of Nick Pace et al unfolded protein state requires more net charges to better interact with denaturant agent for dissolution. These net charges are prepared when pH is lowered.…”

“…They have also revealed that the solubilization of the intended protein by a solution with pH=2.0 and using no urea or GdnHCl has resulted in a maximum release of interferon beta-1b from IB. 21 Furthermore, according to Vallejo's findings, dissolution of the inclusion bodies using a detergent such as urea is sensitive to pH. Vallejo also indicates that the optimal pH for protein dissolution should be determined.…”

Disulfide–bridging PEGylation during refolding for the more efficient production of modified proteins

“…The majority of the studies described here were conducted with PEG‐ mono ‐sulfone 3 that was derived from a 20 kDa PEG precursor (hence PEG 20 ‐ mono ‐sulfone 3 ). IFN‐β1b is produced from E. coli , however it is well known to be a hydrophobic protein that can be difficult to refold from IBs . IFN‐β1b is a cytokine and is an example of an α‐helical barrel protein, of which several are used clinically .…”