Obtaining active recombinant proteins from bacterial inclusion bodies using salt solutions under neutral pH conditions

Eukaryotic recombinant proteins expressed in bacterial cells usually aggregate within the cells as inclusion bodies. Despite the widely-accepted theory considering inclusion bodies as inactive materials, inclusion bodies may contain large amounts of correctly-folded active recombinant proteins. Proteins trapped in inclusion bodies can be released using a high pH solution (pH ≥ 11); however, they may undergo structural changes in such pH conditions that may lead to their inactivation. Shifting in pH alongside the use of metal ions can help recover protein activity. The model protein we used in this study, 9R-Nimo.scFv, is highly active when extracted from bacterial inclusion bodies at high pH condition (pH 12) but loses its activity when pH is reduced to pH 7. We evaluated the capacity of nine salt solutions in terms of recovering protein activity in neutral pH conditions and found that ZnSO4 solution was the best one for this purpose. KNO3 and MnSO4 were also found to have a good capacity for recovering protein activity, as well. •The ability of diverse metal ions in recovering protein activity following pH-shift inactivation is described in this article.•ZnSO4, KNO3 and MnSO4 help restore protein activity after pH-shift inactivation.•CoCl2 and CuSO4 are more suitable for protein precipitation rather than recovering protein activity.