Obtaining active single chain antibody from bacterial inclusion bodies using metal ions

Publication date: Available online 27 January 2020Source: Protein Expression and PurificationAuthor(s): Marzieh Najafi, Yaghoub SafdariAbstractEukaryotic 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. Molecules 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 lead to their protein 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 thatZnSO4 solution was the best one for this purpose. KNO3, and MnSO4 were also found to have a good capacity in recovering protein activity, as well.
Source: Protein Expression and Purification - Category: Biochemistry Source Type: research