Different Approaches to Stabilize a Recombinant Fusion Protein

Abstract

We have used a fusion protein between staphylococcal protein A and E. coli β–galactosidase as a model system to investigate different approaches to stabilize recombinant gene products. First, growth conditions were adapted to preferentially produce insoluble inclusion bodies. This material was resistant to proteolysis, but no active fusion protein could be recovered. Secondly, biochemical characterization revealed that the soluble fusion protein was cleaved by a membrane–bound protease at a Lys–Arg peptide bound in the linker region. A new linker region was therefore engineered lacking the protease sensitive sequence. The new fusion protein was found to be resistant to degradation and high levels of soluble and active fusion protein could be produced. Finally, a mutant E. coli strain lacking the outer membrane protease OmpT was tested as a production strain, and a stable fusion protein could be recovered. These results demonstrate the use of several independent strategies to avoid degradation of recombinant proteins in heterologous hosts.

Cite this article

Hellebust, H., Murby, M., Abrahmsén, L. et al. Different Approaches to Stabilize a Recombinant Fusion Protein. Nat Biotechnol 7, 165–168 (1989). https://doi.org/10.1038/nbt0289-165

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