Document Type

Journal Article

Publisher

Nature Publishing Group

School

Centre of Excellence for Alzheimer’s Disease Research & Care / School of Medical Sciences,

RAS ID

27373

Comments

Originally published as: Khan, I., Krishnaswamy, S., Sabale, M., Groth, D., Wijaya, L., Morici, M., ... & Verdile, G. (2018). Efficient production of a mature and functional gamma secretase protease. Scientific reports, 8(1), 12834. Original article available here.

Abstract

Baculoviral protein expression in insect cells has been previously used to generate large quantities of a protein of interest for subsequent use in biochemical and structural analyses. The MultiBac baculovirus protein expression system has enabled, the use of a single baculovirus to reconstitute a protein complex of interest, resulting in a larger protein yield. Using this system, we aimed to reconstruct the gamma (γ)-secretase complex, a multiprotein enzyme complex essential for the production of amyloid-β (Aβ) protein. A MultiBac vector containing all components of the γ-secretase complex was generated and expression was observed for all components. The complex was active in processing APP and Notch derived γ-secretase substrates and proteolysis could be inhibited with γ-secretase inhibitors, confirming specificity of the recombinant γ-secretase enzyme. Finally, affinity purification was used to purify an active recombinant γ-secretase complex. In this study we demonstrated that the MultiBac protein expression system can be used to generate an active γ-secretase complex and provides a new tool to study γ-secretase enzyme and its variants.

DOI

10.1038/s41598-018-30788-w

Access Rights

free_to_read

Creative Commons License

Creative Commons Attribution 4.0 License
This work is licensed under a Creative Commons Attribution 4.0 License.

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