Rational scaffold design guided by atomic structure determination of Affibody molecule complexes
| Reference number | |
| Coordinator | Affibody AB |
| Funding from Vinnova | SEK 495 356 |
| Project duration | August 2019 - January 2021 |
| Status | Completed |
| Venture | Research infrastructure - utilisation and collaboration |
| Call | Industrial pilot projects for utilisation of neutron- and photon based techniques at large scale infrastructures - spring 2019 |
| End-of-project report | 2019-02559_AffibodyAB.pdf (pdf, 362 kB) |
Important results from the project
The overall objectives were to assess the feasibility and time need for structural determination and use that information to develop methods to guide molecular design of Affibody protein drugs in general, with respect to formatting, stability and affinity. Two subprojects were performed aiming to determine a structure of one or two Affibody molecules in complex with its antigen. Besides obtaining a high-resolution structure of an Affibody molecule, valuable insight in the process of structure determination was gained by Affibody through interaction with SARomics Biostructures
Expected long term effects
Three different Affibody molecules and two different antigens were successfully produced and characterized with regards to stability and binding kinetics. Protein (di- or tri) complexes and single Affibody molecules were set up for crystallization. Well-diffracting crystals were obtained for one antigen and one Affibody molecule, but not of a complex. The structure of a single Affibody molecule was solved and refined to 1.45 Å resolution. This high-resolution structure will serve as a tool in rational protein design to further develop the Affibody platform technology.
Approach and implementation
Affibody molecules and antigens were expressed recombinantly and purified. Affibody molecules were characterized by CD spectroscopy and antigen binding analysed by SPR measurements. Antigen:Affibody complexes were purified by SEC. Complex, antigen and Affibody molecules, respectively, were further analysed by DSF before crystallization experiments were setup. Crystals were analysed at the BioMAX beamline at MAX IV and at station I04 of Diamond Light Source, UK.