Abstract
The ADDomer (Adenovirus-derived dodecamer) is a virus-like particle (VLP) that can be produced recombinantly with the baculovirus-insect cell expression system. This synthetic nanoparticle is based on the penton base protein of the viral capsid of the human Adenovirus serotype 3 (Ad3). Its structural properties make it an ideal subject of protein engineering efforts, as the ~70 kDa monomer has an inherent capacity towards self-organisation forming ~30 nm dodecahedrons spontaneously. The assembled particle comprises 60 penton base monomers and has a highly repetitive surface with flexible loops facing outwards. The sequence and length of these loops are not conserved among different adenovirus strains, and they can be modified without compromising the integrity of the dodecahedron. This structural feature enables the insertion of immunogenic oligo-peptide epitopes into the ADDomer scaffold that can be used for antigen presentation.In this research work, we demonstrate the rational design and production of ADDomer-based vaccine candidate molecules and protein scaffolds. The high-resolution structural information acquired by cryo-electron microscopy enables an iterative design approach towards ultimate thermostability. The varied strain-specific and chimeric scaffold molecules also permit the wider application of ADDomer-based vaccines by circumventing pre-existing immunity.
Further structural analysis of the penton base protein involved the crystallisation of the outwards facing crown domain. The structural analysis by X-ray crystallography resulted in two high-resolution (2.9 and 3.4 Å) structures of the ADDobody, a ~38 kDa monomeric protein. Its variable loops with a binding capacity to antigens demonstrated high affinity and specificity and led to the creation of an ADDobody library that can be used for the selection of synthetic ADDobody binders.
The ADDomer particle and the ADDobody have the perspective for future applications in the field of therapeutics as next-generation vaccines and high-affinity binders.
| Date of Award | 3 Oct 2023 |
|---|---|
| Original language | English |
| Awarding Institution |
|
| Supervisor | Imre Berger (Supervisor), Adrian J Mulholland (Supervisor) & Christiane H Berger-Schaffitzel (Supervisor) |