Calder’ protein engineering technology is based on targeted di-tyrosine (DT) crosslinking, which allows proteins to be locked in desired shapes. We apply this methodology to engineer novel vaccines that are locked in the shapes that elicit the most potently protective antibody responses.
Antibodies recognize 3D shapes and surfaces, and the antibodies that vaccines give rise to need to recognize the same 3D shapes of a protein on the surface of a virus when it challenges the body in order to block infection and protect against the virus. Therefore, the vaccine needs to have the same, natural 3D shape as the viral surface protein, and locking these surface proteins in their most natural shapes makes them much better protective, since the antibodies they trigger are much better able to block virus and prevent infection.
Dityrosine (DT) bonds are observed throughout nature in proteins such as silk, elastin and collagen where they provide strength, toughness and elasticity. Calder has harnessed the natural process of dityrosine crosslink formation and now applies it to the design of vaccines. We target DT bonds within the 3D structures of proteins by (i) introducing Tyr substitutions at positions that are in structural proximity, and (ii) introducing covalent crosslinks between these Tyr pairs, using a mild, natural enzymatic reaction. These DT crosslinks are zero-length carbon-carbon bonds that lock the protein irreversibly in its desired shape, without altering the rest of the molecule. This minimally modifying approach retains the target protein’s structural integrity and confers substantial advantages:
- The crosslinking reaction is targeted: it is extremely specific to Tyrosine residues; and it only results between two Tyrosine residues that are in structural proximity (<3Å). Efficiencies of >90% are routinely achieved.
- DT crosslinks are introduced after the protein is folded, they lock in the protein’s most natural shape without distorting it, and this method can be layered on existing technologies.
Tyrosine and di-tyrosine are both highly UV fluorescent, allowing DT bond formation to be measured highly sensitively, in real-time, allowing attribute-based monitoring of the reaction during manufacturing.
- DT crosslinking results in improved potency and thermo-stability and supports novel composition-of-matter IP.