Investigating Affinity Maturation of Sars-Cov-2 Antibody CC12.1

Antibodies are essential to the body’s adaptive immune response. Certain antibodies known as neutralizing antibodies can neutralize host infection of viruses by binding noncovalently to certain key antigenic regions of the viral surface. The current research landscape has gaps yet to be investigated for how these neutralizing antibodies develop from immature precursors (rearranged from germline precursors). Understanding this development pathway is crucial to vaccine development to optimize viral-specific neutralizing antibodies. In my project I will investigate the likely development pathway of the neutralizing antibody CC12.1 which binds the SARS-CoV-2 spike protein. The binding blocks the spike protein from binding human angiotensin-converting enzyme 2 (ACE2), therefore neutralizing the virus. CC12.1 was isolated from a patient infected with the original strain (Wuhan-Hu-1), but it no longer neutralizes current variants such as Delta and Omicron. I used nicking mutagenesis to make all possible mutations between the immature precursor and matured CC12.1, displayed these mutants as antibody Fab fragments on the surface of yeast, and performed cell surface titrations against the Wuhan-Hu-1 Spike protein. Our results show that mature CC12.1 has a much lower dissociation constant against Spike than the immature precursor, as expected. We also identify key gain of binding affinity mutations in the development pathway.