Directed evolution and optimization of antibodies for enhanced affinity and stability

Julian, Mark C.
Thumbnail Image
Other Contributors
Tessier, Peter M.
Garde, Shekhar
Collins, Cynthia H.
Colón, Wilfredo
Issue Date
Chemical engineering
Terms of Use
This electronic version is a licensed copy owned by Rensselaer Polytechnic Institute, Troy, NY. Copyright of original work retained by author.
Full Citation
Mutational analysis of these evolved antibodies reveals that many affinity-enhancing mutations within the antibody binding loops are destabilizing and that compensatory mutations are necessary to maintain antibody stability. Interestingly, our finding that it is necessary to accumulate compensatory mutations during affinity maturation to maintain thermodynamic stability for antibodies generated in vitro is similar to findings for antibodies generated in vivo, which suggests that the formation of the antigen-binding site is generally a destabilizing process. We have used this knowledge to develop refined mutagenesis and selection techniques for generating and sorting antibody libraries. Our libraries have mutations focused in the most important antibody binding loop and follow patterns of amino acid diversity found across thousands of natural antibody sequences. We have used these novel libraries as well as strong positive and negative selections to identify interesting antibody variants that target aggregated forms of the Alzheimer’s Aβ peptide with high binding affinity and specificity. Further, the stability and specificity of the selected antibodies are similar to those observed for natural antibodies and far superior to the properties of other antibodies we have identified using conventional in vitro methods. Collectively, our work provides new strategies to improve upon the design, evolution and selection of in vitro antibody libraries to yield high affinity antibody variants with favorable biophysical properties.
May 2017
School of Engineering
Dept. of Chemical and Biological Engineering
Rensselaer Polytechnic Institute, Troy, NY
Rensselaer Theses and Dissertations Online Collection
Restricted to current Rensselaer faculty, staff and students. Access inquiries may be directed to the Rensselaer Libraries.