Design and evolution of Alzheimer's and Parkinson's antibodies
Loading...
Authors
Tiller, Kathryn Elizabeth
Issue Date
2017-08
Type
Electronic thesis
Thesis
Thesis
Language
ENG
Keywords
Chemical and biological engineering
Alternative Title
Abstract
We have also developed methods for efficiently enhancing the affinity of antibody variable domains using synthetic libraries based on natural diversity mutagenesis. Our approach involves first performing alanine-scanning mutagenesis to identify CDR positions permissive to mutagenesis. Next, we vary these permissive sites using degenerate codons that encode the wild-type residue and a few (1-5) residues that most frequently occur at each CDR site in natural antibodies. Finally, we use yeast surface display and stringent selection conditions to isolate antibodies with improved affinity and specificity. Using a variable domain specific for the Parkinson’s protein (α-synuclein) as a model system, this strategy led to a high success rate of isolating antibodies with significantly improved affinity while maintaining (or even improving) specificity. Computational modeling revealed that the enhanced affinity was not due to direct contacts between the CDR mutations and target molecule. Instead, the mutations appear to enhance affinity due to subtle structural rearrangements that enhance existing interactions or establish new interactions between wild-type CDR residues and the target molecule. These findings highlight new approaches to overcome challenges associated with in vitro antibody evolution and enable robust generation of antibodies with high specificity and stability in addition to affinity.
Description
August 2017
School of Engineering
School of Engineering
Full Citation
Publisher
Rensselaer Polytechnic Institute, Troy, NY