Understanding differences between synthetic and natural antibodies can help improve antibody engineering

Abstract

Synthetic libraries are a major source of human-like antibody (Ab) drug leads. To assess the similarity between natural Abs and the products of these libraries, we compared large sets of natural and synthetic Abs using "CDRs Analyzer," a tool we introduce for structural analysis of Ab-antigen (Ag) interactions. Natural Abs, we found, recognize their Ags by combining multiple complementarity-determining regions (CDRs) to create an integrated interface. Synthetic Abs, however, rely dominantly, sometimes even exclusively on CDRH3. The increased contribution of CDRH3 to Ag binding in synthetic Abs comes with a substantial decrease in the involvement of CDRH2 and CDRH1. Furthermore, in natural Abs CDRs specialize in specific types of non-covalent interactions with the Ag. CDRH1 accounts for a significant portion of the cation-pi interactions; CDRH2 is the major source of salt-bridges and CDRH3 accounts for most hydrogen bonds. In synthetic Abs this specialization is lost, and CDRH3 becomes the main sources of all types of contacts. The reliance of synthetic Abs on CDRH3 reduces the complexity of their interaction with the Ag: More Ag residues contact only one CDR and fewer contact 3 CDRs or more. We suggest that the focus of engineering attempts on CDRH3 results in libraries enriched with variants that are not naturallike. This may affect not only Ag binding, but also Ab expression, stability and selectivity. Our findings can help guide library design, creating libraries that can bind more epitopes and Abs that better mimic the natural antigenic interactions.

Computational Design of Epitope-Specific Functional Antibodies

“…It has been shown that in naturally occurring antibodies, CDRs specialize in certain types of contacts (Burkovitz and Ofran, 2016;Kunik and Ofran, 2013;Kunik et al, 2012b). Thus, rather than relying on random mutagenesis, library design was based on analysis of the structure and sequence of the antibody.…”

Alanine Mutagenesis in the Complementarity Determining Region 3 of the MTB and HIV-1 Peptide-Bispecific T Cell Receptor Beta Chain Affects Ligand Recognition

“…This could be due to the fact that Asp 114 is likely to form a salt bridge with a positively charged residue or the positively charged antigen in peptide–MHC complex. A lot of antigen–antibody interfaces include one or more salt bridges ( 34 ). The S118A mutation, which removes a single side chain hydroxyl group, eliminated the antigen response, illustrating the significant impact that small structural changes can have on T cell recognition.…”

Construction of Antibody Phage Libraries and Their Application in Veterinary Immunovirology

“…The PCR-amplified V H and V L and V H -C H and V L -C L gene fragments are ligated in a suitable phagemid for single-chain variable fragment (scFv) and Fab library generation, respectively. In the case of synthetic antibody phage libraries, the initial few steps are not needed, and library diversity is increased through the precise introduction of degenerate DNA into CDR encoding regions, thus rivaling or exceeding than that of natural immune repertoire [5].…”

“…Antibodies 2020, 9, x FOR PEER REVIEW 3 of 13 fragment (scFv) and Fab library generation, respectively. In the case of synthetic antibody phage libraries, the initial few steps are not needed, and library diversity is increased through the precise introduction of degenerate DNA into CDR encoding regions, thus rivaling or exceeding than that of natural immune repertoire [5]. The engineered phagemids are transformed into suitable bacteria (e.g., XL1-Blue, TG1, and ER2537), providing a suitable environment for recombination of antibody fragments [6].…”