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The chemical synthesis of knob domain antibody fragments
- Publication Year :
- 2021
- Publisher :
- Cold Spring Harbor Laboratory, 2021.
-
Abstract
- Cysteine-rich knob domains found in the ultralong complementarity determining regions of a subset of bovine antibodies, are capable of functioning autonomously as 3-6 kDa peptides. While they can be expressed recombinantly in cellular systems, in this paper we show that knob domains are also readily amenable to chemical synthesis, with a co-crystal structure of a chemically synthesised knob domain in complex with antigen showing structural equivalence to the biological product. For drug discovery, following immunisation of cattle, knob domain peptides can be synthesised directly from antibody sequence data, combining the power and diversity of the bovine immune repertoire with the ability to rapidly incorporate non-biological modifications. We demonstrate that, through rational design with non-natural amino acids, paratope diversity can be massively expanded, in this case improving the efficacy of an allosteric peptide. As a potential route to further improve stability, we also performed head-to-tail cyclisation, exploiting the unusual proximity of the N- and C-termini to synthesise functional, fully cyclic antibody fragments. Lastly, we highlight the stability of knob domains in plasma and, through pharmacokinetic studies, use palmitoylation as a route to extend the plasma half-life of knob domains in vivo. This study presents an antibody-derived medicinal chemistry platform, with protocols for solid-phase synthesis of knob domains; together with characterisation of their molecular structures, in vitro pharmacology and pharmacokinetics.
- Subjects :
- Male
Models, Molecular
Protein Folding
viruses
Peptide
Complementarity determining region
Computational biology
Peptides, Cyclic
Biochemistry
Rats, Sprague-Dawley
Protein Domains
Palmitoylation
Tandem Mass Spectrometry
parasitic diseases
Animals
Amino Acid Sequence
Immunoglobulin Fragments
Solid-Phase Synthesis Techniques
chemistry.chemical_classification
Drug discovery
Rational design
General Medicine
Biological product
Complementarity Determining Regions
Amino acid
chemistry
Thermodynamics
Molecular Medicine
Cattle
Paratope
Protein Binding
Subjects
Details
- Database :
- OpenAIRE
- Accession number :
- edsair.doi.dedup.....979d9fb522645065a68d3d469815cd27