The proximity of the N- and C- termini of bovine knob domains enable engineering of target specificity into polypeptide chains

Alice Hawkins, Callum Joyce, Kevin Brady, Adam Hold, MIchael Knight, Conor Howard, Jean Van Den Elsen, Alastair D G Lawson, Alex Macpherson

Research output: Contribution to journalArticlepeer-review

7 Citations (SciVal)


Cysteine-rich knob domains can be isolated from the ultralong heavy-chain complementarity-determining region (CDR) 3, which are unique to a subset of bovine antibodies, to create antibody fragments of ~4 kDa. Advantageously, the N- and C- termini of these small binding domains are in close proximity, and we propose that this may offer a practical route to engineer extrinsic binding specificity into proteins. To test this, we transplanted knob domains into various loops of rat serum albumin, targeting sites that were distal to the interface with the neonatal Fc receptor. Using knob domains raised against the clinically validated drug target complement component C5, we produced potent inhibitors, which exhibit an extended plasma half-life in vivo via attenuated renal clearance and neonatal Fc receptor-mediated avoidance of lysosomal catabolism. The same approach was also used to modify a Camelid VHH, targeting a framework loop situated at the opposing end of the domain to the CDRs, to produce a small, single-chain bispecific antibody and a dual inhibitor of Complement C3 and C5. This study presents new protein inhibitors of the complement cascade and demonstrates a broadly applicable method to engineer target specificity within polypeptide chains, using bovine knob domains.
Original languageEnglish
Article number2076295
Issue number1
Early online date29 May 2022
Publication statusPublished - 1 Jun 2022

Bibliographical note

Funding Information:
The author(s) reported there is no funding associated with the work featured in this article.

Publisher Copyright:
© 2022 UCB Biopharma UK. Published with license by Taylor & Francis Group, LLC.


  • Bovine antibodies
  • V
  • bispecific
  • complement C5
  • knob domain
  • serum albumin

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology


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