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In Vitro Maturation of a Humanized Shark VNAR Domain to Improve Its Biophysical Properties

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Book cover Genotype Phenotype Coupling

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2070))

Abstract

VNAR domains are the binding regions of new antigen receptor proteins (IgNAR) which are unique to sharks, skates, and rays (Elasmobranchii). Individual VNAR domains can bind antigens independently and are the smallest reported adaptive immune recognition entities in the vertebrate kingdom. Sharing limited sequence homology with human immunoglobulin domains, their development and use as biotherapeutic agents require that they be humanized to minimize their potential immunogenicity. Efforts to humanize a human serum albumin (HSA)-specific VNAR, E06, resulted in protein molecules that initially had undesirable biophysical properties or reduced affinity for cognate antigen. Two lead humanized anti-HSA clones, v1.10 and v2.4, were subjected to a process of random mutagenesis using error-prone PCR. The mutated sequences for each humanized VNAR variant were screened for improvements in affinity for HSA and biophysical properties, achieved without a predicted increase in overall immunogenicity.

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Authors and Affiliations

  1. Elasmogen Ltd., Aberdeen, UK

    John Steven, Obinna C. Ubah, Magdalena Buschhaus, Marina Kovaleva, Laura Ferguson, Andrew J. Porter & Caroline J. Barelle

  2. Scottish Biologics Facility, School of Medical Sciences, University of Aberdeen, Aberdeen, UK

    Andrew J. Porter

Authors
  1. John Steven
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  2. Obinna C. Ubah
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  3. Magdalena Buschhaus
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  4. Marina Kovaleva
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  5. Laura Ferguson
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  6. Andrew J. Porter
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  7. Caroline J. Barelle
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Corresponding author

Correspondence to Obinna C. Ubah .

Editor information

Editors and Affiliations

  1. Protein Engineering and Antibody Technologies, Merck KGaA, Darmstadt, Germany

    Stefan Zielonka

  2. Protein Engineering and Antibody Technologies, Merck KGaA, Darmstadt, Germany

    Simon Krah

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Steven, J. et al. (2020). In Vitro Maturation of a Humanized Shark VNAR Domain to Improve Its Biophysical Properties. In: Zielonka, S., Krah, S. (eds) Genotype Phenotype Coupling. Methods in Molecular Biology, vol 2070. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9853-1_7

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  • DOI: https://doi.org/10.1007/978-1-4939-9853-1_7

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9852-4

  • Online ISBN: 978-1-4939-9853-1

  • eBook Packages: Springer Protocols

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