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Molecular design of recombinant scFv antibodies for site-specific photocoupling to β-cyclodextrin in solution and onto solid support.

Author:
  • Linn Petersson
  • Lars Wagner Städe
  • Mattias Brofelth
  • Stefanie Gärtner
  • Elin Fors
  • Martin Sandgren
  • Jacob Vallkil
  • Niclas Olsson
  • Kim Lambertsen Larsen
  • Carl Borrebaeck
  • Laurent Duroux
  • Christer Wingren
Publishing year: 2014
Language: English
Pages: 2164-2173
Publication/Series: Biochimica et Biophysica Acta
Volume: 1844
Issue: 12
Document type: Journal article
Publisher: Elsevier

Abstract english

The ability to design and tailor-make antibodies to meet the biophysical demands required by the vast range of current and future antibody-based applications within biotechnology and biomedicine will be essential. In this proof-of-concept study, we have for the first time tailored human recombinant scFv antibodies for site-specific photocoupling through the use of an unnatural amino acid (UAA) and the dock'n'flash technology. In more detail, we have successfully explored the possibility to expand the genetic code of E. coli and introduced the photoreactive UAA p-benzoyl-L-phenylalanine (pBpa), and showed that the mutated scFv antibody could be expressed in E. coli with retained structural and functional properties, as well as binding affinity. The pBpa group was then used for affinity capture of the mutated antibody by β-cyclodextrin (β-CD), which provided the hydrogen atoms to be abstracted in the subsequent photocoupling process upon irradiation at 365nm. The results showed that the pBpa mutated antibody could be site-specifically photocoupled to free and surface (array) immobilized β-CD. Taken together, this paves the way for novel means of tailoring recombinant scFv antibodies for site-specific photochemical-based tagging, functionalization and immobilization in numerous applications.

Keywords

  • Biological Sciences

Other

Published
  • ISSN: 0006-3002
Carl B
Carl Borrebaeck
E-mail: carl.borrebaeck [at] immun.lth.se

Professor

Department of Immunotechnology

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