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

Petersson, Linn LU ; Städe, Lars Wagner; Brofelth, Mattias LU ; Gärtner, Stefanie LU ; Fors, Elin LU ; Sandgren, Martin LU ; Vallkil, Jacob LU ; Olsson, Niclas LU ; Larsen, Kim Lambertsen and Borrebaeck, Carl LU , et al. (2014) In Biochimica et Biophysica Acta 1844(12). p.2164-2173
Abstract
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... (More)
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. (Less)
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publication status
published
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Biochimica et Biophysica Acta
volume
1844
issue
12
pages
2164 - 2173
publisher
Elsevier
external identifiers
  • pmid:25172394
  • wos:000345182800012
  • scopus:84907611606
ISSN
0006-3002
DOI
10.1016/j.bbapap.2014.08.010
language
English
LU publication?
yes
id
1dd665b5-e603-45d0-840a-08b777c3aea9 (old id 4612688)
date added to LUP
2014-09-11 16:55:10
date last changed
2017-07-02 04:02:15
@article{1dd665b5-e603-45d0-840a-08b777c3aea9,
  abstract     = {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.},
  author       = {Petersson, Linn and Städe, Lars Wagner and Brofelth, Mattias and Gärtner, Stefanie and Fors, Elin and Sandgren, Martin and Vallkil, Jacob and Olsson, Niclas and Larsen, Kim Lambertsen and Borrebaeck, Carl and Duroux, Laurent and Wingren, Christer},
  issn         = {0006-3002},
  language     = {eng},
  number       = {12},
  pages        = {2164--2173},
  publisher    = {Elsevier},
  series       = {Biochimica et Biophysica Acta},
  title        = {Molecular design of recombinant scFv antibodies for site-specific photocoupling to β-cyclodextrin in solution and onto solid support.},
  url          = {http://dx.doi.org/10.1016/j.bbapap.2014.08.010},
  volume       = {1844},
  year         = {2014},
}