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Refolding of Aggregation‐Prone ScFv Antibody Fragments Assisted by Hydrophobically Modified Poly(sodium acrylate) Derivatives
Author(s) -
Martin Nicolas,
Costa Narciso,
Wien Frank,
Winnik Françoise M.,
Ortega Céline,
Herbet Amaury,
Boquet Didier,
Tribet Christophe
Publication year - 2017
Publication title -
macromolecular bioscience
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.924
H-Index - 105
eISSN - 1616-5195
pISSN - 1616-5187
DOI - 10.1002/mabi.201600213
Subject(s) - chemistry , circular dichroism , polymer , folding (dsp implementation) , protein aggregation , acrylate , macromolecule , protein folding , biophysics , combinatorial chemistry , polymer chemistry , biochemistry , monomer , organic chemistry , electrical engineering , biology , engineering
ScFv antibody fragments are a promising alternative to full‐length antibodies for both therapeutic and diagnosis applications. They can be overexpressed in bacteria, which enables easy large scale production. Since scFv are artificial constructs, they are poorly soluble and prone to aggregation, which makes them difficult to manipulate and to refold. Here, stabilization and refolding of scFv fragments from urea‐unfolded solutions are reported based on the use of micromolar amounts of polymers playing the role of artificial chaperons. Using fluorescence correlation spectroscopy, the size and aggregation number of complexes of scFv with unmodified or hydrophobically modified poly(sodium acrylate) are determined. The evolution of the secondary structure along the refolding procedure, in the presence or absence of 0.4 m l‐ arginine at scFv:polymer < 1:5 (w/w), is determined by high‐sensitivity synchrotron‐radiation circular dichroism. Measurements reveal that refolding in the presence of polymers yields native‐like secondary structure, though a different folding pathway can be followed compared to refolding in the absence of polymer. This is the first report on the use of macromolecular additives to assist refolding of a multidomain protein of therapeutic interest.