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Guide to the Preparation of Molecularly Imprinted Polymer Nanoparticles for Protein Recognition by Solid-Phase Synthesis.

Authors
  • Xu, Jingjing1
  • Medina-Rangel, Paulina X1
  • Haupt, Karsten2
  • Tse Sum Bui, Bernadette3
  • 1 Sorbonne Universités, Université de Technologie de Compiègne, CNRS Enzyme and Cell Engineering Laboratory, Rue Roger Couttolenc, Compiègne Cedex, France. , (France)
  • 2 Sorbonne Universités, Université de Technologie de Compiègne, CNRS Enzyme and Cell Engineering Laboratory, Rue Roger Couttolenc, Compiègne Cedex, France. Electronic address: [email protected] , (France)
  • 3 Sorbonne Universités, Université de Technologie de Compiègne, CNRS Enzyme and Cell Engineering Laboratory, Rue Roger Couttolenc, Compiègne Cedex, France. Electronic address: [email protected] , (France)
Type
Published Article
Journal
Methods in enzymology
Publication Date
Jan 01, 2017
Volume
590
Pages
115–141
Identifiers
DOI: 10.1016/bs.mie.2017.02.004
PMID: 28411635
Source
Medline
Keywords
License
Unknown

Abstract

Molecularly imprinted polymers (MIPs) are synthetic antibody mimics possessing specific cavities designed for a target molecule. Nowadays, molecular imprinting of proteins still remains a challenge as the generation of selective imprinted cavities is extremely difficult, due to their flexible structure and the presence of a multitude of functional sites. To overcome this difficulty, we propose a solid-phase synthesis strategy to prepare MIPs specific for any protein that can be immobilized in an oriented way on a solid support. Trypsin and kallikrein were used as model proteins. The solid-phase support consists of glass beads functionalized with two affinity ligands of the enzymes, the competitive inhibitor p-aminobenzamidine to orient the enzymes via their active site, or a Cu2+chelate to orient via the surface histidine residues of the enzyme. Thermoresponsive molecularly imprinted polymer nanoparticles (MIP-NPs) are then synthesized around the immobilized enzyme. The MIP-NPs are released by a simple temperature change, resulting in protein-free polymers endowed with improved binding site homogeneity since all binding sites have the same orientation. The MIP-NPs exhibit apparent dissociation constants between 0.02 and 2nM toward their target proteins, which is comparable to those of natural antibodies. Moreover, these water-compatible polymers, targeting different domains of the enzyme, can also function as protective agents (armor), hence preventing the target proteins from denaturation by heat or pH.

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