Influence of Surface‐Imprinted Nanoparticles on Trypsin Activity

Guerreiro, António; Poma, Alessandro; Karim, Kal; Moczko, Ewa; Takarada, Jéssica; Vargas-Sansalvador, Isabel Perez de; Turner, Nicholas W.; Piletska, Elena; Magalhães, Cristiana Schmidt de; Glazova, Natalia; Serkova, Anastasia; Omelianova, Aleksandra; Piletsky, Sergey A.

doi:10.1002/adhm.201300634

Cited by 60 publications

(43 citation statements)

References 16 publications

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“…However, these methods are largely incompatible with proteins due to the reaction environment, which leads to conformational changes or even denaturation of the protein during the imprinting process. 15 Therefore, effective and inexpensive isolation or purification processes are of substantial importance. In particular, due to the robustness of the inorganic core such imprinted hybrid particles may be used as stationary phase in chromatographic applications including SPE and HPLC.…”

Section: Introductionmentioning

confidence: 99%

Inhibitor-assisted synthesis of silica-core microbeads with pepsin-imprinted nanoshells

et al. 2016

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A novel approach for molecularly imprinting proteins, i.e. inhibitor-assisted imprinting, onto silica microspheres is discussed, which provides advanced functional materials addressing prevalent challenges in the field of protein purification and isolation from biotechnologically relevant media. Pepstatin-assisted surface-imprinted core-shell microbeads for the acidic protease pepsin were synthesized serving as selective sorbent material for solid phase extraction (SPE) applications. The inorganic core, i.e., amino-functionalized silica spheres (AFSS), is prepared by co-condensation of tetraethylorthosilicate (TEOS) and (3-aminopropyl) trimethoxysilane (APTMS) in water-in-oil (W/O) emulsion, which is then reacted with pepstatin, a selective inhibitor of pepsin, onto the surface of the AFSS via an amide bond. 3aminophenylboronic acid (APBA) serves as the functional monomer for establishing nanothin imprinted polymer films, i.e., poly (3-aminophenylboronic acid) (pAPBA) at the surface of the pepstatin-immobilized AFSS via oxidation by ammonium persulfate in aqueous solution in the presence (molecularly imprinted polymer, MIP) and absence (non-imprinted polymer; NIP) of pepsin. Thus obtained core-shell microbeads are packaged into SPE cartridges for evaluating the selectivity for pepsin. Each individual synthesis step is thoroughly characterized using x-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), transmission electron microscopy (TEM), and BET methods. Finally, the imprinted core-shell microbeads indeed provide specific binding.

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Section: Introductionmentioning

confidence: 99%

Inhibitor-assisted synthesis of silica-core microbeads with pepsin-imprinted nanoshells

et al. 2016

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“…As was stated above, control experiments were performed with nanoMIPs prepared with the same monomer composition but imprinted against an unrelated template (trypsin)17. These particles are referred to as non-imprinted polymer (NIP) to distinguish them from the target-imprinted materials.…”

Section: Resultsmentioning

confidence: 99%

A comparison of the performance of molecularly imprinted polymer nanoparticles for small molecule targets and antibodies in the ELISA format

Smolińska-Kempisty

Guerreiro

Canfarotta

et al. 2016

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103

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Here we show that molecularly imprinted polymer nanoparticles, prepared in aqueous media by solid phase synthesis with immobilised L-thyroxine, glucosamine, fumonisin B2 or biotin as template, can demonstrate comparable or better performance to commercially produced antibodies in enzyme-linked competitive assays. Imprinted nanoparticles-based assays showed detection limits in the pM range and polymer-coated microplates are stable to storage at room temperature for at least 1 month. No response to analyte was detected in control experiments with nanoparticles imprinted with an unrelated template (trypsin) but prepared with the same polymer composition. The ease of preparation, high affinity of solid-phase synthesised imprinted nanoparticles and the lack of requirement for cold chain logistics make them an attractive alternative to traditional antibodies for use in immunoassays.

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“…Highly accessible cavities with narrow (monoclonal) distribution of binding affinities and low cross-reactivity were formed. Interestingly, in dependence on the method of target immobilization-random or oriented with protected active site-after rebinding to the MIP the enzyme was either inhibited or stabilized and catalytically active [37].…”

Section: "Monoclonal Mips" For Proteinsmentioning

confidence: 98%

Cytochrome c‐Derived Hybrid Systems Based on Moleculary Imprinted Polymers

et al. 2015

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Hybrid architectures which combine a MIP with an immobilized “affinity ligand” or a biocatalyst sum up the advantages of both components. In this paper, hybrid architectures combining a layer of a molecularly imprinted electropolymer with a mini‐enzyme or a self‐assembled monolayer will be presented. (i) Microperoxidase‐11 (MP‐11) catalyzed oxidation of the drug aminopyrine on a product‐imprinted sublayer: The peroxide dependent conversion of the analyte aminopyrine takes place in the MP‐11 containing layer on top of a product‐imprinted electropolymer on the indicator electrode. The hierarchical architecture resulted in the elimination of interfering signals for ascorbic acid and uric acid. An advantage of the new hierarchical structure is the separation of MIP formation by electropolymerization and immobilization of the catalyst. In this way it was for the first time possible to integrate an enzyme with a MIP layer in a sensor configuration. This combination has the potential to be transferred to other enzymes, e.g. P450, opening the way to clinically important analytes. (ii) Epitope‐imprinted poly‐scopoletin layer for binding of the C‐terminal peptide and cytochrome c (Cyt c): The MIP binds both the target peptide and the parent protein almost eight times stronger than the non‐imprinted polymer with affinities in the lower micromolar range. Exchange of only one amino acid in the peptide decreases the binding by a factor of five. (iii) MUA‐poly‐scopoletin MIP for cytochrome c: Cyt c bound to the MIP covered gold electrode exhibits direct electron transfer with a redox potential and rate constant typical for the native protein. The MIP cover layer suppresses the displacement of the target protein by BSA or myoglobin. The combination of protein imprinted polymers with an efficient electron transfer is a new concept for characterizing electroactive proteins such as Cyt c. The competition with other proteins shows that the MIP binds its target Cyt c preferentially and that molecular shape and the charge of protein determine the binding of interfering proteins.

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Influence of Surface‐Imprinted Nanoparticles on Trypsin Activity

Cited by 60 publications

References 16 publications

Inhibitor-assisted synthesis of silica-core microbeads with pepsin-imprinted nanoshells

Inhibitor-assisted synthesis of silica-core microbeads with pepsin-imprinted nanoshells

A comparison of the performance of molecularly imprinted polymer nanoparticles for small molecule targets and antibodies in the ELISA format

Cytochrome c‐Derived Hybrid Systems Based on Moleculary Imprinted Polymers

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