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Proof of concept of a “greener” protein purification/enrichment method based on carboxylate-terminated carbosilane dendrimer-protein interactions

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Abstract

Protein sample preparation is a critical and an unsustainable step since it involves the use of tedious methods that usually require high amount of solvents. The development of new materials offers additional opportunities in protein sample preparation. This work explores, for the first time, the potential application of carboxylate-terminated carbosilane dendrimers to the purification/enrichment of proteins. Studies on dendrimer binding to proteins, based on protein fluorescence intensity and emission wavelengths measurements, demonstrated the interaction between carboxylate-terminated carbosilane dendrimers and proteins at all tested pH levels. Interactions were greatly affected by the protein itself, pH, and dendrimer concentration and generation. Especially interesting was the interaction at acidic pH since it resulted in a significant protein precipitation. Dendrimer-protein interactions were modeled observing stable complexes for all proteins. Carboxylate-terminated carbosilane dendrimers at acidic pH were successfully used in the purification/enrichment of proteins extracted from a complex sample.

Images showing the growing turbidity of solutions containing a mixture of proteins (lysozyme, myoglobin, and BSA) at different protein:dendrimer ratios (1:0, 1:1, 1:8, and 1:20) at acidic pH and SDS-PAGE profiles of the corresponsing supernatants. Comparison of SDS-PAGE profiles for the pellets obtained during the purification of proteins present in a complex sample using a conventional “no-clean” method based on acetone precipitation and the proposed “greener” method using carboxylate-terminated carbosilane dendrimer at a 1:20 protein:dendrimer ratio

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Acknowledgments

This work was supported by the Ministry of Economy and Competitiveness (ref. AGL2012-36362 and CTQ-2014-54004-P), the Comunidad de Madrid and European funding from FEDER program (ref. S2013/ABI-3028, AVANSECAL), and Consortium NANODENDMED ref. S2011/BMD-2351 (CAM). The financial support of the Czech Science Foundation (project no. GA15-05903S) is also acknowledged. E.G.-G. thanks the University of Alcalá for her pre-doctoral contract. CIBER-BBN is an initiative funded by the VI National R&D&i Plan 2008–2011, Iniciativa Ingenio 2010, Consolider Program, CIBER Actions and financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund.

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Authors and Affiliations

  1. Departamento de Química Analítica, Química Física e Ingeniería Química, Universidad de Alcalá, Ctra. Madrid-Barcelona Km. 33.600, 28871, Alcalá de Henares, Madrid, Spain

    Estefanía González-García, María Luisa Marina & María Concepción García

  2. Faculty of Science, J. E. Purkinje University, Ceske mladeze 8, 40096, Usti nad Labem, Czech Republic

    Marek Maly

  3. Departamento de Química Orgánica y Química Inorgánica, Universidad de Alcalá, Ctra. Madrid-Barcelona, Km 33.600, 28871, CIBER-BBN, Alcalá de Henares, Madrid, Spain

    Francisco Javier de la Mata & Rafael Gómez

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  1. Estefanía González-García
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  2. Marek Maly
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  3. Francisco Javier de la Mata
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Correspondence to María Concepción García.

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González-García, E., Maly, M., de la Mata, F.J. et al. Proof of concept of a “greener” protein purification/enrichment method based on carboxylate-terminated carbosilane dendrimer-protein interactions. Anal Bioanal Chem 408, 7679–7687 (2016). https://doi.org/10.1007/s00216-016-9864-6

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  • DOI: https://doi.org/10.1007/s00216-016-9864-6

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