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Required polymer lengths per precipitated protein molecule in protein-polymer interaction

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Abstract

Protein precipitation using non-charged and charged polymers is a common method for protein purification, gaining broader interest among manufacturers in downstream processing. While during polymer- surface interactions, the formation of loops, tails and trains has been known for quite a long time, details of polymer conformation and chain length, interacting with the protein during protein precipitation are not fully discovered. Our research presents a more profound understanding of polymer-protein interaction, combining fluorescence and infrared spectroscopic measurements of proteins and polymer standards with well defined chain length to confirm different models of protein-polymer interaction. Lysozyme, chymotrypsinogen A, myoglobin and a monoclonal antibody, all of different molecular weight, isoelectric point and charge distribution at the protein surface, were used for protein-polymer precipitation. The use of polymers of various charge density and chain length showed that the required polymer length per precipitated protein (Ldef) is up to 25-times larger than the diameter of the corresponding protein, depending on the surface charge distribution of the protein, and its isoelectric point, as well as the charge density of the polymer. Our results support proposed mechanisms of polymer wrapping and loop formation for optimal charge neutralization during complexation and imply interaction of several polymer chains per precipitated protein molecule. Electrophoretic light scattering showed a qualitative correlation of the zeta potential of analyzed polymers with their corresponding Ldef values. Comparing protein precipitation behavior of long and short polymer chains, the latter exhibited reduced precipitation efficiency, visible as elevated Ldef.

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Acknowledgments

The authors thank Merck KGaA for financial and material support. Thanks to Johann Bauer and Stephan von der Au, both Merck KGaA for helpful advice and supplying the Zetasizer Nano. Part of this work was performed within the frame of the project BIOPUR and IOLIPRO, funded by the German Federal Ministry of Education and Research (BMBF). We thank Charly Hohlschuh, Merck KGaA, for language assistance and proof reading.

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Authors and Affiliations

  1. Clemens-Schöpf Institute, Technical University of Darmstadt, Darmstadt, Germany

    Florian Capito & Harald Kolmar

  2. Merck KGaA, Frankfurter Strasse 250, 64293, Darmstadt, Germany

    Florian Capito, Bernd Stanislawski & Romas Skudas

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  1. Florian Capito
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  2. Harald Kolmar
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  3. Bernd Stanislawski
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  4. Romas Skudas
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Correspondence to Florian Capito.

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Capito, F., Kolmar, H., Stanislawski, B. et al. Required polymer lengths per precipitated protein molecule in protein-polymer interaction. J Polym Res 21, 346 (2014). https://doi.org/10.1007/s10965-013-0346-7

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