Polyelectrolyte–protein interaction at low ionic strength: required chain flexibility depending on protein average charge
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The effect of low ionic strength leading to reduced polyelectrolyte–protein interactions has been shown by in silico and in vitro experiments, suggesting polyelectrolyte rigidity increasing at low ionic strength, thus leading to reduced interactions with proteins. This contribution elucidates polyelectrolyte–protein precipitation in the 0–2.6-mS cm−1 ionic strength regime with polyelectrolyte rigidity determinations, using viscosimetry at these conditions, also considering protein charge distributions, using different proteins. Precipitation yields increased from 5 to 40 % at low ionic strength to up to 90 % at intermediate ionic strength, depending on protein and polyelectrolyte type, using lysozyme and three different monoclonal antibodies. Comparing precipitation behavior of the monoclonal antibodies, a qualitative correlation between required polyelectrolyte flexibility to enhance protein precipitation and protein average charge as well as hydrophobicity of the antibodies was discovered. Antibodies with lower average charge and less hydrophobicity required more flexible polyelectrolytes to enhance precipitation behavior by allowing interaction of the polyelectrolytes with proteins, attaching to positively charged protein patches while “circumnavigating” negatively charged protein areas. In contrast, antibodies with higher protein average charge showed increasing precipitation yields up to 90 % already at lower ionic strength, associated with then more rigid polyelectrolyte structures. Therefore, designing polyelectrolytes with specific chain flexibility could help to improve precipitation behavior toward specific target proteins in polyelectrolyte-driven purification techniques.
KeywordsPolyelectrolyte flexibility Proteins Structure–property relations Viscosity
The authors are grateful to Merck KGaA for financial and technical support for this project. Thanks to Merck Millipore for antibody supply. Thanks to Mikhail Kozlov, Merck Millipore, and Johann Bauer, Merck KGaA, for helpful advice on this project, and Alexandra Hill and Simon Geissler, both Merck KGaA, for providing the rheometer.
- 12.Šmigol V, Švec F, Hosoya K, Wang Q, Fréchet JM (1992) Monodisperse polymer beads as packing material for high-performance liquid chromatography. Synthesis and properties of monodisperse polystyrene and poly(methacrylate) latex seeds. Die Angewandte Makromolekulare Chemie 195:151–164. doi: 10.1002/apmc.1992.051950112 CrossRefGoogle Scholar
- 17.Gervais DP, Pfeiffer KA (2010) (Pfizer Limited) US patent 20100204455, August 12Google Scholar
- 18.Fahrner R, Franklin J, McDonald P, Peram T, Sisodiya V, Victa C (2008) (Genentech, Inc.) International patent WO/2008/091,740, January 10Google Scholar
- 19.Gronke RS, Jaquez OA (2009) (Biogen Idec MA Inc.) US Patent 12/425,328, April 16Google Scholar
- 20.Ramanan S, Stenson R (2008) (Amgen Inc.) International patent WO/2008/100,578, August 21Google Scholar
- 26.Hattori T, Hallberg R, Dubin PL (2000) Roles of electrostatic interaction and polymer structure in the binding of β-lactoglobulin to anionic polyelectrolytes: measurement of binding constants by frontal analysis continuous capillary electrophoresis. Langmuir 16:9738–9743. doi: 10.1021/la000648p CrossRefGoogle Scholar
- 43.Drifford M, Delsanti M (2001) Polyelectrolyte solutions with multivalent added salts: stability, structure, and dynamics. In: Radeva T (ed) Physical chemistry of polyelectrolytes, surfactant science series. Marcel Dekker, New York, pp 149–161Google Scholar
- 54.Kogej K, Skerjanc J (2001) Surfactant binding to polyelectrolytes. In: Radeva T (ed) Physical chemistry of polyelectrolytes, surfactant science series. Marcel Dekker, New York, pp 793–828Google Scholar
- 56.Tribet C (2001) Complexation between amphiphilic polyelectrolytes and proteins: from necklaces to gels. In: Radeva T (ed) Physical chemistry of polyelectrolytes, surfactant science series. Marcel Dekker, New York, pp 687–742Google Scholar