Unified superresolution experiments and stochastic theory provide mechanistic insight into protein ion-exchange adsorptive separations

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dc.contributor.authorKisley, Lydia
dc.contributor.authorChen, Jixin
dc.contributor.authorMansur, Andrea P.
dc.contributor.authorShuang, Bo
dc.contributor.authorKourentzi, Katerina D.
dc.contributor.authorPoongavanam, Mohan-Vivekanandan
dc.contributor.authorChen, Wen-Hsiang
dc.contributor.authorDhamane, Sagar
dc.contributor.authorWillson, Richard C.
dc.contributor.authorLandes, Christy F.
dc.date.accessioned2020-03-11T17:16:25Z
dc.date.available2020-03-11T17:16:25Z
dc.date.issued2014-02
dc.description.abstractChromatographic protein separations, immunoassays, and biosensing all typically involve the adsorption of proteins to surfaces decorated with charged, hydrophobic, or affinity ligands. Despite increasingly widespread use throughout the pharmaceutical industry, mechanistic detail about the interactions of proteins with individual chromatographic adsorbent sites is available only via inference from ensemble measurements such as binding isotherms, calorimetry, and chromatography. In this work, we present the direct superresolution mapping and kinetic characterization of functional sites on ion-exchange ligands based on agarose, a support matrix routinely used in protein chromatography. By quantifying the interactions of single proteins with individual charged ligands, we demonstrate that clusters of charges are necessary to create detectable adsorption sites and that even chemically identical ligands create adsorption sites of varying kinetic properties that depend on steric availability at the interface. Additionally, we relate experimental results to the stochastic theory of chromatography. Simulated elution profiles calculated from the molecular-scale data suggest that, if it were possible to engineer uniform optimal interactions into ion-exchange systems, separation efficiencies could be improved by as much as a factor of five by deliberately exploiting clustered interactions that currently dominate the ion-exchange process only accidentally.
dc.identifier.citationCopyright 2014 Proceedings of the Natoinal Academy of Sciences. Recommended citation: Kisley, Lydia, Jixin Chen, Andrea P. Mansur, Bo Shuang, Katerina Kourentzi, Mohan-Vivekanandan Poongavanam, Wen-Hsiang Chen, Sagar Dhamane, Richard C. Willson, and Christy F. Landes. "Unified superresolution experiments and stochastic theory provide mechanistic insight into protein ion-exchange adsorptive separations." Proceedings of the National Academy of Sciences 111, no. 6 (2014): 2075-2080. DOI: 10.1073/pnas.1318405111. URL: https://www.pnas.org/content/111/6/2075.short. Reproduced in accordance with the original publisher's licensing terms and with permission from the authors.
dc.identifier.urihttps://hdl.handle.net/10657/6216
dc.language.isoen_US
dc.publisherProceedings of the National Academy of Sciences
dc.subjection-exchange chromatography
dc.subjectsingle-molecule kinetics
dc.subjectbioseparations
dc.subjectoptical nanoscopy
dc.titleUnified superresolution experiments and stochastic theory provide mechanistic insight into protein ion-exchange adsorptive separations
dc.typeArticle

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