Understanding and enhancing selective Fab separations using multimodal chromatography
March 5-10, 2017
This research seeks to determine what conditions are required to achieve selective separations of similar protein variants and to provide fundamental insight into the mechanisms underlying these separations. The retention of protein libraries on several multimodal cation-exchange systems demonstrated that the retention of many proteins proved to be sensitive to subtle changes in the ligand chemistry and geometrical presentation. All-atom explicit Molecular Dynamics (MD) simulations were then carried out to shed light on the multiple weak interactions that resulted in the unique selectivities achieved in these multimodal chromatographic systems. A range of biophysics techniques including NMR, ITC and AFM were also employed to study the energetics, kinetics and thermodynamics of protein binding to self-assembled monolayers (SAMs) of MM ligands. Finally, quantitative structure activity relationship models were developed using unique molecular descriptors inspired by the biophysics and simulation studies for predicting the chromatographic behavior of proteins on a many multimodal resin systems and to provide insight into the relative importance of various regions on Fabs for interacting with these ligands. This work provides fundamental understanding of the nature of these interactions at the molecular level and insight into the design of MM ligands with important implications for addressing challenging problems in downstream bioprocessing.
Steven Cramer, "Understanding and enhancing selective Fab separations using multimodal chromatography" in "Separations Technology IX: New Frontiers in Media, Techniques, and Technologies", Kamalesh K. Sirkar, New Jersey Institute of Technology, USA Steven M. Crame, Rensselaer Polytechnic Institute, USA João G. Crespo, LAQV-Requimte, FCT-Universidade Nova de Lisboa, Caparica, Portugal Marco Mazzotti, ETH Zurich, Switzerland Eds, ECI Symposium Series, (2017). https://dc.engconfintl.org/separations_technology_ix/43