Chemically Selective Displacers for Protein Purification

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PROPOSAL NUMBER: 0730830PRINCIPAL INVESTIGATOR: Steven CramerINSTITUTION: Rensselaer Polytechnic InstitutePROPOSAL TITLE: Chemically Selective Displacers for Protein PurificationThe effects of protein, displacer structure, mobile phase characteristics and displacer concentration on the efficacy and selectivity of displacement chromatography systems will be examined using an automated liquid handling system to enable multi-dimensional screens. High affinity and selective displacer lead compounds selected from the high throughput screening (HTS) studies will be synthesized. Throughout this synthesis work various approaches to introduce chromophores and/or fluorophores into these displacers will be examined in order to facilitate on-line detection. This work will examine displacers that have the ability to bind to specific classes of proteins. In addition to selective displacers identified from the HTS studies, this work will investigate the a priori design of class selective displacers using known binding ligands tethered to appropriately charged moieties. Displacers developed in this phase of the project will be evaluated for their ability to selectively displace the desired proteins under batch and column experiments. The mode of interaction between selective displacers and proteins will also be investigated using multiple techniques. A four state simulation model based on the steric mass action isotherm will be used to predict column performance of these selective displacers. The proposed research will have an impact on both large scale bioprocessing as well as analytical biotechnology. The development of selective displacers that bind to a specific protein or protein class while also acting as efficient displacers will dramatically increase the selectivity of ion exchange systems. The proposed work could have a major impact on the purification of key proteins involved in cellular regulation. The proposed research will also have an important impact on the education of both graduate and undergraduate chemical engineering students. The students involved in this multidisciplinary project will obtain significant training in chromatographic bioprocessing, modeling, robotics, and a variety of analytical techiques such as capillary zone electrophoresis, surface plasmon resonance and NMR. The research from this project will be incorporated into a course on Chromatographic Separation Processes. Finally, high school students who are carrying out internships in the PI's laboratory will also obtain important laboratory experience. If successful the proposal will have a significant and broad impact on bioprocessing and a number of health related fields.