Protein Production Core

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PROJECT SUMMARY (Protein Production Core) The Protein Production Core (PPC) serves as research support for the Oklahoma COBRE in Structural Biology (OCSB). It is a state-of the art facility for the expression and purification of protein for OCSB-supported researchers and the wider structural biology community. The PPC produces high quality proteins in a reliable and efficient manner on a fee-for-service basis and provides hands-on-training and expert advice in all aspects of protein expression and purification available in the facility. The PPC staff consists of a Director, Dr. Philip Bourne and a Graduate Student training position. The graduate student typically assists in the PPC for a semester. They are trained by the PPC director to use all of the equipment in the facility and assist in the day- to-day activities of the PPC. The PPC is guided by an advisory committee comprised of one junior and two senior faculty researchers. The PPC facility was established using OCSB Phase I funding. Milestones achieved during this period included: (i) renovation of laboratory space to create a stand-alone secure core facility, (ii) establishing the facility with modern instrumentation for protein expression and purification, and (iii) development of a user base. The user base has grown to twenty two research groups, including over 65 individual PPC users. Progress has been demonstrated by evidence of increased productivity of the user base i.e., publications and grant support. The PPC works closely with the investigators and the OCSB-supported Macromolecular Crystallography Laboratory (MCL) to produce homogeneous, highly purified protein for crystallization trials. The PPC is equipped with instrumentation and expertise frequently not found within individual research laboratories. Equipment housed in the PPC includes two refrigerated medium pressure chromatography systems and a large selection of columns for different purification techniques (affinity, hydrophobic, ion-exchange, and size exclusion). Other core equipment includes: an Emulsiflex-C3 homogenizer for lysis of yeast and bacterial cells; a high performance Avanti centrifuge, two freezers (-80 °C and -20 °C), and a BioRad ChemiDoc imager that detects UV, white and chemiluminescence. During the proposed Phase II of the OCSB, planned expansion of the PPC includes the addition of a low pressure chromatography system to ease the burden on the two medium pressure systems and expand the protein purification throughput of the facility. The addition of a small self-contained Dynamic Light Scattering instrument will allow for assessing the quality of samples prior to crystallization trials. This addition is ideal for a multi-user facility where a quick assessment of sample quality prior to crystallization experiments would be extremely useful. The increased protein production capacity and the expanded biophysical characterization will meet the existing demand and will help expand the user base. The larger user base will lead to more income from user fees and this will move the facility closer to self-sufficiency.