1) Bacterial Protein Expression Core (Hill. Director: Schubert. Manager)OverviewThe Bacterial Protein Expression Core is located on the third floor of the EEJMRB at the University ofUtah In close proximity to the labs of Hill, Sundquist, and Kay. It is managed by Heidi Schubert and willbe staffed by two Center-funded technicians. The facility provides efficient construction of expressionvectors, testing of protein expression and solubility, protein purification, and characterization. We haveestablished a synergistic relationship with the Utah Molecular Hematology Protein Expression Core forthe cost effective production of recombinant TEV protease and Pfu polymerase, and employ services ofthe University of Utah core facilities in mass spectrometry, oligonucleotide synthesis, N-terminal proteinsequencing, and DNA sequencing. The Bacterial Protein Expression Core facilities are integrated withoperations of the Eukaryotic Protein Expression Core for bioinformatics, cloning, and protein purification,and there is also a close association with protein structure determination efforts in X-ray and ProteinNMR cores. This core will therefore facilitate structural and biochemical studies on targets identified bythe Center members pursuing questions of HIV/Host biology.In general, the methods used in this Core are standard, but have been adapted to maximize efficiencyon a scale appropriate for the targeted, but ambitious, scope of this application. The basicinstrumentation and protocols are already in place and will be expanded to match increased demand ifthe Center is funded. The increased throughput will be achieved by efficient database management,consolidated bioinformatics resources, use of technical staff, and standardization and refinement ofprotocols. The approaches are an extension of the methodology that has supported structural andbiochemical studies by the Hill and Sundquist labs (see biosketches for publications). Our initial targetcapacity is an average of 24 new expression vectors constructed per week, although the actual rate willfluctuate in response to demand. This level of throughput will allow aggressive approaches to problems,such as optimizing N- and C-termini or making surface mutations to produce crystallizable constructs. Ifdemand exceeds capacity, we can add personnel/instrumentation/supplies on a charge-back basis.
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