Protein Expression and Proteomics

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The Protein Expression and Proteomics (PEP) Core received supplemental funding for one year starting June 1, 2006, to provide an unmet need for our community of investigators. The Protein Expression portion of the proposed pilot Core, under the direction of Dr. John Elder, will provide the appropriate vectors and cell lines needed to facilitate expression cloning of investigator-targeted proteins. The Proteomics portion of the core, under the direction of Dr. Bruce Torbett, will provide protein analysis services for CFAR investigators. These services include protein extraction from viruses, cells, and tissues; protein resolution and quantification using 1- and 2-dimensional (D) electrophoresis and imaging systems; protein identification using mass spectroscopy; and analyses of proteins using Luminex¿ xMAP¿ multiplex detection technology. The unique aspect of the PEP Core is the focus on providing services to investigators requiring expression of proteins from HIV-1 and other viral genes and protein analysis of HIV-1 and other virally infected samples. The Protein Expression portion of the Core will provide the appropriate vectors and cell lines needed to facilitate expression cloning of investigator-targeted proteins, using bacterial, insect cell, or mammalian cell vector systems, as deemed appropriate for the particular application. The Elder laboratory has compiled an extensive panel of vectors for each expression system and has extensive experience in protein expression and purification for ligand binding studies, enzymology, and crystallographic analyses. The laboratory is equipped with incubators required to grow cells at the required temperatures and also centrifuges, FPLC, and HPLC systems to aid in protein purification where necessary. The Core will aid the investigator in vector selection and amplification and cloning of targeted genes. In addition, the Core will help investigators in optimizing protein expression and selection of purification schemes most appropriate for each application. The Proteomics portion of the Core will provide all facets of protein resolution from visualization to identification. CFAR members will have access to the NCI-funded Proteomics Core housed within the Departmental of Molecular and Experimental Medicine at The Scripps Research Institute. Currently the Proteomics Core has available 1- and 2-D gel electrophoresis protein separation instrumentation, IEF Zoom fractionators for solution phase IEF, a state-of-the-art 2D Proteomic Imaging System with a Bio-Rad VersaDoc 1000 imager and imaging software. All imaging software can be utilized free of charge on resident computers housed within the Core. Technical advice is available from the Core technicians, and when needed, Core technicians provide demonstrations and seminars on analysis and imaging software. In addition, Core technicians will provide advice to investigators for optimizing protein extraction procedures and selection of purification schemes most appropriate for each application. The Protein Expression and Proteomics Core will provide informational seminars/sessions on the Core services each year and "new technology" seminars/sessions throughout the funding period.