Core C - Protein Expression and Purification Core The success of the entire Program of Projects hinges on the availability of large amounts of protein reagents as well as on the ability to rapidly clone, express and purify a very diverse set of mutant proteins. For example, hundreds of milligrams of core histones and histone chaperones will be required for the proposed research. Additionally, the Projects demand the flexibility to prepare smaller amounts of specific mutants and truncations of histones and chaperones. Specifically, these reagents will be required for the studies on histone acetylation patterns, histone chaperone interactions, nucleosome dynamics and regulation of higher order chromatin structures. Smaller quantities of highly pure histone chaperones, histone acetyl transferases, transcription factors and coactivators will also be needed. Rigorous quality control of the shared reagents will ensure maximum reproducibility for the research in the three Projects. The Protein Expression and Purification (PEP) Core is fully equipped for the high-throughput preparation of large amounts of proteins. The Director, Dr. Karolin Luger, and the Associate Director, Teri McLain, have many years of experience in the large-scale and small-scale purification of a large variety of proteins. The PEP CORE has the following specific aims:
Aim 1 : Express and purify histones from E. coli.
Aim 2 : Express and purify histone chaperones from E. coli and baculovims-infected Sf9 cells.
Aim 3 : express and purify histone acetyltransferases.
Aim 4 : Express and Purify Activators and Accessory Proteins.
Aim 5 : Extract and Purify Endogenous Proteins from Tissue and Cells.
Aim 6 : Maintain an Up-To-Date Inventory of All Protein Produced by the PEP CORE in the Chromatin Intranet Website.
|Groocock, Lynda M; Nie, Minghua; Prudden, John et al. (2014) RNF4 interacts with both SUMO and nucleosomes to promote the DNA damage response. EMBO Rep 15:601-8|
|Muthurajan, Uma M; Hepler, Maggie R D; Hieb, Aaron R et al. (2014) Automodification switches PARP-1 function from chromatin architectural protein to histone chaperone. Proc Natl Acad Sci U S A 111:12752-7|
|Chodaparambil, Jayanth V; Pate, Kira T; Hepler, Margretta R D et al. (2014) Molecular functions of the TLE tetramerization domain in Wnt target gene repression. EMBO J 33:719-31|
|Kalashnikova, Anna A; Porter-Goff, Mary E; Muthurajan, Uma M et al. (2013) The role of the nucleosome acidic patch in modulating higher order chromatin structure. J R Soc Interface 10:20121022|
|Sheinin, Maxim Y; Li, Ming; Soltani, Mohammad et al. (2013) Torque modulates nucleosome stability and facilitates H2A/H2B dimer loss. Nat Commun 4:2579|
|Rogge, Ryan A; Kalashnikova, Anna A; Muthurajan, Uma M et al. (2013) Assembly of nucleosomal arrays from recombinant core histones and nucleosome positioning DNA. J Vis Exp :|
|Hsieh, Fu-Kai; Kulaeva, Olga I; Patel, Smita S et al. (2013) Histone chaperone FACT action during transcription through chromatin by RNA polymerase II. Proc Natl Acad Sci U S A 110:7654-9|
|Kalashnikova, Anna A; Winkler, Duane D; McBryant, Steven J et al. (2013) Linker histone H1.0 interacts with an extensive network of proteins found in the nucleolus. Nucleic Acids Res 41:4026-35|
|D'Arcy, Sheena; Martin, Kyle W; Panchenko, Tanya et al. (2013) Chaperone Nap1 shields histone surfaces used in a nucleosome and can put H2A-H2B in an unconventional tetrameric form. Mol Cell 51:662-77|
|Elsasser, Simon J; D'Arcy, Sheena (2012) Towards a mechanism for histone chaperones. Biochim Biophys Acta 1819:211-21|
Showing the most recent 10 out of 17 publications