Protein Expression and Purification

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The major objective of this Core is to supply the various Projects and Cores with a continuous supply of highly purified protein in amounts necessary to complete the individual specific aims for each Project and Core, and therefore the long-term objectives of the overall Program Project. The specific aims are to (1) collaborate with Projects 1 & 2 in the cloning of SCLpro and PLpro into over-expression vectors, and to then determine their optimum expression conditions for each enzyme; (2) develop rapid and efficient purification procedures for SCLpro and PLpro enzyme constructs and determine the optimum conditions for their stabilization and storage; and (3) produce large amounts of highly purified enzyme that will be utilized by Projects 1-3, and by computational Core. The ultimate goal is to provide Projects 1-3, and the computational Core with a continuous supply of protein for the purposes of biochemical, kinetic, and biophysical characterization, x-ray structure elucidation, and high-throughput enzymatic and ligand-binding and bioassays.