title: Mouse Aurora A: expression in Escherichia coli and purification.
abstract: Aurora kinases have recently become some of the most intensely pursued oncology targets for the design of small-molecule inhibitors. Most of the active Aurora-A protein variants are currently being expressed from baculoviruses in insect cells, while catalytically impaired proteins can also be generated in and purified from Escherichia coli. In this study we present a method of expressing large quantities of active mouse Aurora-A kinase domain as an N-terminal glutathione-S-transferase fusion protein in bacteria and outline a simple purification method that produces greater than 99% pure protein samples suitable for enzymatic assays and X-ray crystallography. The methods described in this report simplify mouse Aurora-A expression and purification, and may aid in the production of other difficult kinases in prokaryotes.
authors: Department of Protein Sciences and Structural Biology, Sunesis Pharmaceuticals, Inc., 341 Oyster Point Blvd., South San Francisco, CA 94080, USA.
authors: Robert A
authors: RA
authors: Elling
authors: Bradley T
authors: BT
authors: Tangonan
authors: David M
authors: DM
authors: Penny
authors: Jeremy T
authors: JT
authors: Smith
authors: Diana E
authors: DE
authors: Vincent
authors: Stig K
authors: SK
authors: Hansen
authors: Tom
authors: T
authors: O'Brien
authors: Michael J
authors: MJ
authors: Romanowski
mesh_terms: D000818:Animals; D064096:Aurora Kinase A; D064127:Aurora Kinases; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics; D005982:Glutathione Transferase / Q000235:genetics; D051379:Mice; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D005982:Glutathione Transferase; C576918:Aurka protein, mouse; D064096:Aurora Kinase A; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases