Letters to Nature

Nature 407, 395-401 (21 September 2000) | doi:10.1038/35030148; Received 26 April 2000; Accepted 24 July 2000

A chemical switch for inhibitor-sensitive alleles of any protein kinase

Anthony C. Bishop1, Jeffrey A. Ubersax2, Dejah T. Petsch3, Dina P. Matheos4, Nathanael S. Gray5, Justin Blethrow2, Eiji Shimizu4, Joe Z. Tsien4, Peter G. Schultz5, Mark D. Rose4, John L. Wood3, David O. Morgan2 & Kevan M. Shokat1,6

  1. Department of Chemistry, Princeton University, Princeton, New Jersey 08544, USA
  2. Departments of Physiology and Biochemistry & Biophysics, University of California San Francisco, San Francisco , California 94143-0444, USA
  3. Department of Chemistry, Yale University , New Haven, Connecticut 06520-8107, USA
  4. Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA
  5. Genomics Institute of the Novartis Research Foundation, San Diego, California 92121, USA
  6. Present address: Department of Cellular and Molecular Pharmacology, University of California San Francisco, San Francisco, CA 94143-0450, USA

Correspondence to: Kevan M. Shokat1,6 Correspondence and requests for material should be addressed to K.M.S. (e-mail: Email: shokat@cmp.ucsf.edu).

Protein kinases have proved to be largely resistant to the design of highly specific inhibitors, even with the aid of combinatorial chemistry1. The lack of these reagents has complicated efforts to assign specific signalling roles to individual kinases. Here we describe a chemical genetic strategy for sensitizing protein kinases to cell-permeable molecules that do not inhibit wild-type kinases2. From two inhibitor scaffolds, we have identified potent and selective inhibitors for sensitized kinases from five distinct subfamilies. Tyrosine and serine/threonine kinases are equally amenable to this approach. We have analysed a budding yeast strain carrying an inhibitor-sensitive form of the cyclin-dependent kinase Cdc28 (CDK1) in place of the wild-type protein. Specific inhibition of Cdc28 in vivo caused a pre-mitotic cell-cycle arrest that is distinct from the G1 arrest typically observed in temperature-sensitive cdc28 mutants3. The mutation that confers inhibitor-sensitivity is easily identifiable from primary sequence alignments. Thus, this approach can be used to systematically generate conditional alleles of protein kinases, allowing for rapid functional characterization of members of this important gene family.