Abstract:

Tyrosine kinases are an important family of signaling proteins in eukaryotic organisms and are involved in controlling the growth and proliferation of cells. The activity of tyrosine kinases is heavily regulated and the disruption of these control mechanisms frequently leads to cancer. In this thesis I explore two different aspects of tyrosine kinase function. The first question concerns conformational changes within the kinase domain, the catalytic center of tyrosine kinases, and how these conformational changes relate to functional aspects of the protein. This work was performed with the tyrosine kinase c-Abl, a tyrosine kinase responsible for the cancer chronic myeloid leukemia. The results with c-Abl raise the possibility that many aspects of the protein's flexibility are a universal property of the protein kinase superfamily. The second question addressed in this thesis concerns the nature of substrate specificity in a particular tyrosine kinase, called C-terminal Src Kinase (Csk). This protein is the major negative regulator of another family of tyrosine kinases, the Src kinases, which it downregulates by phosphorylating on a C-terminal residue. This modification is unusually specific for tyrosine kinases, and the molecular origin of this specificity is uncovered by the structure of Csk in complex with its substrate protein, c-Src. This structure reveals a unique docking mechanism that orients the tail of c-Src towards the active site of Csk. Specificity is assured by the fact that the usual substrate binding site of tyrosine kinases, located at the base of the active site cleft, is disrupted in Csk, preventing substrates that lack the unique docking site from being phosphorylated by Csk.