Seminars

Chemical tools for studying how a general allosteric site regulates protein kinase function

in Seminars

When: Friday, May 31, 1 pm
Where: ENG room 203, 44 Cummington Street

Title: Chemical tools for studying how a general allosteric
site regulates protein kinase function

Authors: Justin Rettenmaier, Jack Sadowsky, Nathan Thomsen, Allison Doak,
Jim Wells

Small-molecule inhibitors of protein kinases have revolutionized biomedical
science. In the clinic, protein kinase inhibitors have ushered in the era of
personalized medicine by transforming the standard of care for a variety of
cancers. At the bench, the widespread use of protein kinase inhibitors as
research tools has significantly advanced our knowledge of the roles of
protein kinases in signaling pathways. However, the molecular mechanisms that
regulate the function of individual kinases remain poorly understood. We are
developing chemical tools to understand how an allosteric site found in many
kinases regulates their substrate specificity, subcellular localization, and
catalytic activity. In this study, we have focused on the protein kinase PDK1
where this allosteric site, known as the PIF pocket, is required to engage
many of its protein substrates.

The Wells lab previously demonstrated that PDK1 could be activated or
inhibited by directly targeting the PIF pocket with disulfide-trapped small
molecules (Sadowsky et al. 2011, PNAS). Using insights gained from this study,
we designed a high throughput screen and identified molecules that bind this
allosteric site and can activate PDK1 up to 8-fold, or that bind without
altering its kinase activity. We determined the cocrystal structure of PDK1
bound to its native peptide agonist (PIFtide) and compare this to structures
of PDK1 bound to a neutral ligand, a 2-fold activator, or an 8-fold activator.
Finally, we show that these molecules alter PDK1 substrate specificity in
vitro and in cells by inhibiting phosphorylation of some PDK1 substrates while
simultaneously stimulating phosphorylation of others. These findings raise the
possibility that the kinome holds an untapped potential for a much richer
pharmacology than first meets the eye.