Kinase inhibitors are important cancer drugs, but they tend to display limited
target specificity, and their target profiles are often challenging to
rationalize in terms of molecular mechanism. Here we report that the clinical
kinase inhibitor bosutinib recognizes its kinase targets by engaging a pair of
conserved structured water molecules in the active site and that many other
kinase inhibitors share a similar recognition mechanism. Using the nitrile group
of bosutinib as an infrared probe, we show that the gatekeeper residue and one
other position in the ATP-binding site control access of the drug to the
structured water molecules and that the amino acids found at these positions
account for the kinome-wide target spectrum of the drug. Our work highlights the
importance of structured water molecules for inhibitor recognition, reveals a
new role for the kinase gatekeeper and showcases an effective approach for
elucidating the molecular origins of selectivity patterns.
