Abstract
Pseudokinases often operate through functionally related enzymes and receptors. A prime example is the pseudokinase KSR (Kinase Suppressor of RAS), which can act as both an amplifier and inhibitor of members in the RAS-MAPK (Mitogen Activated Protein Kinase) signaling pathway. KSR is structurally related to the active RAF kinases over multiple domains; moreover, the pseudokinase domain of KSR forms physical and regulatory complexes with both RAF and MEK through distinct interfaces. Characterization of small molecule interactions on KSR has been used to uncover novel chemical tools and understand the mechanism of action of clinical drugs. Here, we elaborate on assays and structural methods for measuring binding at orthosteric and interfacial binding sites on KSR. These distinct small molecule pockets provide therapeutic paths for targeting KSR1 and KSR2 pseudokinases in disease, including in RAS and RAF mutant cancers.