Research in my laboratory is focused on using the tools of synthetic organic chemistry, biochemistry, structural biology, and genetics to gain insight into how signaling networks transmit information in normal and disease settings. Our guiding principle is to use chemistry to answer questions that cannot be addressed by the use of biochemistry or genetics alone—in this way we seek to provide tools which fill in the gaps left behind by more traditional approaches.
A major thrust of the lab is to develop chemical tools enabling the identification of the direct substrates of individual protein kinases. There are at least two challenges to identifying the direct substrates of kinases: (1) all kinases accept ATP as the phosphodonor, making distinction between individual kinases based on the cofactor ATP impossible, and (2) substrates of kinases are typically present in low abundance, making their purification and isolation from cells a challenge. We solved the first problem by developing a modified form of ATP which is only accepted by a mutated form of the kinase of interest. We have solved the second problem more recently by using thiophosphate as a tag for direct substrates through development of both antibody and chemical capture methods for isolation of the thiophosphate modified substrates in cells. Using these methods we and others have identified hundreds of direct kinase substrates.
In an attempt to develop a drug to treat Parkinson's disease we have designed a small molecule which helps protect neurons from dying due to various types of stress. Mitochondria have long been implicated in the pathogenesis of Parkinson's disease (PD). Mutations in the mitochondrial kinase PINK1 that reduce kinase activity are associated with mitochondrial defects and result in an autosomal-recessive form of early-onset PD. Therapeutic approaches for enhancing the activity of PINK1 have not been considered because no allosteric regulatory sites for PINK1 are known. We have shown that an alternative strategy, a neo-substrate approach involving the ATP analog kinetin triphosphate (KTP), can be used to increase the activity of both PD-related mutant PINK1G309D and wild-type PINK1. Discovery of neo-substrates for kinases could provide a novel modality for regulating kinase activity.
In the area of cancer, we have recently made a breakthrough by discovering a way to block the function of the GTPase, K-Ras. Somatic mutations in the K-Ras gene are the most common activating lesions found in human cancer, and are generally associated with poor response to standard therapies. Efforts to directly target this oncogene have faced difficulties due to its picomolar affinity for GTP/GDP and the absence of known allosteric regulatory sites. We developed small molecules that irreversibly bind to a common oncogenic mutant, K-RasG12C. These compounds rely on the mutant cysteine for binding and therefore do not affect the wild type protein (WT). Using crystallography, we identified a new pocket beneath the effector binding switch-II region of Ras that was not apparent in previous structures of the molecule. Our results provide structure-based validation of a novel allosteric regulatory site on Ras that is targetable in a mutant-specific manner.