Understanding the structural and dynamical basis of allosteric Signaling across the membrane

Although kinases are believed to function as signaling switches, they are not turned on and off by one element. Rather, an intricate circuitry of intra- and inter-molecular allosteric networks in the regulatory and the catalytic modules control the kinase activity. It is not known how these non-covalent interactions influence the transition from an inactive state to an active state. The mechanism of allosteric regulation of the kinase is further complicated by the emerging role of plasma membrane. The long-term goal of my laboratory will be to establish interdisciplinary research focused toward understanding the general mechanism in allosteric regulation of kinases, particularly at the membrane.  In a recent study with epidermal growth factor receptor (EGFR), we found that EGF-binding to the receptor allows the association of the transmembrane helices that in turn reorients the juxtamembrane segment. This coupled structural rearrangement peels the intracellular module off the membrane allowing the formation of an active EGFR dimer. While these studies have explained the mechanism of the structural coupling between the extracellular and intracellular modules, it is not clear if this mechanism of transmembrane-juxtamembrane coupling is applicable to other members of receptor tyrosine kinases such as Vascular Endothelial Growth Factor Receptor (VEGFR). The VEGFR kinases are popular targets for treating many forms of cancer, but the structural and functional biology of these receptors is not fully understood. Although VEGFR is also activated by ligand dependent dimerization, several features of this receptor kinase make it unique. In contrast to the mammalian system, Plasmodium falciparum, a protozoan parasite, has no known receptor kinase. How do protozoa transmit extracellular signals across the membrane? Several non-receptor Ser/Thr kinases, such as pfPK9 and calcium dependent kinases, were shown to localize at the boundary of host RBC membrane and parasitophorous vacuolar membrane (PVM) during different stages of Plasmodium life cycle. It was speculated that such membrane localization might be essential for transducing signals across the host and parasite membranes. However, functional and structural details of such a mechanism have not yet been established. In this presentation I will discuss my research proposal to study the VEGFR and pfPK9 kinases as model systems to investigate signaling at the membrane.  I will approach this problem by using a combination of structural biology techniques such as NMR spectroscopy and crystallography, biochemistry, molecular biology and single cell microscopy.