Structural biology, NMR spectroscopy, protein structure and dynamics, structure-based drug design
Our lab is fundamentally interested in understanding, from a structural and biophysical perspective, the functioning of proteins involved in regulating transcription, particularly those involved in the dysregulation associated with the development of cancer. Structural and functional characterization of the native forms of these proteins and their relevant complexes via NMR spectroscopy, X-ray crystallography, and a variety of other techniques provides a baseline of understanding. Subsequent characterization of the oncoprotein forms then provides a detailed understanding of the molecular mechanism of oncogenesis associated with altered forms of these proteins. Such knowledge leads to novel avenues for the design of therapeutic agents to treat the cancers associated with these particular oncoproteins.
One current focus is structural studies of a novel transcriptional enhancer referred to as the core-binding factor (CBF). This heterodimeric protein is essential for hematopoietic development. Gene translocations associated with the genes coding for the two subunits of CBF produce novel fusion proteins which have been implicated as playing a role in more than 30% of acute leukemias. We are carrying structural (NMR spectroscopy and X-ray crystallography) and functional studies of the oncoprotein forms of the two subunits of CBF that are associated with leukemia to gain an understanding of their roles in the development of leukemia. A second area of focus in this area is on fusion proteins involving the transcription factor MLL, which are implicated in a very high percentage of pediatric leukemias.
A second focus is the development of highly targeted small molecule inhibitors of the oncoprotein forms of CBF and MLL. Using structural information on the proteins, virtual screening, NMR and fluorescence-based assays, and medicinal chemistry, we have developed the first small molecule inhibitors of these proteins. This is a collaborative effort with outside investigators at Harvard, University of Pennsylvania, and NIH which has been supported by a Specialized Center of Research (SCOR) grant from the Leukemia and Lymphoma Society.
A third focus for the lab is the application of solution NMR methods to the structure determination of membrane proteins. The vast majority of drug targets are membrane- embedded proteins. This class of proteins has presented significant challenges for structure determination by any method. We completed the structure determination of the largest helical membrane protein to be solved by NMR spectroscopy thus far. This structure established a paradigm for tackling this class of proteins by solution NMR. We are currently examining additional technical improvements in this area as well as targeting several new systems for structure determination.
Application of fragment-based drug discovery to membrane proteins: identification of ligands of the integral membrane enzyme DsbB. Früh V, Zhou Y, Chen D, Loch C, Ab E, Grinkova YN, Verheij H, Sligar SG, Bushweller JH, Siegal G. Chem Biol. 17:881-91 (2010).
Structure of the AML1-ETO NHR3-PKA(RIIalpha) Complex and Its Contribution to AML1-ETO Activity. Corpora T, Roudaia L, Oo ZM, Chen W, Manuylova E, Cai X, Chen MJ, Cierpecki T, Speck NA, Bushweller JH. J Mol Biol. [Epub ahead of print] (2010).
The PHD3 domain of MLL acts as a CYP33-regulated switch between MLL-mediated activation and repression. Park S, Osmers U, Raman G, Schwantes RH, Diaz MO, Bushweller JH. Biochemistry. 49:6576-86 (2010).
Structure of the MLL CXXC domain-DNA complex and its functional role in MLL-AF9 leukemia. Cierpicki T, Risner LE, Grembecka J, Lukasik SM, Popovic R, Omonkowska M, Shultis DD, Zeleznik-Le NJ, Bushweller JH. Nat Struct Mol Biol. 17, 62-68 (2010).
The role of CBFbeta in AML1-ETO’s activity. Park S, Speck NA, Bushweller JH. Blood. 114, 2849-50 (2009).
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