Research in the McEvoy lab uses a combination of structural and biochemical approaches to address how protein structure relates to function. Two biological systems are presently being addressed.
Regulation of Intracellular Copper Levels in Bacteria
Copper is required at low levels for proper cellular function; however, excess copper can be toxic due to the production of reactive species. Elaborate systems have evolved that maintain the proper levels of copper and other metals as well. We are studying one of the two chromosomally encoded systems that is responsible for copper homeostasis in E. coli, the cus system. This system encodes a CBA transporter, which is a three component substrate/proton antiporter that crosses both the inner and outer membrane of E. coli, as well as a fourth novel periplasmic component, CusF. We are characterizing the structure of CusF by NMR spectroscopy and xray crystallography in the various metal oxidation states. We are also describing the ligand specificity, affinity, stoichiometry and coordination environment, by such techniques as isothermal titration calorimetry, fluorescence, analytical ultracentrifugation and EPR spectroscopy.
Protein-protein Interactions in Complexes Involved in Asymmetric Cell Division
In Drosophila neural stem cells, protein complexes form at opposite poles of the cell at specific times during the cell cycle. The proper localization of these complexes is crucial to generation of cellular diversity. At present, little structural information is known about these proteins and how their complexes form. Using a combination of NMR spectroscopy and biochemistry we seek to understand the complexes that are forming. In the long term, we hope to obtain a complete picture, at the molecular level, of the protein machinery specifying asymmetric localization of factors which determine cell fate in Drosophila neuroblasts, and to generalize these results in other systems involving homologous components.
Any link on the below references will take you off
of the BMCB site and to an abstract of that particular paper.
Loftin, I.R., S. Franke, S.A. Roberts, A. Weichsel, A. Heroux, W.R. Montfort, C. Rensing, and M.M. McEvoy. 2005. A novel copper-binding fold for the periplasmic copper resistance protein CusF. Biochemistry 44: 10533-10540.
Astashkin, A.V., A.M. Raitsimring, F.A. Walker, C. Rensing, and M.M. McEvoy. 2005. Characterization of the copper(II) binding site in the pink copper binding protein CusF by electron paramagnetic resonance spectroscopy. Journal of Biological Inorganic Chemistry 10: 221-230.
Dyer, C.M., M.L. Quillin, A. Campos, J. Lu, M.M. McEvoy, A.C. Hausrath, E.M. Westbrook, P. Matsumura, B.W. Matthews BW, and F.W. Dahlquist. 2004. Structure of the constitutively active double mutant CheYD13K Y106W alone and in complex with a FliM peptide. Journal of Molecular Biology 342: 1325-35.
Halkides, C.J., M.M. McEvoy, E. Casper, P. Matsumura, K. Volz, and F.W. Dahlquist. 2000. The 1.9 A resolution crystal structure of phosphono-CheY, an analogue of the active form of the response regulator, CheY. Biochemistry 39: 5280-5286.
McEvoy, M.M., A. Bren, M. Eisenbach, and F.W. Dahlquist. 1999. Identification of the binding interfaces on CheY for two of its targets, the phosphatase CheZ and the flagellar switch protein, FliM. Journal of Molecular Biology 289: 1423-1433.
McEvoy, M.M., A.C. Hausrath, G.B. Randolph, S.J. Remington, and F.W. Dahlquist. 1998. Two binding modes reveal flexibility in kinase/response regulator interactions in the bacterial chemotaxis pathway. Proceedings of the National Academy of Sciences of the United States of America 95: 7333-7338.
McEvoy, M.M., A. de la Cruz, and F.W. Dahlquist. 1997. Large modular proteins by NMR. Nature Structural Biology 4: 9.
McEvoy, M.M., and F.W. Dahlquist. 1997. Phosphohistidines in bacterial signaling. Current Opinion in Structural Biology 7: 793-797.
Zhou, H., M.M. McEvoy, D.L. Lowry, R.V. Swanson, M.I. Simon, and F.W. Dahlquist. 1996. Phosphotransfer and CheY-binding domains of the histidine autokinase CheA are joined by a flexible linker. Biochemistry 35: 433-443.
Zhou, H., M.M. McEvoy, D.L. Lowry, R.V. Swanson, M.I. Simon, and F.W. Dahlquist. 1996. Phosphotransfer and CheY-binding domains of the histidine autokinase CheA are joined by a flexible linker. Biochemistry 35: 433-443.
McEvoy, M.M., D.R. Muhandiram, L.E. Kay and F.W. Dahlquist. 1996. Structure and dynamics of a CheY-binding domain of the chemotaxis kinase CheA determined by nuclear magnetic resonance spectroscopy. Biochemistry 35: 5633-5640.
McEvoy, M.M., H. Zhou, A.F. Roth, D.F. Lowry, T.B. Morrison, L.E. Kay, and F.W. Dahlquist. 1995. Nuclear magnetic resonance assignments and global fold of a CheY-binding domain in CheA, the chemotaxis-specific kinase of Escherichia coli. Biochemistry 34: 13871-13880.
Swanson, R.V., D.F. Lowry, P. Matsumura, M.M. McEvoy, M.I. Simon, and F.W. Dahlquist. 1995. Localized perturbations in CheY structure monitored by NMR identify a CheA binding interface. Nature Structural Biology 2: 906-910.
Zaug, A.J., M.M. McEvoy, and T.R. Cech. 1993. Self-splicing of the group I intron from anabaena pre-tRNA: requirement for base-pairing of the exon in the anticodon stem. Biochemistry 32: 7946-7953.
