Structural Biology and Biophysics
Co-workers: Dr. George Hatzopoulos, Ms. Leanne Slater, Mr. Kacper Rogala, Ms. Erin Cutts, Ms. Margaret McGuire, Ms. Alison Inglis, Mr. Martin Masik
Structural biology has been immensely successful in elucidating the high-resolution structures of macromolecular compounds. The challenge we face now is assembling these components into larger units. The group focuses on two such large assemblies of medical relevance involved in malaria-induced cytoadherence and centriole duplication, respectively.
Malaria is a parasitic disease virtually absent in the developed world, but that still affects developing countries strongly. Most malaria deaths are caused by a single parasite species, Plasmodium falciparum, and many can be attributed to obstruction of small blood vessels in tissues by red blood cell clumps. Cytoadherence of infected erythrocytes is mediated by a system unique to P. falciparum that includes the PfEMP1 protein family and other parasite components.
Our research focus is on understanding how this parasite system alters erythrocyte properties including their shape, flexibility and adhesion to epithelial cells. Large protrusions, called “knobs”, develop on the erythrocyte surface and are essential for disease pathology. Yet, we do not know how “knobs” assemble or their internal structure. Important questions to be addressed include how PfEMP1 anchors to the erythrocyte cytoskeleton, how it localizes in “knobs” and how KHARP, another parasite component, alters the mechanical properties of the erythrocyte cytoskeleton.
Centrioles, the major component of animal centrosomes, must duplicate once per cell cycle in order to maintain their number. Abnormalities in this duplication cycle can lead to medical conditions such ciliopathies, male sterility and cancer. It is therefore important to understand how these structures assemble, and how their assembly is regulated by the cell. Genetic analysis uncovered a number of proteins, including SAS-6, SAS-4/CPAP and SAS-5/STIL, as being important for this process. Structural biologists need to place these components now in the context of the centriolar assembly.
Recently, we and co-workers showed that SAS-6 is the first structural component in this assembly process. SAS-6 forms a large, multimeric, wheel-like structure that acts as a framework for other centriolar proteins. We need to address now how this process is regulated, and where the remaining centriolar components fit in.
To address these questions, the laboratory uses NMR spectroscopy, X-ray crystallography and other biophysical techniques. To bridge the gaps between high-resolution structures, large assemblies and whole cells we also collaborate with groups doing electron microscopy, tomography and in vivo molecular biology.
Recent news items
Selected recent publications1. Hatzopoulos, G.N., Erat, M.C., Cutts, E., Rogala, K.B., Slater, L.M., Stansfeld, P.J., Vakonakis, I. (2013) Structural analysis of the G-Box domain of the microcephaly protein CPAP suggests a role in centriole architecture. Structure in press
2. Hilbert, M., Erat, M.C., Hachet, V., Guichard, P., Blank, I.D., Flückiger, I., Slater, L., Lowe, E.D., Hatzopoulos, G.N., Steinmetz, M.O., Gönczy, P. and Vakonakis, I. (2013) The Caenorhabditis elegans centriolar protein SAS-6 can form a spiral that is consistent with imparting a 9-fold symmetry. Proc. Natl. Acad. Sci. U.S.A. 110, 11373-8.
3. Erat, M.C., Sladek, B., Campbell, I.D. and Vakonakis, I. (2013) Structural analysis of collagen type I interactions with human fibronectin reveals a cooperative binding mode. J Biol Chem. 288, 17441-50.
4. Mayer, C., Slater, L., Erat, M.C., Konrat, R., Vakonakis, I. (2012) Structural analysis of the Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) intracellular domain reveals a conserved interaction epitope. J Biol Chem. 287, 7182-9.
5. Kitagawa, D., Vakonakis, I., Olieric, N., Hilbert, M., Keller, D., Olieric, V., Bortfeld, M., Erat, M.C., Flückiger, I., Gönczy, P., Steinmetz, M.O. (2011) Structural Basis of the 9-Fold Symmetry of Centrioles. Cell. 144, 364-75.
6. Erat, M.C., Schwarz-Linek, U., Pickford, A.R., Farndale, R.W., Campbell, I.D., Vakonakis, I. (2010) Implications for collagen binding from the crystallographic structure of fibronectin 6FnI1-2FnII7FnI. J Biol Chem. 285, 33764-70.
7. Vakonakis, I., Staunton, D., Ellis, I.R., Sarkies, P., Flanagan, A., Schor, A.M., Schor, S.L. and Campbell, I.D. (2009) Motogenic sites in human fibronectin are masked by long range interactions. J Biol Chem. 284, 15668-75.
8. Erat, M.C., Slatter, D.A., Lowe, E.D., Millard, C.J., Farndale, R.W., Campbell, I.D. and Vakonakis, I. (2009) Identification and structural analysis of type I collagen sites in complex with fibronectin fragments. Proc. Natl. Acad. Sci. U.S.A. 106, 4195-200.
Cartoon showing how the nine-fold symmetric cartwheel model determines the overall symmetry of centrioles, drawn as ensembles of microtubule blades. An electron micrograph of C. reinhardtii SAS-6 protein with clearly visible ring structures is shown in the background.
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