研究领域

Biological and Physical Chemistry

The long term goal of our research is to understand how the environment of biological membrane affects function of proteins residing in it. Membrane proteins represent about 30% of all proteome and perform essential functions, including energy conversion, signal transduction, and chemical transport in and out of cells. Numerous diseases are associated with reduction or loss of function in membrane proteins, and many pathogens exploit membrane receptors to facilitate entrance inside the cells. About 60% of approved drugs on the market act on membrane proteins. Rational design of new drugs with better efficacy and selectivity requires knowledge of a detailed 3-dimensional structure of the target protein at atomic resolution, however, the number of solved membrane protein structures is far smaller than that of soluble proteins. Success in crystallization of eukaryotic and, in even larger degree, of human membrane proteins is limited by the challenges in heterologous expression of functional proteins and by the low stability of these proteins when purified in detergent solution. Both of these problems relate to intimately more complex nature of biological membranes of higher organisms. Multiple lipid components and intrinsic heterogeneity of biological membranes provide tight control of function and selective stabilization of embedded proteins. One of the main players in maintaining heterogeneity in biological membranes is cholesterol, an ubiquitous lipid in eukaryotic cells. Activities of a large number of proteins from different families were found to be directly modulated by cholesterol, yet there is no clear understanding of a structural mechanism of cholesterol action on membrane proteins. In our research we use lipidic cubic phase (LCP) as a primary tool to study lipid/protein interactions and to crystallize membrane proteins directly from the lipidic environment. Our targets include G protein-coupled receptors (GPCR), ion channels and transporters. In order to achieve our goals we are developing an LCP-toolchest: collection of assays and methods to interrogate behavior of proteins and their interaction with lipids in LCP. In addition to providing insights into the lipid/protein interactions these assays implement convenient metrics to guide protein engineering, selection of ligands and lipids for improving chances for crystallization and expanding high-resolution structural coverage of elusive human membrane proteins.

近期论文

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G. Fenalti, N.A. Zatsepin, C. Betti, P. Giguere, G.W. Han, A. Ishchenko, W. Liu, K. Guillemyn, H. Zhang, D. James, D. Wang, U. Weierstall, J.C.H. Spence, S. Boutet, M. Messerschmidt, G.J. Williams, C. Gati, O.M. Yefanov, T.A. White, D. Oberthuer, M. Metz, C.H. Yoon, A. Barty, H.N. Chapman, S. Basu, J. Coe, C.E. Conrad, R. Fromme, P. Fromme, D. Tourwe, P.W. Schiller, B.L. Roth, S. Ballet, V. Katritch, R.C. Stevens, V. Cherezov (2015) Structural basis for bifunctional peptide recognition at human delta-opioid receptor. Nat Struct Mol Biol 22: 265-268; PDB ID: 4RWA, 4RWD L. Qin, I. Kufareva, L.G. Holden, C. Wang, Y. Zheng, C. Zhao, G. Fenalti, H. Wu, G.W. Han, V. Cherezov, R. Abagyan, R.C. Stevens, T.M. Handel (2015) Crystal structure of the chemokine receptor CXCR4 in complex with a viral chemokine. Science 347: 1117-1122; PDB ID: 4RWS P. Nogly, D. James, D. Wang, T.A. White, N. Zatsepin, A. Shilova, G. Nelson, H. Liu, L. Johansson, M. Heymann, K. Jaeger, M. Metz, C. Wickstrand, W. Wu, P. Bath, P. Berntsen, D. Oberthuer, V. Panneels, V. Cherezov, H. Chapman, G. Schertler, R. Neutze, J. Spence, I. Moraes, M. Burghammer, J. Standfuss, U. Weierstall (2015) Lipidic cubic phase serial millisecond crystallography using synchrotron radiation. IUCrJ 2: 168-176; PDB ID: 4X31, 4X32 V. Isberg, C. de Graaf, A. Bortolato, V. Cherezov, V. Katritch, F.H. Marshall, S. Mordalski, J. Pin, R.C. Stevens, G. Vriend, D.E. Gloriam (2015) Generic GPCR residue numbers - aligning topology maps while minding the gaps. Trends Pharmacol Sci 36: 22-31; D.L. Gater, O. Saurel, I. Iordanov, W. Liu, V. Cherezov, A. Milon (2014) Two classes of cholesterol binding sites for the b2AR revealed by thermostability and NMR Biophysics J 107: 2305-2312; M.A. Goren, T. Morizumi, I. Menon, J.S. Joseph, J.S. Dittman, V. Cherezov, R.C. Stevens, O.P. Ernst, A.K. Menon (2014) Constitutive phospholipid scramblase activity of a G protein-coupled receptor. Nat Commun 5: 5115; W. Liu, A. Ishchenko, V. Cherezov (2014) Preparation of microcrystals in lipidic cubic phase for serial femtosecond crystallography. Nat Protoc 9: 2123-2134; C. Wang, H. Wu, T. Evron, E. Vardy, G.W. Han, X.P., Huang, S.J. Hufeisen, T.J. Mangano, D.J. Urban, V. Katritch, V. Cherezov, M.G. Caron, B.L. Roth, R.C. Stevens (2014) Structural basis for Smoothened receptor modulation and chemoresistance to anticancer drugs. Nat Commun 5: 4355; PDB ID: 4N4W, 4QIN, 4QIM W. Liu, D. Wacker, C. Wang, E. Abola, V. Cherezov (2014) Femtosecond crystallography of membrane proteins in the lipidic cubic phase. Phil Trans R Soc B 369: 20130314; A. Ishchenko, E. Abola, V. Cherezov (2014) Lipidic Cubic Phase Technologies for Structural Studies of Membrane Proteins. in Membrane Protein Production for Structural Analysis, ed. I. Mus-Veteau, pages 289-314 (Springer); J. Zhang, K. Zhang, Z.G. Gao, S. Paoletta, D. Zhang, G.W. Han, T. Li, L. Ma, W. Zhang, C.E. Muller, H. Yang, H. Jiang, V. Cherezov, V. Katritch, K.A. Jacobson, R.C. Stevens, B. Wu, Q.Zhao (2014) Agonist-bound structure of the human P2Y12 receptor. Nature 509: 119-122; PDB ID: 4PXZ, 4PY0 K. Zhang, J. Zhang, Z.G. Gao, D. Zhang, L. Zhu, G.W. Han, S.M. Moss, S. Paoletta, E. Kiselev, W. Lu, G. Fenalti, W. Zhang, C.E. Muller, H. Yang, H. Jiang, V. Cherezov, V. Katritch, K.A. Jacobson, R.C. Stevens, B. Wu, Q.Zhao (2014) Structure of the human P2Y12 receptor in complex with an antithrombotic drug. Nature 509: 115-118; PDB ID: 4NTJ V. Katritch, G. Fenalti, E.E. Abola, B.L. Roth, V. Cherezov, R.C. Stevens (2014) Allosteric sodium in class A GPCR signaling. Trends Biochem Sci 39: 233-244; H. Wu, C. Wang, K.J. Gregory, G.W. Han, H.P. Cho, Y. Xia, C.M. Niswender, V. Katritch, J. Meiler, V. Cherezov, P.J. Conn, R.C. Stevens (2014) Structure of a class C GPCR metabotropic glutamate receptor 1 bound to an allosteric modulator. Science 344: 58-64; PDB ID: 4OR2 U. Weierstall, D. James, C. Wang, T.A. White, D. Wang, W. Liu, J.C. Spence, R.B. Doak, G. Nelson, P. Fromme, R. Fromme, I. Grotjohann, C. Kupitz, N.A. Zatsepin, H. Liu, S. Basu, D. Wacker, G.W. Han, V. Katritch, S. Boutet, M. Messerschmidt, G.J. Williams, J.E. Koglin, M.M. Seibert, M. Klinker, C. Gati, R.L., Shoeman, A. Barty, H.N. Chapman, R.A. Kirian, K.R. Beyerlein, R.C. Stevens, D. Li, S.T. Shah, N. Howe, M. Caffrey, V. Cherezov (2014) Lipidic cubic phase injector facilitates membrane protein serial femtosecond crystallography. Nat Commun 5: 3309; PDB ID: 4O9R G. Fenalti, P.M. Giguere, V. Katritch, X.P. Huang, A.A. Thompson, V. Cherezov, B.L. Roth, R.C. Stevens (2014) Molecular control of delta-opioid receptor signalling. Nature 506: 191-196; PDB ID: 4N6H K. Niescierowicz, L. Caro, V. Cherezov, M. Vivaudou, C.J. Moreau (2014) Functional assay for T4 lysozyme-engineered G protein-coupled receptors with an ion channel reporter. Structure 22: 149-155;