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Non-Equilibrium Molecular Dynamics to Simulate Shear Stress on Angiotensin II Type 1 (AT1) Receptor
Monday, February 29, 2016
325a
that visualizes kinetic features from both simulation and experiment in terms of peaks with specific amplitude and timescale, to identify the residues that are most strongly coupled to distinct activation pathways and suggest labeling sites for future kinetic experiments.
molecular dynamics (aMD) which confirmed that aMD spaned more conformational space than cMD and gMD. Priliminary analysis showed that those predicted residues sites could be a binding pocket for the drugable small molecules.
1596-Pos Board B573 Potential of Mean Force Calculations and Isothermal Titration Calorimetry Measurements of the Human Cardiac Troponin C / Calcium Interaction Reveal Affinity Changes as a Function of Familial Hypertrophic Cardiomyopathy Associated Mutations Charles M. Stevens1,2, Kaveh Rayani1, Gurpreet Singh3, D. Peter Tieleman3, Glen F. Tibbits1,2. 1 Biomedical Physiology & Kinesiology, Simon Fraser University, Burnaby, BC, Canada, 2Cardiovascular Science, Child & Family Research Institute, Vancouver, BC, Canada, 3Biological Science, University of Calgary, Calgary, AB, Canada. Cardiac muscle contraction is regulated by the calcium-binding component of the Troponin Complex, cardiac Troponin C (cTnC), in response to an increase in cytosolic calcium levels. Interaction with calcium is followed by a conformational change, which alters the interactions between troponin complex proteins and mediates the interaction between the thin and thick filaments. Mutations that affect the ability of cTnC to bind calcium are hypothesized to increase or decrease calcium affinity, and thereby predicted to induce hypertrophic or dilated cardiomyopathies, respectively. Cardiomyopathy-associated mutations in the N-Domain of cTnC were selected for this study. These mutations include A8V, L29Q, A31S, L48Q, and C84Y. Equilibrium molecular dynamics simulations were used to model the structural effects of these mutations, and their functional impact assessed using Potential of Mean Force (PMF) calculations, and Isothermal Titration Calorimetry (ITC). Very little structural variation was observed in the mutant protein models. Despite this, large differences in the free energy of calcium binding were observed as a function of these sequence substitutions. The results from ITC are, in some cases, at odds with those derived from MD simulations. These differences may be due to the effect of the mutations on the energetics of the transition between the open and closed conformation of the N-cTnC molecule that are not accessible to the timescales of the MD simulations, but will be readily measured by ITC. Taken together, these results demonstrate that mutations associated with Familial Hypertrophic Cardiomyopathy can act through the alterations in the thermodynamic properties of calcium binding and conformational changes.
1599-Pos Board B576 Conformational Changes in Antigen-Antibody Binding: Molecular Dynamics Study Keiko Shinoda, Hideaki Fujitani. LSBM, RCAST, Univ. of Tokyo, Tokyo, Japan. Proline cis-trans isomerization have emerged as an effective regulatory mechanism in a wide range of biological processes related to Alzheimer disease and cancer. However, the details of the mechanism still remain to be clarified. The cis-trans isomerization has been observed in an antigenantibody system, epiregulin (EPR) and its antibody by X-ray crystallography. Anti EPR antibody, 9E5 has a proline at the residue 103 in the third complementarity-determining region of the heavy chain (CDR-H3), which is in the cis conformation for EPR free 9E5 (apo 9E5) and the trans one for the complex of EPR-9E5 obtained from X-ray crystallography (we refer this complex as ‘‘trans-Complex’’). Because the cis-trans isomerization is known to occur slowly, with the duration ranging from several minutes to hours, we investigated whether EPR forms a stable complex with apo 9E5 before the isomerization occurs. To identify a stable EPR-binding structure with apo 9E5, we conducted extensive binding molecular dynamics (MD) simulations and additional long MD simulations. An EPR-9E5 complex of which the bound structure is highly similar to that of the trans-Complex was found. We refer this complex as ‘‘cis-Complex’’. We then investigated the structural stability and conformational changes of the cis-Complex. The cis-Complex is stable for at least 4 ms. It is found that the CDR-H3 loop plays important role for the EPR binding process in which the CDR-H3 loop interacts with EPR strongly. The interaction energy between the EPR and 9E5 is lower than that for the trans-Complex by more than 100 kJ/mol. From 10 ms-MD simulations of the cis-Complex, it is also found that the slight conformal changes of the Pro103 occur slowly during 10 ms.
1597-Pos Board B574 Computational Evaluation of Mutational Effects on Kinase Dynamics Mohammad M. Sultan, Vijay Pande. Chemistry, Stanford University, Stanford, CA, USA. Kinases are enzymes that control a large number of biochemical pathways and their misregulation, in particular via mutations, is linked to a number of diseases including cancers. In this Molecular Dynamics (MD) study, we investigate the effect of a large number of mutations on the dynamics of protein kinases including the Src kinase family. We run long time-scale MD simulations and combine the dynamics from various mutants into a single Markov state model(MSM). We then use our MD models to investigate the atomistic effects these mutations have on dynamics. We discover several intermediate states within our ensembles including potentially druggable conformations. We also find that the effects of some of these mutations on the conformational landscape can be captured by our MSMs. 1598-Pos Board B575 Dynamics of C-terminus Motion of Norwalk Virus Capsid by Molecular Dynamics (All-Atom & Coarse Grained) Simulation Mahendra B. Thapa1, Jarek Meller2, Mark Rance3. 1 Physics Department, University of Cincinnati, Cincinnati, OH, USA, 2 Depts. of Environmental Health and Biomedical Eng, University of Cincinnati, Cincinnati, OH, USA, 3Dept. of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati, Cincinnati, OH, USA. Dynamics of C-terminus arginine residues of Norwalk Virus capsid were investigated by molecular dynamics (All atom and Coarse grained) simulations. AMBER simulation suit 12 with the use of GPU card was employed for the simulation and the online server CABS Flex server was used for the coarse grained simulation.It was observed that these arginine residues interacted with specified residues of the capsid protein. Most of the interacting residues predicted by conventional molecular dynamics (cMD) and coarse grained molecualar dynamics (gMD) were subset of residues predicted by accelerated
1600-Pos Board B577 Non-Equilibrium Molecular Dynamics to Simulate Shear Stress on Angiotensin II Type 1 (AT1) Receptor Matheus Malta de Sa1, Silvestre Massimo Modestia2, Carlota Oliveira Rangel-Yagui2, Jose´ Eduardo Krieger1. 1 Heart Institute, University of Sao Paulo, Sao Paulo, Brazil, 2Department of Biochemical and Pharmaceutical Technology, School of Pharmaceutical Sciences, University of Sao Paulo, Sao Paulo, Brazil. AT1 is a G protein-coupled receptor responsible for controlling blood pressure in the cardiovascular system. The binding of angiotensin II (AngII) causes Gq recruitment, while shear stress (SS) activates AT1 and causes ERK1/2 phosphorylation independently of Gaq. In order to investigate the latter, atomistic non-equilibrium molecular dynamics simulations explored AT1 conformational changes under SS. The complete structure of AT1 inserted in a POPC bilayer and solvated in water, was simulated for 1 ms at 1 bar and 310 K. SS was applied deforming the simulation box separately in the x and y directions for 100 ns, at a rate of 1 pm/ps. AT1 without SS presents increased flexibility in the intracellular loop (ICL) 2, ICL 3, and helix 8. The root mean square fluctuation of the C-terminal (Leu317-Glu359) is higher without SS (0.61-1.64 nm) versus SS (0.32-0.76 nm). The radius of gyration of the C-terminal is smaller without SS (1.04-1.19 nm) compared to SS (1.36-1.40 nm), suggesting this region is more extended under SS. C-terminal is also more exposed to water in SS, as the average solvent accessible surface area is higher (44.55 2.2 nm2) compared to AT1 without SS (38.551.6 nm2), including the residues prone to phosporylation by GRK (Thr332-Ser338). Collectively, the findings suggest SS exposes and makes the C-terminal more stable to GRK binding, leading to b-arrestin recruitment and, ultimately, ERK1/2 phosphorylation. 1601-Pos Board B578 Computational Study on Flexible Dynamics of Histone Tails Sotaro Fuchigami. Yokohama City University, Yokohama, Japan. Post-translational modifications of histone tails regulate DNA transcription in eukaryotic cells. The flexibility of the histone tails is considered to be crucial for the transcription regulation, but details of their molecular mechanism remain unclear. In the present study, we characterized conformational dynamics of flexible histone tails using molecular dynamics (MD) simulations. We selected histone tails of H2A/H2B dimer as targets, and performed one microsecond MD simulations in explicit water using MARBLE and the
325a
that visualizes kinetic features from both simulation and experiment in terms of peaks with specific amplitude and timescale, to identify the residues that are most strongly coupled to distinct activation pathways and suggest labeling sites for future kinetic experiments.
molecular dynamics (aMD) which confirmed that aMD spaned more conformational space than cMD and gMD. Priliminary analysis showed that those predicted residues sites could be a binding pocket for the drugable small molecules.
1596-Pos Board B573 Potential of Mean Force Calculations and Isothermal Titration Calorimetry Measurements of the Human Cardiac Troponin C / Calcium Interaction Reveal Affinity Changes as a Function of Familial Hypertrophic Cardiomyopathy Associated Mutations Charles M. Stevens1,2, Kaveh Rayani1, Gurpreet Singh3, D. Peter Tieleman3, Glen F. Tibbits1,2. 1 Biomedical Physiology & Kinesiology, Simon Fraser University, Burnaby, BC, Canada, 2Cardiovascular Science, Child & Family Research Institute, Vancouver, BC, Canada, 3Biological Science, University of Calgary, Calgary, AB, Canada. Cardiac muscle contraction is regulated by the calcium-binding component of the Troponin Complex, cardiac Troponin C (cTnC), in response to an increase in cytosolic calcium levels. Interaction with calcium is followed by a conformational change, which alters the interactions between troponin complex proteins and mediates the interaction between the thin and thick filaments. Mutations that affect the ability of cTnC to bind calcium are hypothesized to increase or decrease calcium affinity, and thereby predicted to induce hypertrophic or dilated cardiomyopathies, respectively. Cardiomyopathy-associated mutations in the N-Domain of cTnC were selected for this study. These mutations include A8V, L29Q, A31S, L48Q, and C84Y. Equilibrium molecular dynamics simulations were used to model the structural effects of these mutations, and their functional impact assessed using Potential of Mean Force (PMF) calculations, and Isothermal Titration Calorimetry (ITC). Very little structural variation was observed in the mutant protein models. Despite this, large differences in the free energy of calcium binding were observed as a function of these sequence substitutions. The results from ITC are, in some cases, at odds with those derived from MD simulations. These differences may be due to the effect of the mutations on the energetics of the transition between the open and closed conformation of the N-cTnC molecule that are not accessible to the timescales of the MD simulations, but will be readily measured by ITC. Taken together, these results demonstrate that mutations associated with Familial Hypertrophic Cardiomyopathy can act through the alterations in the thermodynamic properties of calcium binding and conformational changes.
1599-Pos Board B576 Conformational Changes in Antigen-Antibody Binding: Molecular Dynamics Study Keiko Shinoda, Hideaki Fujitani. LSBM, RCAST, Univ. of Tokyo, Tokyo, Japan. Proline cis-trans isomerization have emerged as an effective regulatory mechanism in a wide range of biological processes related to Alzheimer disease and cancer. However, the details of the mechanism still remain to be clarified. The cis-trans isomerization has been observed in an antigenantibody system, epiregulin (EPR) and its antibody by X-ray crystallography. Anti EPR antibody, 9E5 has a proline at the residue 103 in the third complementarity-determining region of the heavy chain (CDR-H3), which is in the cis conformation for EPR free 9E5 (apo 9E5) and the trans one for the complex of EPR-9E5 obtained from X-ray crystallography (we refer this complex as ‘‘trans-Complex’’). Because the cis-trans isomerization is known to occur slowly, with the duration ranging from several minutes to hours, we investigated whether EPR forms a stable complex with apo 9E5 before the isomerization occurs. To identify a stable EPR-binding structure with apo 9E5, we conducted extensive binding molecular dynamics (MD) simulations and additional long MD simulations. An EPR-9E5 complex of which the bound structure is highly similar to that of the trans-Complex was found. We refer this complex as ‘‘cis-Complex’’. We then investigated the structural stability and conformational changes of the cis-Complex. The cis-Complex is stable for at least 4 ms. It is found that the CDR-H3 loop plays important role for the EPR binding process in which the CDR-H3 loop interacts with EPR strongly. The interaction energy between the EPR and 9E5 is lower than that for the trans-Complex by more than 100 kJ/mol. From 10 ms-MD simulations of the cis-Complex, it is also found that the slight conformal changes of the Pro103 occur slowly during 10 ms.
1597-Pos Board B574 Computational Evaluation of Mutational Effects on Kinase Dynamics Mohammad M. Sultan, Vijay Pande. Chemistry, Stanford University, Stanford, CA, USA. Kinases are enzymes that control a large number of biochemical pathways and their misregulation, in particular via mutations, is linked to a number of diseases including cancers. In this Molecular Dynamics (MD) study, we investigate the effect of a large number of mutations on the dynamics of protein kinases including the Src kinase family. We run long time-scale MD simulations and combine the dynamics from various mutants into a single Markov state model(MSM). We then use our MD models to investigate the atomistic effects these mutations have on dynamics. We discover several intermediate states within our ensembles including potentially druggable conformations. We also find that the effects of some of these mutations on the conformational landscape can be captured by our MSMs. 1598-Pos Board B575 Dynamics of C-terminus Motion of Norwalk Virus Capsid by Molecular Dynamics (All-Atom & Coarse Grained) Simulation Mahendra B. Thapa1, Jarek Meller2, Mark Rance3. 1 Physics Department, University of Cincinnati, Cincinnati, OH, USA, 2 Depts. of Environmental Health and Biomedical Eng, University of Cincinnati, Cincinnati, OH, USA, 3Dept. of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati, Cincinnati, OH, USA. Dynamics of C-terminus arginine residues of Norwalk Virus capsid were investigated by molecular dynamics (All atom and Coarse grained) simulations. AMBER simulation suit 12 with the use of GPU card was employed for the simulation and the online server CABS Flex server was used for the coarse grained simulation.It was observed that these arginine residues interacted with specified residues of the capsid protein. Most of the interacting residues predicted by conventional molecular dynamics (cMD) and coarse grained molecualar dynamics (gMD) were subset of residues predicted by accelerated
1600-Pos Board B577 Non-Equilibrium Molecular Dynamics to Simulate Shear Stress on Angiotensin II Type 1 (AT1) Receptor Matheus Malta de Sa1, Silvestre Massimo Modestia2, Carlota Oliveira Rangel-Yagui2, Jose´ Eduardo Krieger1. 1 Heart Institute, University of Sao Paulo, Sao Paulo, Brazil, 2Department of Biochemical and Pharmaceutical Technology, School of Pharmaceutical Sciences, University of Sao Paulo, Sao Paulo, Brazil. AT1 is a G protein-coupled receptor responsible for controlling blood pressure in the cardiovascular system. The binding of angiotensin II (AngII) causes Gq recruitment, while shear stress (SS) activates AT1 and causes ERK1/2 phosphorylation independently of Gaq. In order to investigate the latter, atomistic non-equilibrium molecular dynamics simulations explored AT1 conformational changes under SS. The complete structure of AT1 inserted in a POPC bilayer and solvated in water, was simulated for 1 ms at 1 bar and 310 K. SS was applied deforming the simulation box separately in the x and y directions for 100 ns, at a rate of 1 pm/ps. AT1 without SS presents increased flexibility in the intracellular loop (ICL) 2, ICL 3, and helix 8. The root mean square fluctuation of the C-terminal (Leu317-Glu359) is higher without SS (0.61-1.64 nm) versus SS (0.32-0.76 nm). The radius of gyration of the C-terminal is smaller without SS (1.04-1.19 nm) compared to SS (1.36-1.40 nm), suggesting this region is more extended under SS. C-terminal is also more exposed to water in SS, as the average solvent accessible surface area is higher (44.55 2.2 nm2) compared to AT1 without SS (38.551.6 nm2), including the residues prone to phosporylation by GRK (Thr332-Ser338). Collectively, the findings suggest SS exposes and makes the C-terminal more stable to GRK binding, leading to b-arrestin recruitment and, ultimately, ERK1/2 phosphorylation. 1601-Pos Board B578 Computational Study on Flexible Dynamics of Histone Tails Sotaro Fuchigami. Yokohama City University, Yokohama, Japan. Post-translational modifications of histone tails regulate DNA transcription in eukaryotic cells. The flexibility of the histone tails is considered to be crucial for the transcription regulation, but details of their molecular mechanism remain unclear. In the present study, we characterized conformational dynamics of flexible histone tails using molecular dynamics (MD) simulations. We selected histone tails of H2A/H2B dimer as targets, and performed one microsecond MD simulations in explicit water using MARBLE and the