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Garnier Norbert


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Maître de conférence

Groupe thématique : Réparation de l’ADN : structure, fonction et dynamique

Publications

2016   Références trouvées : 1

Jacob, L. Sawma, P. Garnier, N. Meyer, L. A. Fritz, J. Hussenet, T. Spenle, C. Goetz, J. Vermot, J. Fernandez, A. Baumlin, N. Aci-Seche, S. Orend, G. Roussel, G. Cremel, G. Genest, M. Hubert, P. Bagnard, D.  (2016)

Inhibition of PlexA1-mediated brain tumor growth and tumor-associated angiogenesis using a transmembrane domain targeting peptide

Oncotarget (2016) 7 (36) 57851-57865 - doi : 10.18632/oncotarget.11072
The neuropilin-plexin receptor complex regulates tumor cell migration and proliferation and thus is an interesting therapeutic target. High expression of neuropilin-1 is indeed associated with a bad prognosis in glioma patients. Q-RTPCR and tissue-array analyses showed here that Plexin-A1 is highly expressed in glioblastoma and that the highest level of expression correlates with the worse survival of patients. We next identified a developmental and tumor-associated pro-angiogenic role of Plexin-A1. Hence, by using molecular simulations and a two-hybrid like assay in parallel with biochemical and cellular assays we developed a specific Plexin-A1 peptidic antagonist disrupting transmembrane domain-mediated oligomerization of the receptor and subsequent signaling and functional activity. We found that this peptide exhibits anti-tumor activity in vivo on different human glioblastoma models including glioma cancer stem cells. Thus, screening Plexin-A1 expression and targeting Plexin-A1 in glioblastoma patients exhibit diagnostic and therapeutic value.

The neuropilin-plexin receptor complex regulates tumor cell migration and proliferation and thus is an interesting therapeutic target. High expression of neuropilin-1 is indeed associated with a bad prognosis in glioma patients. Q-RTPCR and tissue-array analyses showed here that Plexin-A1 is highly expressed in glioblastoma and that the highest level of expression correlates with the worse survival of patients. We next identified a developmental and tumor-associated pro-angiogenic role of Plexin-A1. Hence, by using molecular simulations and a two-hybrid like assay in parallel with biochemical and cellular assays we developed a specific Plexin-A1 peptidic antagonist disrupting transmembrane domain-mediated oligomerization of the receptor and subsequent signaling and functional activity. We found that this peptide exhibits anti-tumor activity in vivo on different human glioblastoma models including glioma cancer stem cells. Thus, screening Plexin-A1 expression and targeting Plexin-A1 in glioblastoma patients exhibit diagnostic and therapeutic value.


2014   Références trouvées : 3

Ziani, W., Maillard, A. P., Petit-Hartlein, I., Garnier, N., Crouzy, S., Girard, E. and Coves, J.  (2014)

The X-ray Structure of NccX from Cupriavidus metallidurans 31A Illustrates Potential Dangers of Detergent Solubilization When Generating and Interpreting Crystal Structures of Membrane Proteins

The Journal of Biological Chemistry (2014) 289 (45) 31130-31172 - doi : 10.1074/jbc.M114.586537
The x-ray structure of NccX, a type II transmembrane metal sensor, from Cupriavidus metallidurans 31A has been determined at a resolution of 3.12 A. This was achieved after solubilization by dodecylphosphocholine and purification in the presence of the detergent. NccX crystal structure did not match the model based on the extensively characterized periplasmic domain of its closest homologue CnrX. Instead, the periplasmic domains of NccX appeared collapsed against the hydrophobic transmembrane segments, leading to an aberrant topology incompatible with membrane insertion. This was explained by a detergent-induced redistribution of the hydrophobic interactions among the transmembrane helices and a pair of hydrophobic patches keeping the periplasmic domains together in the native dimer. Molecular dynamics simulations performed with the full-length protein or with the transmembrane segments were used along with in vivo homodimerization assays (TOXCAT) to evaluate the determinants of the interactions between NccX protomers. Taken as a whole, computational and experimental results are in agreement with the structural model of CnrX where a cradle-shaped periplasmic metal sensor domain is anchored into the inner membrane by two N-terminal helices. In addition, they show that the main determinant of NccX dimerization is the periplasmic soluble domain and that the interaction between transmembrane segments is highly dynamic. The present work introduces a new crystal structure for a transmembrane protein and, in line with previous studies, substantiates the use of complementary theoretical and in vivo investigations to rationalize a three-dimensional structure obtained in non-native conditions.

The x-ray structure of NccX, a type II transmembrane metal sensor, from Cupriavidus metallidurans 31A has been determined at a resolution of 3.12 A. This was achieved after solubilization by dodecylphosphocholine and purification in the presence of the detergent. NccX crystal structure did not match the model based on the extensively characterized periplasmic domain of its closest homologue CnrX. Instead, the periplasmic domains of NccX appeared collapsed against the hydrophobic transmembrane segments, leading to an aberrant topology incompatible with membrane insertion. This was explained by a detergent-induced redistribution of the hydrophobic interactions among the transmembrane helices and a pair of hydrophobic patches keeping the periplasmic domains together in the native dimer. Molecular dynamics simulations performed with the full-length protein or with the transmembrane segments were used along with in vivo homodimerization assays (TOXCAT) to evaluate the determinants of the interactions between NccX protomers. Taken as a whole, computational and experimental results are in agreement with the structural model of CnrX where a cradle-shaped periplasmic metal sensor domain is anchored into the inner membrane by two N-terminal helices. In addition, they show that the main determinant of NccX dimerization is the periplasmic soluble domain and that the interaction between transmembrane segments is highly dynamic. The present work introduces a new crystal structure for a transmembrane protein and, in line with previous studies, substantiates the use of complementary theoretical and in vivo investigations to rationalize a three-dimensional structure obtained in non-native conditions.

Arpel, A. Sawma, P. Spenle, C. Fritz, J. Meyer, L. Garnier, N. Velazquez-Quesada, I. Hussenet, T. Aci-Seche, S. Baumlin, N. Genest, M. Brasse, D. Hubert, P. Cremel, G. Orend, G. Laquerriere, P. Bagnard, D.  (2014)

Transmembrane domain targeting peptide antagonizing ErbB2/Neu inhibits breast tumor growth and metastasis

Cell Reports (2014) 8 (6) 1714-1721 - doi : 10.1016/j.celrep.2014.07.044
Breast cancer is still a deadly disease despite major achievements in targeted therapies designed to block ligands or ligand-binding subunits of major tyrosine kinase receptors. Relapse is significant and metastases deleterious, which demands novel strategies for fighting this disease. Here, we report a proof-of-concept experiment demonstrating that small peptides interfering with the transmembrane domain of the tyrosine kinase epidermal growth factor receptor ErbB2 exhibit anticancer properties when used at micromolar dosages in a genetically engineered mouse model of breast cancer. Different assays demonstrate the specificity of the ErbB2-targeting peptide, which induces long-term reduction of ErbB2 phosphorylation and Akt signaling consistent with reduced tumor cell proliferation and increased survival. Microcomputed tomography analysis established the antimetastatic activity of the peptide and its impact on primary tumor growth. This reveals the interior of the cell membrane as an unexplored dimension for drug design.

Breast cancer is still a deadly disease despite major achievements in targeted therapies designed to block ligands or ligand-binding subunits of major tyrosine kinase receptors. Relapse is significant and metastases deleterious, which demands novel strategies for fighting this disease. Here, we report a proof-of-concept experiment demonstrating that small peptides interfering with the transmembrane domain of the tyrosine kinase epidermal growth factor receptor ErbB2 exhibit anticancer properties when used at micromolar dosages in a genetically engineered mouse model of breast cancer. Different assays demonstrate the specificity of the ErbB2-targeting peptide, which induces long-term reduction of ErbB2 phosphorylation and Akt signaling consistent with reduced tumor cell proliferation and increased survival. Microcomputed tomography analysis established the antimetastatic activity of the peptide and its impact on primary tumor growth. This reveals the interior of the cell membrane as an unexplored dimension for drug design.

Aci-Sèche S., Sawma P., Hubert P., Sturgis J.N., Bagnard D., Jacob L. Genest M. and Garnier N.  (2014)

Transmembrane Recognition of the Semaphorin Co- Receptors Neuropilin 1 and Plexin A1 : Coarse-Grained Simulations

PLoS One (2014) 9 (5) e97779 - doi : doi:10.1371/journal.pone.0097779
The cancer associated class 3 semaphorins require direct binding to neuropilins and association to plexins to trigger cell signaling. Here, we address the role of the transmembrane domains of neuropilin 1 and plexin A1 for the dimerization of the

two receptors by characterizing the assembly in lipid bilayers using coarse-grained molecular dynamics simulations. From experimental evidence using a two-hybrid system showing the biochemical association of the two receptors transmembrane domains, we performed molecular simulations in DOPC and POPC demonstrating spontaneously assembly to form homodimers and heterodimers with a very high propensity for right-handed packing of the helices. Inversely, lefthanded packing was observed with a very low propensity. This mode of packing was observed uniquely when the plexin A1

transmembrane domain was involved in association. Potential of mean force calculations were used to predict a hierarchy of self-association for the monomers : the two neuropilin 1 transmembrane domains strongly associated, neuropilin 1 and plexin A1 transmembrane domains associated less and the two plexin A1 transmembrane domains weakly but significantly associated. We demonstrated that homodimerization and heterodimerization are driven by GxxxG motifs, and that the sequence context modulates the packing mode of the plexin A1 transmembrane domains. This work presents major

advances towards our understanding of membrane signaling platforms assembly through membrane domains and provides exquisite information for the design of antagonist drugs defining a novel class of therapeutic agents.

The cancer associated class 3 semaphorins require direct binding to neuropilins and association to plexins to trigger cell signaling. Here, we address the role of the transmembrane domains of neuropilin 1 and plexin A1 for the dimerization of the
two receptors by characterizing the assembly in lipid bilayers using coarse-grained molecular dynamics simulations. From experimental evidence using a two-hybrid system showing the biochemical association of the two receptors transmembrane domains, we performed molecular simulations in DOPC and POPC demonstrating spontaneously assembly to form homodimers and heterodimers with a very high propensity for right-handed packing of the helices. Inversely, lefthanded packing was observed with a very low propensity. This mode of packing was observed uniquely when the plexin A1
transmembrane domain was involved in association. Potential of mean force calculations were used to predict a hierarchy of self-association for the monomers : the two neuropilin 1 transmembrane domains strongly associated, neuropilin 1 and plexin A1 transmembrane domains associated less and the two plexin A1 transmembrane domains weakly but significantly associated. We demonstrated that homodimerization and heterodimerization are driven by GxxxG motifs, and that the sequence context modulates the packing mode of the plexin A1 transmembrane domains. This work presents major
advances towards our understanding of membrane signaling platforms assembly through membrane domains and provides exquisite information for the design of antagonist drugs defining a novel class of therapeutic agents.


2013   Références trouvées : 1

Senille V., Lelievre D., Paquet F., Garnier N., Lamb N., Legrand A., Delmas A.F., Landon C.  (2013)

The Addressing Fragment of Mitogaligin : First Insights into Functional and Structural Properties

ChemBioChem 14 (6) 711-720
Mitogaligin is a mitochondrion-targeting protein involved in cell death. The sequence of the protein is unrelated to that of any known pro- or antiapoptotic protein. Mitochondrial targeting is controlled by an internal sequence from residues 31 to 53, and although this sequence is essential and sufficient to provoke cell death, the precise mechanism of action at the mitochondrial membrane remains to be elucidated. Here, by focusing on the [31–53] fragment, we first assessed and confirmed its cell cytotoxicity by microinjection. Subsequently, with the aid of membrane models, we evaluated the impact of the membrane environment on the 3D structure of the peptide and on how the peptide is embedded and oriented within membranes. The fragment is well organized, even though it does not contain a canonical secondary structure, and adopts an interfacial location. Structural comparison with other membrane-interacting Trp-rich peptides demonstrated similarities with the antimicrobial peptide tritrpcidin.

Mitogaligin is a mitochondrion-targeting protein involved in cell death. The sequence of the protein is unrelated to that of any known pro- or antiapoptotic protein. Mitochondrial targeting is controlled by an internal sequence from residues 31 to 53, and although this sequence is essential and sufficient to provoke cell death, the precise mechanism of action at the mitochondrial membrane remains to be elucidated. Here, by focusing on the [31–53] fragment, we first assessed and confirmed its cell cytotoxicity by microinjection. Subsequently, with the aid of membrane models, we evaluated the impact of the membrane environment on the 3D structure of the peptide and on how the peptide is embedded and oriented within membranes. The fragment is well organized, even though it does not contain a canonical secondary structure, and adopts an interfacial location. Structural comparison with other membrane-interacting Trp-rich peptides demonstrated similarities with the antimicrobial peptide tritrpcidin.


2011   Références trouvées : 2

Aci-Seche, S., Genest, M. & Garnier, N.  (2011)

Ligand entry pathways in the ligand binding domain of PPAR gamma receptor

FEBS Letters 585 (16) 2599-2603
To address the question of ligand entry process, we report targeted molecular dynamics simulations of the entry of the flexible ionic ligand GW0072 in the ligand binding domain of the nuclear receptor PPARγ. Starting with the ligand outside the receptor the simulations led to a ligand docked inside the binding pocket resulting in a structure very close to the holo-form of the complex. The results showed that entry process is guided by hydrophobic interactions and that entry pathways are very similar to exit pathways. We suggest that TMD method may help in discriminating between ligands generated by in silico docking.

To address the question of ligand entry process, we report targeted molecular dynamics simulations of the entry of the flexible ionic ligand GW0072 in the ligand binding domain of the nuclear receptor PPARγ. Starting with the ligand outside the receptor the simulations led to a ligand docked inside the binding pocket resulting in a structure very close to the holo-form of the complex. The results showed that entry process is guided by hydrophobic interactions and that entry pathways are very similar to exit pathways. We suggest that TMD method may help in discriminating between ligands generated by in silico docking.

Garnier, N., Loth, K., Coste, F. Augustyniak, R., Nadan, V., Damblon, C. & Castaing, B..  (2011)

An alternative flexible conformation of the E. coli HUβ(2) protein : structural, dynamics, and functional aspects.

European Biophysics Journal 40 (2) 117-129
The histone-like HU protein is the major nucleoid-associated protein involved in the dynamics and structure of the bacterial chromosome. Under physiological conditions, the three possible dimeric forms of the E. coli HU protein (EcHUα₂, EcHUβ₂, and EcHUαβ) are in thermal equilibrium between two dimeric conformations (N₂ ↔ I₂) varying in their secondary structure content. High-temperature molecular dynamics simulations combined with NMR experiments provide information about structural and dynamics features at the atomic level for the N₂ to I₂ thermal transition of the EcHUβ₂ homodimer. On the basis of these data, a realistic 3D model is proposed for the major I₂ conformation of EcHUβ₂. This model is in agreement with previous experimental data.

The histone-like HU protein is the major nucleoid-associated protein involved in the dynamics and structure of the bacterial chromosome. Under physiological conditions, the three possible dimeric forms of the E. coli HU protein (EcHUα₂, EcHUβ₂, and EcHUαβ) are in thermal equilibrium between two dimeric conformations (N₂ ↔ I₂) varying in their secondary structure content. High-temperature molecular dynamics simulations combined with NMR experiments provide information about structural and dynamics features at the atomic level for the N₂ to I₂ thermal transition of the EcHUβ₂ homodimer. On the basis of these data, a realistic 3D model is proposed for the major I₂ conformation of EcHUβ₂. This model is in agreement with previous experimental data.


2010   Références trouvées : 1

Aci, S., Garnier, N., Goffinont, S., Genest, D., Spotheim-Maurizot , M. & Genest, M.  (2010)

Comparing native and irradiated E. coli lactose repressor-operator complex by molecular dynamics simulation.

European Biophysics Journal, 39, 1375-1384.


2009   Références trouvées : 1

Aci-Seche, S., Garnier, N., Genest, D., Bourg, S., Marot, C., Morin-Allory, L. & Genest, M.  (2009)

A Restrained Molecular Dynamics Empirical Approach for Generating a Small Set of Structures Representative of the Internal Flexibility of a Receptor.

QSAR Combinatorial Science, 28, 959-968.
Multiple protein structure methods have been proposed for incorporating protein flexibility in molecular docking. One approach for docking ligands onto a rigid receptor is to use an ensemble of multiple rigid structures determined experimentally by X-ray or NMR spectroscopy or generated by numerical simulations. In this work we present all empirical method for generating a wide range of conformational states of a wobbling receptor using restrained Molecular Dynamics simulations (MD) and we propose a partitioning protocol for selecting a few representative conformations of the binding site from restrained MID sampling. 

Defining a large number of protein structures is computationally expensive when the MD simulations use an explicit solvent representation. For computational efficiency, solvent effect is therefore represented by an ensemble of restraints applied on a subset of specific atoms, using a distance-dependent permittivity function. The parameters used for the restraints and the permittivity are described. Several 100 ns restrained MD simulations are performed using different sets of parameters. In order to optimize the parameters, the results are compared to a 30 ns MD simulation in explicit solvent. Conformational sampling is speeded up by a factor of around 10-20 when performing restrained MD simulations. A partitioning k-means algorithm is applied to select representative structures of the receptor binding site. The methodology was evaluated on the ligand binding domain of the flexible Peroxysome Proliferator-Activated Receptor-gamma (PPAR gamma).

Multiple protein structure methods have been proposed for incorporating protein flexibility in molecular docking. One approach for docking ligands onto a rigid receptor is to use an ensemble of multiple rigid structures determined experimentally by X-ray or NMR spectroscopy or generated by numerical simulations. In this work we present all empirical method for generating a wide range of conformational states of a wobbling receptor using restrained Molecular Dynamics simulations (MD) and we propose a partitioning protocol for selecting a few representative conformations of the binding site from restrained MID sampling.

Defining a large number of protein structures is computationally expensive when the MD simulations use an explicit solvent representation. For computational efficiency, solvent effect is therefore represented by an ensemble of restraints applied on a subset of specific atoms, using a distance-dependent permittivity function. The parameters used for the restraints and the permittivity are described. Several 100 ns restrained MD simulations are performed using different sets of parameters. In order to optimize the parameters, the results are compared to a 30 ns MD simulation in explicit solvent. Conformational sampling is speeded up by a factor of around 10-20 when performing restrained MD simulations. A partitioning k-means algorithm is applied to select representative structures of the receptor binding site. The methodology was evaluated on the ligand binding domain of the flexible Peroxysome Proliferator-Activated Receptor-gamma (PPAR gamma).


2008   Références trouvées : 2

Genest, D ; Garnier, N ; Arrault, A ; Marot, C ; Morin-Allory, L ; Genest, M   (2008)

Ligand-escape pathways from the Ligand Binding Domain of PPAR receptor as probed by molecular dynamics simulations

European Biophysics Journal 37 (4) 369-379
Conformational rearrangements of peroxysome proliferator activated receptor (PPAR ?) ligand-binding domain (LBD) that accompany the release and binding of ligands are not well understood. To determine the major events associated with the escape of the partial agonist GW0072, molecular dynamic (MD) simulations were performed using two different methods : reversed targeted molecular dynamics (TMD-1) and time-dependent distance restraints (TDR) using as restraints either the root mean square deviation from a reference structure (TMD-1) or the distance between the geometrical centers of the binding pocket and of the ligand (TDR). Both methods do not assume any a priori route for ligand extraction.

Conformational rearrangements of peroxysome proliferator activated receptor (PPAR ?) ligand-binding domain (LBD) that accompany the release and binding of ligands are not well understood. To determine the major events associated with the escape of the partial agonist GW0072, molecular dynamic (MD) simulations were performed using two different methods : reversed targeted molecular dynamics (TMD-1) and time-dependent distance restraints (TDR) using as restraints either the root mean square deviation from a reference structure (TMD-1) or the distance between the geometrical centers of the binding pocket and of the ligand (TDR). Both methods do not assume any a priori route for ligand extraction.

Samna Soumana, O ; Garnier, N ; Genest, M   (2008)

Insight into the recognition patterns of the ErbB receptor family transmembrane domains : heterodimerization models through MD search

European Biophysics Journal, 37 (6) 851-864.
ErbB receptors undergo a complex interaction network defining hierarchical and competition relationships. Dimerization is driven entirely by receptor-receptor interactions and the transmembrane domains play a role in modulating the specificity and the selection of the partners during signal transduction. To shed light on the role of the GxxxG-like dimerization motifs in the formation of ErbB transmembrane heterodimers, we propose structural models resulting from conformational search method combined with molecular dynamics simulations. Left-handed structures of the transmembrane heterodimers are found preponderant over right-handed structures. All heterotypic heterodimers undergo two modes of association either via the N-terminal motif or the C-terminal motif. The transmembrane domain of ErbB3 impairs this C-terminal motif but also associates with the other partners owing to the presence of Gly residues. The two dimerization modes involve different orientations of the two helices. Thus, a molecular-switch model allowing the transition between the two dimerizing states may apply to the heterodimers and could help interpret receptor competition for the formation of homodimers and heterodimers. The comparison between experimental and theoretical results on the dimerization hierarchy of the transmembrane domains is not straightforward. However, we demonstrate that the intrinsic properties of the transmembrane sequences are an important component in heterodimer formation and that the ErbB2 and ErbB3 transmembrane domains have a strong power for heterodimerization as observed experimentally..

ErbB receptors undergo a complex interaction network defining hierarchical and competition relationships. Dimerization is driven entirely by receptor-receptor interactions and the transmembrane domains play a role in modulating the specificity and the selection of the partners during signal transduction. To shed light on the role of the GxxxG-like dimerization motifs in the formation of ErbB transmembrane heterodimers, we propose structural models resulting from conformational search method combined with molecular dynamics simulations. Left-handed structures of the transmembrane heterodimers are found preponderant over right-handed structures. All heterotypic heterodimers undergo two modes of association either via the N-terminal motif or the C-terminal motif. The transmembrane domain of ErbB3 impairs this C-terminal motif but also associates with the other partners owing to the presence of Gly residues. The two dimerization modes involve different orientations of the two helices. Thus, a molecular-switch model allowing the transition between the two dimerizing states may apply to the heterodimers and could help interpret receptor competition for the formation of homodimers and heterodimers. The comparison between experimental and theoretical results on the dimerization hierarchy of the transmembrane domains is not straightforward. However, we demonstrate that the intrinsic properties of the transmembrane sequences are an important component in heterodimer formation and that the ErbB2 and ErbB3 transmembrane domains have a strong power for heterodimerization as observed experimentally..


2007   Références trouvées : 1

Samna Soumana, O ; Garnier, N ; Genest, M  (2007)

Molecular dynamics simulation approach for the prediction of transmembrane helix-helix heterodimers assembly.

European Biophysics Journal, 36, 1071-1082
Computational methods are useful to identify favorable structures of transmembrane (TM) helix oligomers when experimental data are not available or when they cannot help to interpret helix-helix association. We report here a global search method using molecular dynamics (MD) simulations to predict the structures of transmembrane homo and heterodimers. The present approach is based only on sequence information without any experimental data and is first applied to glycophorin A to validate the protocol and to the HER2-HER3 heterodimer receptor.

Computational methods are useful to identify favorable structures of transmembrane (TM) helix oligomers when experimental data are not available or when they cannot help to interpret helix-helix association. We report here a global search method using molecular dynamics (MD) simulations to predict the structures of transmembrane homo and heterodimers. The present approach is based only on sequence information without any experimental data and is first applied to glycophorin A to validate the protocol and to the HER2-HER3 heterodimer receptor.


2006   Références trouvées : 1

Aller, P ; Garnier, N ; Genest, M  (2006)

Transmembrane helix packing of ErbB/Neu receptor in membrane environment : A molecular dynamics study

Journal of Biomolecular Structure & Dynamics 24 (3) 209-228
Dimerization or oligomerization of the ErbB/Neu receptors are necessary but not sufficient for initiation of receptor signaling. The two intracellular domains must be properly oriented for the juxtaposition of the kinase domains allowing trans-phosphorylation. This suggests that the transmembrane (TM) domain acts as a guide for defining the proper orientation of the intracellular domains. Two structural models, with the two helices either in left-handed or in right-handed coiling have been proposed as the TM domain structure of the active receptor.

Dimerization or oligomerization of the ErbB/Neu receptors are necessary but not sufficient for initiation of receptor signaling. The two intracellular domains must be properly oriented for the juxtaposition of the kinase domains allowing trans-phosphorylation. This suggests that the transmembrane (TM) domain acts as a guide for defining the proper orientation of the intracellular domains. Two structural models, with the two helices either in left-handed or in right-handed coiling have been proposed as the TM domain structure of the active receptor.


2005   Références trouvées : 2

Samna Soumana, O ; Aller, P ; Garnier, N ; Genest, M  (2005)

Transmembrane peptides from tyrosine kinase receptor. Mutation-related behavior in a lipid bilayer investigated by molecular dynamics simulations

Journal of Biomolecular Structure & Dynamics 23 (1) 91-100
Polar mutations in transmembrane a helices may alter the structural details of the hydrophobic sequences and control intermolecular contacts. We have performed molecular dynamics simulations on the transmembrane domain of the proto-oncogenic and the oncogenic forms of the Neu receptor in a fluid DMPC bilayer to test whether the Gin mutation which replaces the Val residue at position 664 may alter the helical structure and its insertion in the membrane. The simulations show that the wild and the mutant forms of the transmembrane domain have a different behavior in the bilayer. The native transmembrane sequence is found to be more flexible than in the presence of the Glu mutation, characterized by a tendency to pi deformation to accommodate the helix length to the membrane thickness. The mutant form of this domain does not evidence helical deformation in the present simulation.

Polar mutations in transmembrane a helices may alter the structural details of the hydrophobic sequences and control intermolecular contacts. We have performed molecular dynamics simulations on the transmembrane domain of the proto-oncogenic and the oncogenic forms of the Neu receptor in a fluid DMPC bilayer to test whether the Gin mutation which replaces the Val residue at position 664 may alter the helical structure and its insertion in the membrane. The simulations show that the wild and the mutant forms of the transmembrane domain have a different behavior in the bilayer. The native transmembrane sequence is found to be more flexible than in the presence of the Glu mutation, characterized by a tendency to pi deformation to accommodate the helix length to the membrane thickness. The mutant form of this domain does not evidence helical deformation in the present simulation.

Aller, P ; Voiry, L ; Garnier, N ; Genest, M  (2005)

Molecular dynamics (MD) investigations of preformed structures of the transmembrane domain of the oncogenic Neu receptor dimer in a DMPC bilayer

Biopolymers 77 (4) 184-197
The critical Val/Glu mutation in the membrane spanning domain of the rat Neu receptor confers the ability for ligand-independent signaling and leads to increased dimerization and transforming ability. There is evidence that the two transmembrane interacting helices play a role in receptor activation by imposing orientation constraints to the intracellular tyrosine kinase domains. By using MD simulations we have attempted to discriminate between correct and improper helix-helix packing by examining the structural and energetic properties of preformed left-handed and right-handed structures in a fully hydrated DMPC bilayer. The best energetic balance between the residues at the helix-helix interface and the residues exposed to the lipids is obtained for helices in symmetrical left-handed interactions packed together via Glu side chain/Ala backbone interhelical hydrogen bonds.

The critical Val/Glu mutation in the membrane spanning domain of the rat Neu receptor confers the ability for ligand-independent signaling and leads to increased dimerization and transforming ability. There is evidence that the two transmembrane interacting helices play a role in receptor activation by imposing orientation constraints to the intracellular tyrosine kinase domains. By using MD simulations we have attempted to discriminate between correct and improper helix-helix packing by examining the structural and energetic properties of preformed left-handed and right-handed structures in a fully hydrated DMPC bilayer. The best energetic balance between the residues at the helix-helix interface and the residues exposed to the lipids is obtained for helices in symmetrical left-handed interactions packed together via Glu side chain/Ala backbone interhelical hydrogen bonds.


2003   Références trouvées : 1

Garnier, N., Crouzy, S. & Genest, M.  (2003)

Molecular Dynamics Simulations of the Transmembrane Domain of the Oncogenic ErbB2 Receptor Dimer in a DMPC Bilayer.

Journal of Biomolecular Structure and Dynamics 21, 179-199.


1999   Références trouvées : 2

Duneau, JP ; Crouzy, S ; Garnier, N ; Chapron, Y ; Genest, M  (1999)

Molecular dynamics simulations of the ErbB-2 transmembrane domain within an explicit membrane environment : comparison with vacuum simulations

Biophysical Chemistry 76 (1) 35-53
Two 500-ps molecular dynamics simulations performed on the single transmembrane domain of the ErbB-2 tyrosine kinase receptor immersed in a fully solvated dilauroylphosphatidyl-ethanolamine bilayer (DLPE) are compared to vacuum simulations. One membrane simulation shows that the initial alpha helix undergoes a local pi helix conversion in the peptide part embedded in the membrane core similar to that found in simulation vacuum. Lipid/water/peptide interaction analysis shows that in the helix core, the intramolecular peptide interactions are largely dominant compared to the interactions with water and lipids whereas the helix extremities are much more sensitive to these interactions at the membrane interfaces. Our results suggest that simulations in a lipid environment are required to understand the dynamics of transmembrane helices, but can be reasonably supplemented by in vacuo simulations to explore rapidly its conformational space and to describe the internal deformation of the hydrophobic core. (C) 1999 Elsevier Science B.V. All rights reserved.

Two 500-ps molecular dynamics simulations performed on the single transmembrane domain of the ErbB-2 tyrosine kinase receptor immersed in a fully solvated dilauroylphosphatidyl-ethanolamine bilayer (DLPE) are compared to vacuum simulations. One membrane simulation shows that the initial alpha helix undergoes a local pi helix conversion in the peptide part embedded in the membrane core similar to that found in simulation vacuum. Lipid/water/peptide interaction analysis shows that in the helix core, the intramolecular peptide interactions are largely dominant compared to the interactions with water and lipids whereas the helix extremities are much more sensitive to these interactions at the membrane interfaces. Our results suggest that simulations in a lipid environment are required to understand the dynamics of transmembrane helices, but can be reasonably supplemented by in vacuo simulations to explore rapidly its conformational space and to describe the internal deformation of the hydrophobic core. (C) 1999 Elsevier Science B.V. All rights reserved.

Sajot, N ; Garnier, N ; Genest, M  (1999)

Dimer models for ErbB-2/neu transmembrane domains from molecular dynamics simulations

Theoretical Chemistry Accounts 101 (1-3) 67-72
Interest in the transmembrane receptors tyrosine kinase of the erbB family is high due to the involvement of some of the members in human cancers. The original oncogenic alleles of neu discovered in rat neuroectodermal tumors lead to single Va1664Glu substitution within the predicted transmembrane domain : Identical substitution at the homologous position 659 constitutively activates the oncogenic potential of the human ErbB-2 receptor by enhanced receptor dimer formation. The precise molecular details of receptor dimerization are still unknown and to acquire more knowledge of the mechanisms involved, molecular dynamics simulations are undertaken to study transmembrane dimer association. Transmembrane helices are predicted to associate in left-handed coiled-coil structures stabilized by Glu-Glu interhelix hydrogen bonds in the mutated form. The internal dynamics reveals pi helix deformations which modify the helix-helix interface. Predicted models agree with those suggested from polarized IR and magic-angle spinning NMR spectroscopy.

Interest in the transmembrane receptors tyrosine kinase of the erbB family is high due to the involvement of some of the members in human cancers. The original oncogenic alleles of neu discovered in rat neuroectodermal tumors lead to single Va1664Glu substitution within the predicted transmembrane domain : Identical substitution at the homologous position 659 constitutively activates the oncogenic potential of the human ErbB-2 receptor by enhanced receptor dimer formation. The precise molecular details of receptor dimerization are still unknown and to acquire more knowledge of the mechanisms involved, molecular dynamics simulations are undertaken to study transmembrane dimer association. Transmembrane helices are predicted to associate in left-handed coiled-coil structures stabilized by Glu-Glu interhelix hydrogen bonds in the mutated form. The internal dynamics reveals pi helix deformations which modify the helix-helix interface. Predicted models agree with those suggested from polarized IR and magic-angle spinning NMR spectroscopy.


1998   Références trouvées : 1

Duneau, JP ; Garnier, N ; Cremel, G ; Nullans, G ; Hubert, P ; Genest, D ; Vincent, M ; Gallay, J ; Genest, M  (1998)

Time resolved fluorescence properties of phenylalanine in different environments. Comparison with molecular dynamics simulation

Biophysical Chemistry 73 (1-2) 109-119
Time resolved fluorescence of the phenylalanine residue (Phe) alone and included in the transmembrane domain (TMD) sequences of the epidermal growth factor receptor (EGFR) and ErbB-2 was studied using the synchrotron radiation source of light, and compared to molecular dynamics (MD) simulations. The fluorescence intensity decay is strongly sensitive to the environment. A mono-exponential decay was obtained for Phe amino acid alone in two different solvents and for Phe included in EGFR transmembrane sequence, with fluorescence lifetime values varying from 1.7 ns (EGFR) to 7.4 ns (Phe dissolved in water). In ErbB-2 transmembrane sequence three lifetimes were detected.

Time resolved fluorescence of the phenylalanine residue (Phe) alone and included in the transmembrane domain (TMD) sequences of the epidermal growth factor receptor (EGFR) and ErbB-2 was studied using the synchrotron radiation source of light, and compared to molecular dynamics (MD) simulations. The fluorescence intensity decay is strongly sensitive to the environment. A mono-exponential decay was obtained for Phe amino acid alone in two different solvents and for Phe included in EGFR transmembrane sequence, with fluorescence lifetime values varying from 1.7 ns (EGFR) to 7.4 ns (Phe dissolved in water). In ErbB-2 transmembrane sequence three lifetimes were detected.


1997   Références trouvées : 2

Garnier, N ; Genest, D ; Duneau, JP ; Genest, M  (1997)

Molecular modeling of c-erbB2 receptor dimerization : Coiled-coil structure of wild and oncogenic transmembrane domains - Stabilization by interhelical hydrogen bonds in the oncogenic form

Biopolymers 42 (2) 157-168
Dimerization models of c-erbB2 transmembrane domains (Leu651-Ile675) are studied by molecular mechanics and molecular dynamics simulations. Both wild and Glu mutated transmembrane helices exhibit the same relative orientation for favorable associations and dimerize preferentially in left-handed coiled-coil structures. The mutation point 659 belongs to the interfacing residues, and in the transforming domain, symmetric hydrogen bonds between Glu carboxylic groups stabilize the dimeric structure. The same helix packing found for the wild dimers, except side-chain-side-chain hydrogen bonds, suggests that the transmembrane domains dimerize according to similar process. Structural and energetical characterization of the models are presented. (C) 1997 John Wiley & Sons, Inc.

Dimerization models of c-erbB2 transmembrane domains (Leu651-Ile675) are studied by molecular mechanics and molecular dynamics simulations. Both wild and Glu mutated transmembrane helices exhibit the same relative orientation for favorable associations and dimerize preferentially in left-handed coiled-coil structures. The mutation point 659 belongs to the interfacing residues, and in the transforming domain, symmetric hydrogen bonds between Glu carboxylic groups stabilize the dimeric structure. The same helix packing found for the wild dimers, except side-chain-side-chain hydrogen bonds, suggests that the transmembrane domains dimerize according to similar process. Structural and energetical characterization of the models are presented. (C) 1997 John Wiley & Sons, Inc.

Duneau, JP ; Garnier, N ; Genest, M  (1997)

Insight into signal transduction : Structural alterations in transmembrane helices probed by multi-1 ns molecular dynamics simulations.

Journal of Biomolecular Structure & Dynamics, 15 (3) 555-572
The hypothesis of structural alteration in transmembrane helices for signal transduction process is viewed by molecular dynamics simulation techniques. For the c-erbB-2 transmembrane domain involved in oncogenicity, the occurrence of conformational changes has been previously described as transition from the alpha to pi helix. This dynamical feature is thoroughly analyzed for the wild phenotype and oncogenic sequences from a series of 18 simulations carried out on one nanosecond time scale. We show that these structural events do not depend upon the conditions of simulations like force field or starting helix coordinates. We demonstrate that the oncogenic mutations Val659 Glu, Gln and Asp do not prevent the transition. Furthermore, we show that beta branched residues, in conjunction with Gly residues in the c-erbB-2 sequence, act as destabilizers for the alpha helix structure. pi deformations are tightly related to other local structural motifs found in soluble and membrane proteins. These structural alterations are discussed in term of structure-activity relationships for the c-erbB-2 activating mechanism mediated by transmembrane domain dimerization.

The hypothesis of structural alteration in transmembrane helices for signal transduction process is viewed by molecular dynamics simulation techniques. For the c-erbB-2 transmembrane domain involved in oncogenicity, the occurrence of conformational changes has been previously described as transition from the alpha to pi helix. This dynamical feature is thoroughly analyzed for the wild phenotype and oncogenic sequences from a series of 18 simulations carried out on one nanosecond time scale. We show that these structural events do not depend upon the conditions of simulations like force field or starting helix coordinates. We demonstrate that the oncogenic mutations Val659 Glu, Gln and Asp do not prevent the transition. Furthermore, we show that beta branched residues, in conjunction with Gly residues in the c-erbB-2 sequence, act as destabilizers for the alpha helix structure. pi deformations are tightly related to other local structural motifs found in soluble and membrane proteins. These structural alterations are discussed in term of structure-activity relationships for the c-erbB-2 activating mechanism mediated by transmembrane domain dimerization.


1996   Références trouvées : 1

Garnier, N ; Genest, D ; Genest, M  (1996)

Correlated motions and propagation of the effect of a local conformational change in the transmembrane helix of the c-erbB2 encoded protein and in its V659E mutant, studied by molecular dynamics simulations

Biophysical Chemistry 58 (3) 225-237
A detailed study of the dynamical behavior of the 29-residue peptide including the transmembrane domain of p185(c-erbB2) oncogene-encoded protein and of its V659E mutant is presented. In a first part of this work we analyse equal time correlation coefficients between the backbone dihedral angle fluctuations. Concerted motions are observed in the wild type transmembrane a-helix but not in the corresponding V659E intramembrane domain. The difference observed in the correlation pattern is attributed to the single amino acid replacement. In a second part, we investigate the propagation of the effect of a local conformational change along the transmembrane segment, one of the dominant hypotheses for signal transduction mechanisms of transmembrane receptors. The analysis of angular time correlation functions together with that of the response of the different residues to a local disturbance applied at the N-terminal side evidences a propagation phenomenon for the wild type peptide. This effect is much less clear for the mutated peptide. Furthermore we show that the first one is much more flexible than the second one.

A detailed study of the dynamical behavior of the 29-residue peptide including the transmembrane domain of p185(c-erbB2) oncogene-encoded protein and of its V659E mutant is presented. In a first part of this work we analyse equal time correlation coefficients between the backbone dihedral angle fluctuations. Concerted motions are observed in the wild type transmembrane a-helix but not in the corresponding V659E intramembrane domain. The difference observed in the correlation pattern is attributed to the single amino acid replacement. In a second part, we investigate the propagation of the effect of a local conformational change along the transmembrane segment, one of the dominant hypotheses for signal transduction mechanisms of transmembrane receptors. The analysis of angular time correlation functions together with that of the response of the different residues to a local disturbance applied at the N-terminal side evidences a propagation phenomenon for the wild type peptide. This effect is much less clear for the mutated peptide. Furthermore we show that the first one is much more flexible than the second one.


1994   Références trouvées : 2

Garnier, N ; Genest, D ; Hebert, E ; Genest, M  (1994)

Influence of a mutation in the transmembrane domain of the p185(c-erbb2) oncogene-encoded protein studied by molecular-dynamics simulations

Journal of Biomolecular Structure & Dynamics 11 (5) 983-1002
The c-erbB2 proto-oncogene encodes for a protein of 185kDa(p185) which becomes transforming upon the Val—>Glu transmembrane amino acid substitution. The transforming ability seems to be due to a substitution-resulting constitutive activation of the tyrosine kinase cytosolic domain of the protein. These observations prompted us to evaluate the structural and dynamical behavior of the transmembrane region of the wild and transforming p185 protein in order to understand the role of this region in the transduction mechanism. 160 ps molecular dynamics simulations in vacuo have been performed on two peptides corresponding to the sequence [651-679] of p185(c-erbB2) protein and its transforming mutant Val(659)—>Glu(659).

The c-erbB2 proto-oncogene encodes for a protein of 185kDa(p185) which becomes transforming upon the Val—>Glu transmembrane amino acid substitution. The transforming ability seems to be due to a substitution-resulting constitutive activation of the tyrosine kinase cytosolic domain of the protein. These observations prompted us to evaluate the structural and dynamical behavior of the transmembrane region of the wild and transforming p185 protein in order to understand the role of this region in the transduction mechanism. 160 ps molecular dynamics simulations in vacuo have been performed on two peptides corresponding to the sequence [651-679] of p185(c-erbB2) protein and its transforming mutant Val(659)—>Glu(659).

Garnier, N ; Genest, D ; Genest, M   (1994)

Motions and correlations of the transmembrane domain of a protein studied by molecular dynamics simulation

Non Linear Exitations in Biomolecules - Peyrard, M - 241-246 - Les Editions de physique Springer


1991   Références trouvées : 3

Garnier, N.  (1991)

Brownian dynamics close to a wall, measured by quasi-elastic light scattering from an evanescent wave.

Progress in Colloid and Polymer Science 84, 371-376

Ostrowsky, N ; Garnier, N  (1991)

Quasi-elastic light-scattering from an evanescent wave to probe particle wall interactions

Biochemical Society Transactions 19 (2) 500-501

Garnier, N ; Ostrowsky, N  (1991)

Brownian dynamics in a confined geometry - experiments and numerical simulations

Journal de Physique 1 (10) 1221-1232
The Brownian dynamics of a colloidal suspension is measured in the immediate vicinity of a rigid surface by the Evanescent Quasielastic Light Scattering Technique. A net decrease of the measured diffusion coefficient is observed, due to the hydrodynamic slowing down of the particles very close to the wall. This effect is all the more important when the particles are allowed to get closer to the wall, i.e. when the range of the static wall/particle repulsive interaction decreases. It thus provides a mean for testing the particle/wall static interactions via a dynamic light scattering measurement. The data are analysed by a Brownian dynamic simulation which is proven to be quite valuable to interpret light scattering data from

The Brownian dynamics of a colloidal suspension is measured in the immediate vicinity of a rigid surface by the Evanescent Quasielastic Light Scattering Technique. A net decrease of the measured diffusion coefficient is observed, due to the hydrodynamic slowing down of the particles very close to the wall. This effect is all the more important when the particles are allowed to get closer to the wall, i.e. when the range of the static wall/particle repulsive interaction decreases. It thus provides a mean for testing the particle/wall static interactions via a dynamic light scattering measurement. The data are analysed by a Brownian dynamic simulation which is proven to be quite valuable to interpret light scattering data from


1989   Références trouvées : 1

Ostrowsky, N., Garnier, N. & Sornette, D.  (1989)

The use of dynamic light scattering in studies of vesicles interactions.

Journal of Dispersion Science and Technology 10, 277-284.