Sampling of conformational ensemble for virtual screening using molecular dynamics simulations and normal mode analysis

Aim: Molecular dynamics simulations and normal mode analysis are well-established approaches to generate receptor conformational ensembles (RCEs) for ligand docking and virtual screening. Here, we report new fast molecular dynamics-based and normal mode analysis-based protocols combined with conformational pocket classifications to efficiently generate RCEs. Materials \& methods: We assessed our protocols on two well-characterized protein targets showing local active site flexibility, dihydrofolate reductase and large collective movements, CDK2. The performance of the RCEs was validated by distinguishing known ligands of dihydrofolate reductase and CDK2 among a dataset of diverse chemical decoys. Results \& discussion: Our results show that different simulation protocols can be efficient for generation of RCEs depending on different kind of protein flexibility

Duloxetine in the Treatment of Major Depressive Disorder: An Open-Label Study

Background: Major depressive disorder (MDD) is a chronic and highly disabling condition. Existing pharmacotherapies produce full remission in only 30% to 40% of treated patients. Antidepressants exhibiting dual reuptake inhibition of both serotonin (5-HT) and norepinephrine (NE) may achieve higher rates of remission compared with those acting upon a single neurotransmitter. In this study, the safety and efficacy of duloxetine, a potent dual reuptake inhibitor of 5-HT and NE, were examined. Methods: Patients (N = 533) meeting DSM-IV criteria for MDD received open-label duloxetine (60 mg once a day [QD]) for 12 weeks during the initial phase of a relapse prevention trial. Patients were required to have a 17-item Hamilton Rating Scale for Depression (HAMD17) total score ≥18 and a Clinical Global Impression of Severity (CGI-S) score ≥4 at baseline. Efficacy measures included the HAMD17 total score, HAMD17 subscales, the CGI-S, the Patient Global Impression of Improvement (PGI-I) scale, Visual Analog Scales (VAS) for pain, and the Symptom Questionnaire, Somatic Subscale (SQ-SS). Quality of life was assessed using the Sheehan Disability Scale (SDS) and the Quality of Life in Depression Scale (QLDS). Safety was evaluated by recording spontaneously-reported treatment-emergent adverse events, changes in vital signs and laboratory analytes, and the Patient Global Impression of Sexual Function (PGI-SF) scale. Results: The rate of discontinuation due to adverse events was 11.3%. Treatment-emergent adverse events reported by ≥10% duloxetine-treated patients were nausea, headache, dry mouth, somnolence, insomnia, and dizziness. Following 12 weeks of open-label duloxetine therapy, significant improvements were observed in all assessed efficacy and quality of life measures. In assessments of depression severity (HAMD17, CGI-S) the magnitude of symptom improvement continued to increase at each study visit, while for painful physical symptoms the onset of improvement was rapid and reached a maximum after 2 to 3 weeks of treatment. Conclusion: In this open-label phase of a relapse prevention study, duloxetine (60 mg QD) was shown to be safe and effective in the treatment of MDD. Trial registration: NCT00036309

Unexpected Gating Behaviour of an Engineered Potassium Channel Kir

In this study, we investigated the dynamics and functional characteristics of the KirBac3.1 S129R, a mutated bacterial potassium channel for which the inner pore-lining helix (TM2) was engineered so that the bundle crossing is trapped in an open conformation. The structure of this channel has been previously determined at high atomic resolution. We explored the dynamical characteristics of this open state channel using an in silico method MDeNM that combines molecular dynamics simulations and normal modes. We captured the global and local motions at the mutation level and compared these data with HDX-MS experiments. MDeNM provided also an estimation of the probability of the different opening states that are in agreement with our electrophysiological experiments. In the S129R mutant, the Arg129 mutation releases the two constriction points in the channel that existed in the wild type but interestingly creates another restriction point

Small GTPase Ran: Depicting the nucleotide-specific conformational landscape of the functionally important C-terminus

The small GTPase Ran is the main regulator of the nucleo-cytoplasmic import and export through the nuclear pore complex. It functions as a molecular switch cycling between the GDP-bound inactive and GTP-bound active state. It consists of a globular (G) domain and a C-terminal region, which is bound to the G-domain in the inactive, GDP-bound states. Crystal structures of the GTP-bound active form complexed with Ran binding proteins (RanBP) show that the C-terminus undergoes a large conformational change, embracing Ran binding domains (RanBD). Whereas in the crystal structures of macromolecular complexes not containing RanBDs the structure of the C-terminal segment remains unresolved, indicating its large conformational flexibility. This movement could not have been followed either by experimental or simulation methods. Here, starting from the crystal structure of Ran in both GDP- and GTP-bound forms we show how rigid the C-terminal region in the inactive structure is during molecular dynamics (MD) simulations. Furthermore, we show how MD simulations of the active form are incapable of mapping the open conformations of the C-terminus. By using the MDeNM (Molecular Dynamics with excited Normal Modes) method, we were able to widely map the conformational surface of the C-terminus of Ran in the active GTP-bound form, which allows us to envisage how it can embrace RanBDs

Duloxetine in the treatment of major depressive disorder: an open-label study

<p>Abstract</p> <p>Background</p> <p>Major depressive disorder (MDD) is a chronic and highly disabling condition. Existing pharmacotherapies produce full remission in only 30% to 40% of treated patients. Antidepressants exhibiting dual reuptake inhibition of both serotonin (5-HT) and norepinephrine (NE) may achieve higher rates of remission compared with those acting upon a single neurotransmitter. In this study, the safety and efficacy of duloxetine, a potent dual reuptake inhibitor of 5-HT and NE, were examined.</p> <p>Methods</p> <p>Patients (N = 533) meeting DSM-IV criteria for MDD received open-label duloxetine (60 mg once a day [QD]) for 12 weeks during the initial phase of a relapse prevention trial. Patients were required to have a 17-item Hamilton Rating Scale for Depression (HAMD₁₇) total score ≥18 and a Clinical Global Impression of Severity (CGI-S) score ≥4 at baseline. Efficacy measures included the HAMD₁₇total score, HAMD₁₇subscales, the CGI-S, the Patient Global Impression of Improvement (PGI-I) scale, Visual Analog Scales (VAS) for pain, and the Symptom Questionnaire, Somatic Subscale (SQ-SS). Quality of life was assessed using the Sheehan Disability Scale (SDS) and the Quality of Life in Depression Scale (QLDS). Safety was evaluated by recording spontaneously-reported treatment-emergent adverse events, changes in vital signs and laboratory analytes, and the Patient Global Impression of Sexual Function (PGI-SF) scale.</p> <p>Results</p> <p>The rate of discontinuation due to adverse events was 11.3%. Treatment-emergent adverse events reported by ≥10% duloxetine-treated patients were nausea, headache, dry mouth, somnolence, insomnia, and dizziness. Following 12 weeks of open-label duloxetine therapy, significant improvements were observed in all assessed efficacy and quality of life measures. In assessments of depression severity (HAMD₁₇, CGI-S) the magnitude of symptom improvement continued to increase at each study visit, while for painful physical symptoms the onset of improvement was rapid and reached a maximum after 2 to 3 weeks of treatment.</p> <p>Conclusion</p> <p>In this open-label phase of a relapse prevention study, duloxetine (60 mg QD) was shown to be safe and effective in the treatment of MDD.</p> <p>Trial registration</p> <p>NCT00036309.</p

Targeting STAT1 by myricetin and delphinidin provides efficient protection of the heart from ischemia/reperfusion-induced injury

AbstractFlavonoids exhibit a variety of beneficial effects in cardiovascular diseases. Although their therapeutic properties have been attributed mainly to their antioxidant action, they have additional protective mechanisms such as inhibition of signal transducer and activator of transcription 1 (STAT1) activation. Here, we have investigated the cardioprotective mechanisms of strong antioxidant flavonoids such as quercetin, myricetin and delphinidin. Although all of them protect the heart from ischemia/reperfusion-injury, myricetin and delphinidin exert a more pronounced protective action than quercetin by their capacity to inhibit STAT1 activation. Biochemical and computer modeling analysis indicated the direct interaction between STAT1 and flavonoids with anti-STAT1 activity

Mutation D816V Alters the Internal Structure and Dynamics of c-KIT Receptor Cytoplasmic Region: Implications for Dimerization and Activation Mechanisms

The type III receptor tyrosine kinase (RTK) KIT plays a crucial role in the transmission of cellular signals through phosphorylation events that are associated with a switching of the protein conformation between inactive and active states. D816V KIT mutation is associated with various pathologies including mastocytosis and cancers. D816V-mutated KIT is constitutively active, and resistant to treatment with the anti-cancer drug Imatinib. To elucidate the activating molecular mechanism of this mutation, we applied a multi-approach procedure combining molecular dynamics (MD) simulations, normal modes analysis (NMA) and binding site prediction. Multiple 50-ns MD simulations of wild-type KIT and its mutant D816V were recorded using the inactive auto-inhibited structure of the protein, characteristic of type III RTKs. Computed free energy differences enabled us to quantify the impact of D816V on protein stability in the inactive state. We evidenced a local structural alteration of the activation loop (A-loop) upon mutation, and a long-range structural re-organization of the juxta-membrane region (JMR) followed by a weakening of the interaction network with the kinase domain. A thorough normal mode analysis of several MD conformations led to a plausible molecular rationale to propose that JMR is able to depart its auto-inhibitory position more easily in the mutant than in wild-type KIT and is thus able to promote kinase mutant dimerization without the need for extra-cellular ligand binding. Pocket detection at the surface of NMA-displaced conformations finally revealed that detachment of JMR from the kinase domain in the mutant was sufficient to open an access to the catalytic and substrate binding sites

Study of protein-ligand interactions by quantum and molecular mechanical approaches

ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

Etude théorique des propriétés énergétiques et structurales de modèles du centre de fixation de l'hémoglobine

Dans ce travail, différentes propriétés du centre de fixation de l'oxygène dans l'hémoglobine (Hb) sont étudiées avec des méthodes de mécanique quantique pure, et une méthode hybride de mécanique quantique/mécanique moléculaire: IMOMM. Deux types de modèles de ce centre sont utilisés: les modèles minimaux qui sont constitués d'une partie de l'hème (porphyrine sans substituant (P) et atome de Fe(II)) et des 2 ligands axiaux du fer (une base aux propriétés voisines de l'histidine proximale (B) et l'O2), et les modèles étendus qui incluent l'hème dans son inte gralité et trois résidus distaux de l'Hb. Les trois degrés de coordination du fer sont considérés: 1-[Fe(II)(P)], 2-[Fe(II)(P)(B)] et 3-[Fe(II)(P)(B)(O2)]. La première propriété étudiée est le coût énergétique de la rotation des ligands axiaux du fer pour les deux degrés de coordination 2 et 3. Il ressort que les barrières de rotation sont très faibles. Ceci permet de comprendre le grand nombre d'orientations observées dans les structures biologiques. La deuxième propriété étudiée est le coût énergétique du déplacement du fer hors du plan de l'hème. Celui-ci est faible pour un large intervalle pour les modèles [Fe(II)(P)] et [Fe(II)(P)(B)]alors qu'il est très élevé pour le modèle [Fe(II)(P)(B)(O2)]. En absence d'oxygène le fer peut donc se déplacer aisément alors que la fixation de l'oxygène induit une contrainte importante. La dernière propriété étudiée est l'influence des résidus distaux de l'hème sur l'affinité de la protéine. Le calcul de l'énergie d'oxygénation sur les modèles étendus permet de tenir compte de l'état allostérique (T ou R) et du type de chaîne (a ou b) de l'Hb. Il apparaît que les résidus distaux sont responsables de la différence d'affinité observée d'une part entre les états T et R et d'autre part entre les chaînes a et b dans l'état T. La transition allostérique de T à R apparaît alors nécessaire pour que l'oxygène puisse se fixer sur la sous-unité b.In this work, different features of the oxygen fixation centre of hemoglobin (Hb) are studied using pure quantum mechanics methods and hybrid quantum mechanics/molecular mechanics method : IMOMM. Two types of models are used for the fixation centre: "minimal" models, in which the atoms in the vicinity of the iron atom are inserted (the naked porphyrine "P", a base with properties close to those of the proximal histidine "B" and the oxygen molecule), and "extended" models, where the porphyrine is complete and three distal residues are included. The three most common coordinations of the iron are considered: [Fe(II)(P)], [Fe(II)(P)(B)] and [Fe(II)(P)(B)(O2]. All the optimised structures and electronic states obtained from the theoretical methods are in good agreement with the experimental data. The first feature we studied was the rotation of the axial ligands of the iron. In all models the rotation barriers were very low. This result explains the variety of orientations seen in the experimental structures. The second feature was the energetic cost of the displacement of the iron out of the porphyrine plane. This cost was very low for the [Fe(II)(P)] and [Fe(II)(P)(B)] models but very high for the [Fe(II)(P)(B)(O2)] one. So, in the protein, the iron would appear to fluctuate easily in the absence of oxygen but an important constraint arises when O2 is fixed. The last feature we studied was the influence of the distal residues on the affinity of Hb. The extended models allowed the determination of the energy difference for oxygenation depending on the allosteric state (T=deoxy form, R=oxy form) and the subunit (a or b) environment. These calculations show that the distal residues stabilize oxygenation by about 5 kcal/mol for the aR, bR, and aT structures but destabilize it for the bT one. These results suggest that the allosteric transition from T to R is necessary for fixation of the oxygen to the b subunit which leads to an increase of the oxygen affinity of the protein.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF