scholarly journals A structural model of the human serotonin transporter in an outward-occluded state

2019 ◽  
Author(s):  
Eva Hellsberg ◽  
Gerhard F. Ecker ◽  
Anna Stary-Weinzinger ◽  
Lucy R. Forrest
Keyword(s):  
Serotonin Transporter ◽  
Conformational Changes ◽  
Structural Model ◽  
Antidepressant Drugs ◽  
Homology Model ◽  
Synaptic Cleft ◽  
Open Conformation ◽  
Starting Point ◽  
Ligand Interactions ◽  
Dynamics Simulations

AbstractThe human serotonin transporter hSERT facilitates the reuptake of its endogenous substrate serotonin from the synaptic cleft into presynaptic neurons after signaling. Reuptake regulates the availability of this neurotransmitter and therefore hSERT plays an important role in balancing human mood conditions. In 2016, the first 3D structures of this membrane transporter were reported in an inhibitor-bound, outward-open conformation. These structures revealed valuable information about interactions of hSERT with antidepressant drugs. Nevertheless, the question remains how serotonin facilitates the specific conformational changes that open and close pathways from the synapse and to the cytoplasm as required for transport. Here, we present a serotonin-bound homology model of hSERT in an outward-occluded state, a key intermediate in the physiological cycle, in which the interactions with the substrate are likely to be optimal. Our approach uses two template structures and includes careful refinement and comprehensive computational validation. According to microsecond-long molecular dynamics simulations, this model exhibits interactions between the gating residues in the extracellular pathway, and these interactions differ from those in an outward-open conformation of hSERT bound to serotonin. Moreover, we predict several features of this state by monitoring the intracellular gating residues, the extent of hydration, and, most importantly, protein-ligand interactions in the central binding site. The results illustrate common and distinct characteristics of these two transporter states and provide a starting point for future investigations of the transport mechanism in hSERT.

A structural model of a Ras-Raf signalosome

2020 ◽  
Author(s):  
Venkatesh P. Mysore ◽  
Zhi-Wei Zhou ◽  
Chiara Ambrogio ◽  
Lianbo Li ◽  
Jonas N. Kapp ◽  
...  
Keyword(s):  
Structural Model ◽  
Mapk Pathway ◽  
Large Body ◽  
Mapk Signaling ◽  
Effector Proteins ◽  
Ras Proteins ◽  
Starting Point ◽  
Galectin 3 ◽  
Dynamics Simulations ◽  
Composite Interfaces

AbstractThe protein K-Ras functions as a molecular switch in signaling pathways regulating cell growth. In the MAPK pathway, which is implicated in many cancers, multiple K-Ras proteins are thought to assemble at the cell membrane with Ras-effector proteins from the Raf family. Here we propose an atomistic structural model for such an assembly. Our starting point was an asymmetric, GTP-mediated K-Ras dimer model, which we generated using unbiased molecular dynamics simulations and verified with mutagenesis experiments. Adding further K-Ras monomers in a head-to-tail fashion led to a compact helical assembly, a model we validated using electron microscopy and cell-based experiments. This assembly stabilizes K-Ras in its active state and presents composite interfaces to facilitate Raf binding. Guided by existing experimental data, we then positioned C-Raf, the downstream kinase MEK1, and accessory proteins (Galectin-3 and 14-3-3σ) on the helical assembly. The resulting Ras-Raf signalosome model offers an explanation for a large body of data on MAPK signaling.

scholarly journals In vitro study of Hesperetin and Hesperidin as inhibitors of zika and chikungunya virus proteases

PLoS ONE ◽  
2021 ◽  
Vol 16 (3) ◽  
pp. e0246319
Author(s):  
Raphael J. Eberle ◽  
Danilo S. Olivier ◽  
Carolina C. Pacca ◽  
Clarita M. S. Avilla ◽  
Mauricio L. Nogueira ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Chikungunya Virus ◽  
In Vitro Study ◽  
3D Models ◽  
Binding Pocket ◽  
Starting Point ◽  
Low Cytotoxicity ◽  
Inhibitory Potential ◽  
Dynamics Simulations

The potential outcome of flavivirus and alphavirus co-infections is worrisome due to the development of severe diseases. Hundreds of millions of people worldwide live under the risk of infections caused by viruses like chikungunya virus (CHIKV, genus Alphavirus), dengue virus (DENV, genus Flavivirus), and zika virus (ZIKV, genus Flavivirus). So far, neither any drug exists against the infection by a single virus, nor against co-infection. The results described in our study demonstrate the inhibitory potential of two flavonoids derived from citrus plants: Hesperetin (HST) against NS2B/NS3pro of ZIKV and nsP2pro of CHIKV and, Hesperidin (HSD) against nsP2pro of CHIKV. The flavonoids are noncompetitive inhibitors and the determined IC50 values are in low µM range for HST against ZIKV NS2B/NS3pro (12.6 ± 1.3 µM) and against CHIKV nsP2pro (2.5 ± 0.4 µM). The IC50 for HSD against CHIKV nsP2pro was 7.1 ± 1.1 µM. The calculated ligand efficiencies for HST were > 0.3, which reflect its potential to be used as a lead compound. Docking and molecular dynamics simulations display the effect of HST and HSD on the protease 3D models of CHIKV and ZIKV. Conformational changes after ligand binding and their effect on the substrate-binding pocket of the proteases were investigated. Additionally, MTT assays demonstrated a very low cytotoxicity of both the molecules. Based on our results, we assume that HST comprise a chemical structure that serves as a starting point molecule to develop a potent inhibitor to combat CHIKV and ZIKV co-infections by inhibiting the virus proteases.

scholarly journals Computational modeling reveals a hydrophobic force-induced pore-forming mechanism for cholesterol-dependent cytolysins

2019 ◽  
Author(s):  
Shichao Pang ◽  
Junchen Yang ◽  
Jingfang Wang
Keyword(s):  
Free Energy ◽  
Conformational Changes ◽  
Structural Model ◽  
Hydrophobic Interactions ◽  
Free Energy Calculations ◽  
Forming Process ◽  
Hydrophobic Force ◽  
Forming Mechanism ◽  
Hydrophobic Pore ◽  
Dynamics Simulations

ABSTRACTDuring the pore-forming process, cholesterol-dependent cytolysins (CDCs) bind to cholesterol-rich membranes and subsequently undergo a series of conformational changes, predominantly involving in the collapse of the protein and the transformation from α helices to β-hairpins to form a large hydrophobic pore. In the current study, we reconstructed a structural model for both the prepore and pore-forming complexes of PFO based on the cryo-EM data of pneumolysin and performed molecular dynamics simulations and free energy calculations to study the conformational changes in the PFO prepore-to-pore conversion. Our simulations indicate that D2 cannot collapse spontaneously due to the hydrogen bonding and pi-pi interactions between domains D2 and D3, which are partially weakened by binding to cell membranes and oligomerization. The free energy landscape for the prepore-to-pore conversion reveals that an additional force is required for the collapse of D2 to overcome an energy barrier of ∼ 24 kcal/mol. Based on these computational results, we proposed a hydrophobic force-induced pore-forming mechanism to explain the pore-forming process of CDCs. In this mechanism, the hydrophobic interactions between the TMHs and membranes are essential for the prepore-to-pore conversion. The hydrophobic force generated by the TMHs-membrane interactions drives the conformational changes in domains D2 and D3. These findings well explain how the conformational changes within two distant domains synergistically occur, and fits well for the previous biophysical and biochemical data.

Chloride-dependent conformational changes in the GlyT1 glycine transporter

2020 ◽  
Author(s):  
Yuan-Wei Zhang ◽  
Stacy Uchendu ◽  
Vanessa Leone ◽  
Richard T. Bradshaw ◽  
Ntumba Sangwa ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Ion Pair ◽  
Cysteine Residues ◽  
Glycine Transporter ◽  
Open Conformation ◽  
Transporter Family ◽  
Transport Cycle ◽  
Dynamics Simulations ◽  
Insight Into

AbstractThe human GlyT1 glycine transporter requires chloride for its function. However, the mechanism by which Cl- exerts its influence is unknown. To examine the role that Cl- plays in the transport cycle, we measured the effect of Cl- on both glycine binding and conformational changes. The ability of glycine to displace the high-affinity radioligand [3H]CHIBA-3007 required Na+ and was potentiated over 1000-fold by Cl-. We generated GlyT1b mutants containing reactive cysteine residues in either the extracellular or cytoplasmic permeation pathways and measured changes in the reactivity of those cysteine residues as indicators of conformational changes in response to ions and substrate. Na+ increased accessibility in the extracellular pathway and decreased it in the cytoplasmic pathway, consistent with stabilizing an outward-open conformation as observed in other members of this transporter family. In the presence of Na+, both glycine and Cl- independently shifted the conformation of GlyT1b toward an outward-closed conformation. Together, Na+, glycine and Cl- stabilized an inward-open conformation of GlyT1b. We then examined whether Cl- acts by interacting with a conserved glutamine to allow formation of an ion pair that stabilizes the closed state of the extracellular pathway. Molecular dynamics simulations of a GlyT1 homologue indicated that this ion pair is formed more frequently as that pathway closes. Mutation of the glutamine blocked the effect of Cl-, and substituting it with glutamate or lysine resulted in outward- or inward-facing transporter conformations, respectively. These results provide novel and unexpected insight into the role of Cl- in this family of transporters.

scholarly journals Ligand Induced Conformational Changes of the Human Serotonin Transporter Revealed by Molecular Dynamics Simulations

PLoS ONE ◽  
2013 ◽  
Vol 8 (6) ◽  
pp. e63635 ◽  
Author(s):  
Heidi Koldsø ◽  
Henriette Elisabeth Autzen ◽  
Julie Grouleff ◽  
Birgit Schiøtt
Keyword(s):  
Molecular Dynamics ◽  
Molecular Dynamics Simulations ◽  
Serotonin Transporter ◽  
Conformational Changes ◽  
Dynamics Simulations

scholarly journals Ligand binding promiscuity of αVβ3 integrin is enlarged in response to mechanical force

2017 ◽  
Author(s):  
Michael Bachmann ◽  
Markus Schäfer ◽  
Vasyl V. Mykuliak ◽  
Marta Ripamonti ◽  
Lia Heiser ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Mechanical Load ◽  
Matrix Proteins ◽  
Αvβ3 Integrin ◽  
Superresolution Imaging ◽  
Hybrid Domain ◽  
Open Conformation ◽  
Dynamics Simulations ◽  
Integrin Ligand ◽  
Ligand Promiscuity

AbstractαVβ3 integrin recognizes multiple extracellular matrix proteins, including vitronectin (Vn) and fibronectin (Fn). However, cell experiments are frequently performed on homogenously coated substrates with only one integrin ligand present. Here, we employed binary-choice substrates of Fn and Vn to dissect αVβ3 integrin-mediated binding to both ligands on the subcellular scale. Superresolution imaging revealed that αVβ3 integrin preferred binding to Vn under various conditions. In contrast, binding to Fn required mechanical load on αVβ3 integrin. Integrin mutations, structural analysis, and molecular dynamics simulations established a model where the extended-closed conformation of αVβ3 integrin binds Vn but not Fn. Force-mediated hybrid domain swing-out characterizes the extended-open conformation needed for efficient Fn binding. Thus, force-dependent conformational changes in αVβ3 integrin increase the number of available ligands and therefore the ligand promiscuity of this integrin. These findings for αVβ3 integrin were shown to regulate cell migration and mechanotransduction differentially on Fn compared to Vn and therefore to regulate cell behavior.

scholarly journals Exploring the Early Stages of the Amyloid Aβ(1–42) Peptide Aggregation Process: An NMR Study

2021 ◽  
Vol 14 (8) ◽  
pp. 732
Author(s):  
Angelo Santoro ◽  
Manuela Grimaldi ◽  
Michela Buonocore ◽  
Ilaria Stillitano ◽  
Anna Maria D’Ursi
Keyword(s):  
Conformational Changes ◽  
Amyloid Fibrils ◽  
Structural Model ◽  
Conformational Transition ◽  
Helical Structure ◽  
Aggregation Process ◽  
Nmr Study ◽  
Aβ Aggregation ◽  
Dynamics Simulations ◽  
Β Sheet

Alzheimer’s disease (AD) is a neurodegenerative pathology characterized by the presence of neurofibrillary tangles and amyloid plaques, the latter mainly composed of Aβ(1–40) and Aβ(1–42) peptides. The control of the Aβ aggregation process as a therapeutic strategy for AD has prompted the interest to investigate the conformation of the Aβ peptides, taking advantage of computational and experimental techniques. Mixtures composed of systematically different proportions of HFIP and water have been used to monitor, by NMR, the conformational transition of the Aβ(1–42) from soluble α-helical structure to β-sheet aggregates. In the previous studies, 50/50 HFIP/water proportion emerged as the solution condition where the first evident Aβ(1–42) conformational changes occur. In the hypothesis that this solvent reproduces the best condition to catch transitional helical-β-sheet Aβ(1–42) conformations, in this study, we report an extensive NMR conformational analysis of Aβ(1–42) in 50/50 HFIP/water v/v. Aβ(1–42) structure was solved by us, giving evidence that the evolution of Aβ(1–42) peptide from helical to the β-sheet may follow unexpected routes. Molecular dynamics simulations confirm that the structural model we calculated represents a starting condition for amyloid fibrils formation.

scholarly journals Chloride-dependent conformational changes in the GlyT1 glycine transporter

2021 ◽  
Vol 118 (10) ◽  
pp. e2017431118 ◽  
Author(s):  
Yuan-Wei Zhang ◽  
Stacy Uchendu ◽  
Vanessa Leone ◽  
Richard T. Bradshaw ◽  
Ntumba Sangwa ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Ion Pair ◽  
Cysteine Residues ◽  
Glycine Transporter ◽  
Open Conformation ◽  
Transporter Family ◽  
Transport Cycle ◽  
Dynamics Simulations ◽  
Insight Into

The human GlyT1 glycine transporter requires chloride for its function. However, the mechanism by which Cl− exerts its influence is unknown. To examine the role that Cl− plays in the transport cycle, we measured the effect of Cl− on both glycine binding and conformational changes. The ability of glycine to displace the high-affinity radioligand [3H]CHIBA-3007 required Na+ and was potentiated over 1,000-fold by Cl−. We generated GlyT1b mutants containing reactive cysteine residues in either the extracellular or cytoplasmic permeation pathways and measured changes in the reactivity of those cysteine residues as indicators of conformational changes in response to ions and substrate. Na+ increased accessibility in the extracellular pathway and decreased it in the cytoplasmic pathway, consistent with stabilizing an outward-open conformation as observed in other members of this transporter family. In the presence of Na+, both glycine and Cl− independently shifted the conformation of GlyT1b toward an outward-closed conformation. Together, Na+, glycine, and Cl− stabilized an inward-open conformation of GlyT1b. We then examined whether Cl− acts by interacting with a conserved glutamine to allow formation of an ion pair that stabilizes the closed state of the extracellular pathway. Molecular dynamics simulations of a GlyT1 homolog indicated that this ion pair is formed more frequently as that pathway closes. Mutation of the glutamine blocked the effect of Cl−, and substituting it with glutamate or lysine resulted in outward- or inward-facing transporter conformations, respectively. These results provide an unexpected insight into the role of Cl− in this family of transporters.

COMPUTATIONAL STUDY OF THE DYNAMICS OF PEPTIDE DEFORMYLASE COMPLEX FROM LEPTOSPIRA INTERROGANS: EXPLORING THE CONFORMATIONAL CHANGES OF THE SUBSTRATE POCKET

2008 ◽  
Vol 07 (05) ◽  
pp. 911-922
Author(s):  
QIANG WANG ◽  
JIANWU WANG ◽  
ZHENGTING CAI ◽  
WEIREN XU
Keyword(s):  
Molecular Dynamics ◽  
Conformational Changes ◽  
Computational Study ◽  
Dynamics Simulation ◽  
Peptide Deformylase ◽  
Leptospira Interrogans ◽  
Hydrophobic Cluster ◽  
Open Conformation ◽  
Ligand Free ◽  
Dynamics Simulations

The peptide deformylase from Leptospira interrogans (LiPDF) shows many unusual characteristics. The substrate pocket of formate-bound complex adopts an open conformation. However, in the actinonin-bound LiPDF complex, a slightly open substrate pocket is observed. The opening is not large enough for the inhibitor, because the CD-loop restricts the access to the active site. To explore the conformational changes of the substrate pocket, we perform a 16,000 ps molecular dynamics simulation separately on the ligand-free LiPDF and actinonin-bound LiPDF. During the molecular dynamics simulations, extensive conformational changes have taken place. The comparison of the two MD results shows that the CD-loop, hydrophilic inhibitor, and hydrophobic cluster are necessary for the reopening of the substrate pocket. In addition, Tyr71 plays an important role in mediating the movements of CD-loop, and the transition of the substrate pocket from open to semi-open only occurs in the presence of an inhibitor, which are consistent with the experiment very well.

scholarly journals Dynamic Community Composition Unravels Allosteric Communication in PDZ3

2020 ◽  
Author(s):  
Tandac F. Guclu ◽  
Ali Rana Atilgan ◽  
Canan Atilgan
Keyword(s):  
Community Composition ◽  
Conformational Changes ◽  
Structural Changes ◽  
Distal Segment ◽  
Allosteric Communication ◽  
C Terminus ◽  
N Terminus ◽  
Ligand Interactions ◽  
Binding Cavity ◽  
Dynamics Simulations

ABSTRACTThe third domain of PSD-95 (PDZ3) is a model for investigating allosteric communication in protein and ligand interactions. While motifs contributing to its binding specificity have been scrutinized, a conformational dynamical basis is yet to be established. Despite the miniscule structural changes due to point mutants, the observed significant binding affinity differences have previously been assessed with a focus on two α-helices located at the binding groove (α2) and the C-terminus (α3). Here, we employ a new computational approach to develop a generalized view on the molecular basis of PDZ3 binding selectivity and interaction communication for a set of point mutants of the protein (G330T, H372A, G330T-H372A) and its ligand (CRIPT named L1 and its T-2F variant L2) along with the wild type (WT). To analyze the dynamical aspects hidden in the conformations that are produced by molecular dynamics simulations, we utilize variations in community composition calculated based on the betweenness centrality measure from graph theory. We find that the highly charged N-terminus which is located far from the ligand has the propensity to share the same community with the ligand in the biologically functional complexes, indicating a distal segment might mediate the binding dynamics. N- and C-termini of PDZ3 share communities, and α3 acts as a hub for the whole protein by sustaining the communication with all structural segments, albeit being a trait not unique to the functional complexes. Moreover, α2 which lines the binding cavity frequently parts communities with the ligand and is not a controller of the binding but is rather a slave to the overall dynamics coordinated by the N-terminus. Thus, ligand binding fate in PDZ3 is traced to the population of community compositions extracted from dynamics despite the lack of significant conformational changes.

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