scholarly journals Cysteine Mutagenesis and Computer Modeling of the S6 Region of an Intermediate Conductance IKCa Channel

2002 ◽  
Vol 120 (1) ◽  
pp. 99-116 ◽  
Author(s):  
Manuel Simoes ◽  
Line Garneau ◽  
Hélène Klein ◽  
Umberto Banderali ◽  
Fadi Hobeila ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Structural Data ◽  
Structural Features ◽  
K Channel ◽  
Crystallographic Structure ◽  
Channel Pore ◽  
State Dependent ◽  
Cysteine Scanning Mutagenesis ◽  
Three Dimensional Models ◽  
State Configuration

Cysteine-scanning mutagenesis (SCAM) and computer-based modeling were used to investigate key structural features of the S6 transmembrane segment of the calcium-activated K+ channel of intermediate conductance IKCa. Our SCAM results show that the interaction of [2-(trimethylammonium)ethyl] methanethiosulfonate bromide (MTSET) with cysteines engineered at positions 275, 278, and 282 leads to current inhibition. This effect was state dependent as MTSET appeared less effective at inhibiting IKCa in the closed (zero Ca2+ conditions) than open state configuration. Our results also indicate that the last four residues in S6, from A283 to A286, are entirely exposed to water in open IKCa channels, whereas MTSET can still reach the 283C and 286C residues with IKCa maintained in a closed state configuration. Notably, the internal application of MTSET or sodium (2-sulfonatoethyl) methanethiosulfonate (MTSES) caused a strong Ca2+-dependent stimulation of the A283C, V285C, and A286C currents. However, in contrast to the wild-type IKCa, the MTSET-stimulated A283C and A286C currents appeared to be TEA insensitive, indicating that the MTSET binding at positions 283 and 286 impaired the access of TEA to the channel pore. Three-dimensional structural data were next generated through homology modeling using the KcsA structure as template. In accordance with the SCAM results, the three-dimensional models predict that the V275, T278, and V282 residues should be lining the channel pore. However, the pore dimensions derived for the A283–A286 region cannot account for the MTSET effect on the closed A283C and A286 mutants. Our results suggest that the S6 domain extending from V275 to V282 possesses features corresponding to the inner cavity region of KcsA, and that the COOH terminus end of S6, from A283 to A286, is more flexible than predicted on the basis of the closed KcsA crystallographic structure alone. According to this model, closure by the gate should occur at a point located between the T278 and V282 residues.

scholarly journals Three-Dimensional Structures of Carbohydrates and Where to Find Them

2020 ◽  
Vol 21 (20) ◽  
pp. 7702 ◽  
Author(s):  
Sofya I. Scherbinina ◽  
Philip V. Toukach
Keyword(s):  
Experimental Data ◽  
Molecular Modeling ◽  
Computational Methods ◽  
Three Dimensional ◽  
Structural Diversity ◽  
Structural Data ◽  
Structural Features ◽  
Data Validation ◽  
Data Generation ◽  
Efficient Treatment

Analysis and systematization of accumulated data on carbohydrate structural diversity is a subject of great interest for structural glycobiology. Despite being a challenging task, development of computational methods for efficient treatment and management of spatial (3D) structural features of carbohydrates breaks new ground in modern glycoscience. This review is dedicated to approaches of chemo- and glyco-informatics towards 3D structural data generation, deposition and processing in regard to carbohydrates and their derivatives. Databases, molecular modeling and experimental data validation services, and structure visualization facilities developed for last five years are reviewed.

scholarly journals Membrane Protein Complex ExbB4-ExbD1-TonB1from Escherichia coli Demonstrates Conformational Plasticity

2015 ◽  
Vol 197 (11) ◽  
pp. 1873-1885 ◽  
Author(s):  
Aleksandr Sverzhinsky ◽  
Jacqueline W. Chung ◽  
Justin C. Deme ◽  
Lucien Fabre ◽  
Kristian T. Levey ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Iron Acquisition ◽  
Proton Translocation ◽  
Three Dimensional ◽  
Structural Data ◽  
Structural Features ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Content Type

ABSTRACTIron acquisition at the outer membrane (OM) of Gram-negative bacteria is powered by the proton motive force (PMF) of the cytoplasmic membrane (CM), harnessed by the CM-embedded complex of ExbB, ExbD, and TonB. Its stoichiometry, ensemble structural features, and mechanism of action are unknown. By panning combinatorial phage libraries, periplasmic regions of dimerization between ExbD and TonB were predicted. Using overexpression of full-length His6-taggedexbB-exbDand S-taggedtonB, we purified detergent-solubilized complexes of ExbB-ExbD-TonB fromEscherichia coli. Protein-detergent complexes of ∼230 kDa with a hydrodynamic radius of ∼6.0 nm were similar to previously purified ExbB4-ExbD2complexes. Significantly, they differed in electronegativity by native agarose gel electrophoresis. The stoichiometry was determined to be ExbB4-ExbD1-TonB1. Single-particle electron microscopy agrees with this stoichiometry. Two-dimensional averaging supported the phage display predictions, showing two forms of ExbD-TonB periplasmic heterodimerization: extensive and distal. Three-dimensional (3D) particle classification showed three representative conformations of ExbB4-ExbD1-TonB1. Based on our structural data, we propose a model in which ExbD shuttles a proton across the CM via an ExbB interprotein rearrangement. Proton translocation would be coupled to ExbD-mediated collapse of extended TonB in complex with ligand-loaded receptors in the OM, followed by repositioning of TonB through extensive dimerization with ExbD. Here we present the first report for purification of the ExbB-ExbD-TonB complex, molar ratios within the complex (4:1:1), and structural biology that provides insights into 3D organization.IMPORTANCEReceptors in the OM of Gram-negative bacteria allow entry of iron-bound siderophores that are necessary for pathogenicity. Numerous iron-acquisition strategies rely upon a ubiquitous and unique protein for energization: TonB. Complexed with ExbB and ExbD, the Ton system links the PMF to OM transport. Blocking iron uptake by targeting a vital nanomachine holds promise in therapeutics. Despite much research, the stoichiometry, structural arrangement, and molecular mechanism of the CM-embedded ExbB-ExbD-TonB complex remain unreported. Here we demonstratein vitroevidence of ExbB4-ExbD1-TonB1complexes. Using 3D EM, we reconstructed the complex in three conformational states that show variable ExbD-TonB heterodimerization. Our structural observations form the basis of a model for TonB-mediated iron acquisition.

scholarly journals Structure-aided prediction of mammalian transcription factor complexes in conserved non-coding elements

2013 ◽  
Vol 368 (1632) ◽  
pp. 20130029 ◽  
Author(s):  
Harendra Guturu ◽  
Andrew C. Doxey ◽  
Aaron M. Wenger ◽  
Gill Bejerano
Keyword(s):  
Transcription Factor ◽  
Binding Site ◽  
Protein Interactions ◽  
Binding Sites ◽  
Three Dimensional ◽  
Structural Data ◽  
Regulatory Elements ◽  
Computational Method ◽  
Web Resource ◽  
Three Dimensional Models

Mapping the DNA-binding preferences of transcription factor (TF) complexes is critical for deciphering the functions of cis -regulatory elements. Here, we developed a computational method that compares co-occurring motif spacings in conserved versus unconserved regions of the human genome to detect evolutionarily constrained binding sites of rigid TF complexes. Structural data were used to estimate TF complex physical plausibility, explore overlapping motif arrangements seldom tackled by non-structure-aware methods, and generate and analyse three-dimensional models of the predicted complexes bound to DNA. Using this approach, we predicted 422 physically realistic TF complex motifs at 18% false discovery rate, the majority of which (326, 77%) contain some sequence overlap between binding sites. The set of mostly novel complexes is enriched in known composite motifs, predictive of binding site configurations in TF–TF–DNA crystal structures, and supported by ChIP-seq datasets. Structural modelling revealed three cooperativity mechanisms: direct protein–protein interactions, potentially indirect interactions and ‘through-DNA’ interactions. Indeed, 38% of the predicted complexes were found to contain four or more bases in which TF pairs appear to synergize through overlapping binding to the same DNA base pairs in opposite grooves or strands. Our TF complex and associated binding site predictions are available as a web resource at http://bejerano.stanford.edu/complex .

scholarly journals Structural features of the fly chromatin colors revealed by automatic three-dimensional modeling.

2016 ◽  
Author(s):  
François Serra ◽  
Davide Baù ◽  
Guillaume Filion ◽  
Marc A. Marti-Renom
Keyword(s):  
Spatial Organization ◽  
Three Dimensional ◽  
Structural Features ◽  
Three Dimensions ◽  
Chromosome Conformation ◽  
Three Dimensional Modeling ◽  
Genomic Technologies ◽  
A Genome ◽  
Contact Data ◽  
Three Dimensional Models

The sequence of a genome is insufficient to understand all genomic processes carried out in the cell nucleus. To achieve this, the knowledge of its three- dimensional architecture is necessary. Advances in genomic technologies and the development of new analytical methods, such as Chromosome Conformation Capture (3C) and its derivatives, now permit to investigate the spatial organization of genomes. However, inferring structures from raw contact data is a tedious process for shortage of available tools. Here we present TADbit, a computational framework to analyze and model the chromatin fiber in three dimensions. To illustrate the use of TADbit, we automatically modeled 50 genomic domains from the fly genome revealing differential structural features of the previously defined chromatin colors, establishing a link between the conformation of the genome and the local chromatin composition. More generally, TADbit allows to obtain three-dimensional models ready for visualization from 3C-based experiments and to characterize their relation to gene expression and epigenetic states. TADbit is open-source and available for download from http://www.3DGenomes.org.

scholarly journals Conserved Ca2+-antagonist-binding properties and putative folding structure of a recombinant high-affinity dihydropyridine-binding domain

2000 ◽  
Vol 347 (3) ◽  
pp. 829-836 ◽  
Author(s):  
Irene HUBER ◽  
Edwin WAPPL ◽  
Alexander HERZOG ◽  
Jörg MITTERDORFER ◽  
Hartmut GLOSSMANN ◽  
...  
Keyword(s):  
Dissociation Constants ◽  
Three Dimensional ◽  
Structural Data ◽  
Binding Pocket ◽  
K Channel ◽  
Interaction Domain ◽  
Ki 67 ◽  
Reasonable Approach ◽  
Binding Residues ◽  
Ca2 Channels

Sensitivity to 1,4-dihydropyridines (DHPs) can be transferred from L-type (α1C) to non-L-type (α1A) Ca2+ channel α1 subunits by the mutation of nine pore-associated non-conserved amino acid residues, yielding mutant α1ADHP. To determine whether the hallmarks of reversible DHP binding to L-type Ca2+ channels (nanomolar dissociation constants, stereoselectivity and modulation by other chemical classes of Ca2+ antagonist drugs) were maintained in α1ADHP, we analysed the pharmacological properties of (+)-[3H]isradipine-labelled α1ADHP Ca2+ channels after heterologous expression. Binding of (+)-isradipine (Ki 7.4 nM) and the non-benzoxadiazole DHPs nifedipine (Ki 86 nM), (±)-nitrendipine (Ki 33 nM) and (±)-nimodipine (Ki 67 nM) to α1ADHP occurred at low nanomolar Ki values. DHP binding was highly stereoselective [25-fold higher affinity for (+)-isradipine]. As with native channels it was stimulated by (+)-cis-diltiazem, (+)-tetrandrine and mibefradil. This suggested that the three-dimensional architecture of the channel pore was maintained within the non-L-type α1A subunit. To predict the three-dimensional arrangement of the DHP-binding residues we exploited the X-ray structure of a recently crystallized bacterial K+ channel (KcsA) as a template. Our model is based on the assumption that the Ca2+ channel S5 and S6 segments closely resemble the KcsA transmembrane folding architecture. In the absence of three-dimensional structural data for the α1 subunit this is currently the most reasonable approach for modelling this drug-interaction domain. Our model predicts that the previously identified DHP-binding residues form a binding pocket large enough to co-ordinate a single DHP molecule. It also implies that the four homologous Ca2+ channel repeats are arranged in a clockwise manner.

A Computer Graphic Method to Enhance the Display and Interpretation of Three-Dimensional Data

1990 ◽  
Vol 48 (1) ◽  
pp. 540-541
Author(s):  
Kelly A. Dryden
Keyword(s):  
Effective Means ◽  
Three Dimensional ◽  
Image Data ◽  
Video Camera ◽  
Structural Features ◽  
Refresh Rate ◽  
Raster Graphics ◽  
Digital Equipment Corp ◽  
Complex Structural ◽  
Three Dimensional Models

Three-dimensional density information derived from electron micrographs may reveal complex structural features which are difficult to interpret from one or more two-dimensional views. Displaying a series of static views on a graphics device in quick succession provides an effective means to examine the structure dynamically and in a way which enhances the depth perception.Digital images derived from three-dimensional models are transfered to the memory of a 1280x1024 resolution (8-bit pixel) color raster graphics device (Lexidata 3400, Adage Inc., Billerica, MA) and displayed on a 19 in. color monitor at a 25-30 Hz interlaced refresh rate. Individual pixels are displayed as one of 256 different colors or grey levels from a palette of up to 224—1 combinations. An entire series of images is loaded into the graphics memory in a sequential order, usually in rows. The displayed series is zoomed up to focus on a single frame and separate frames are displayed in appropriate sequence by panning to different regions of the graphics memory. The frequency at which each frame can be displayed is adjustable and only limited by the refresh rate of the monitor. Such dynamic image sequences can be photographed with a standard video camera and replayed on a television monitor with a video cassette recorder. The programs for displaying image data are written in FORTRAN and have been implemented on a VAX/VMS 8550 minicomputer (Digital Equipment Corp., Maynard, MA).

Mechanisms of atrial fibrillation termination by rapidly unbinding Na+ channel blockers: insights from mathematical models and experimental correlates

2008 ◽  
Vol 295 (4) ◽  
pp. H1489-H1504 ◽  
Author(s):  
Philippe Comtois ◽  
Masao Sakabe ◽  
Edward J. Vigmond ◽  
Mauricio Munoz ◽  
Anne Texier ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Mathematical Models ◽  
Three Dimensional ◽  
Dominant Frequency ◽  
Na Channel ◽  
Channel Blockers ◽  
Antiarrhythmic Agents ◽  
State Dependent ◽  
Three Dimensional Models

Atrial fibrillation (AF) is the most common sustained clinical arrhythmia and is a problem of growing proportions. Recent studies have increased interest in fast-unbinding Na+ channel blockers like vernakalant (RSD1235) and ranolazine for AF therapy, but the mechanism of efficacy is poorly understood. To study how fast-unbinding INa blockers affect AF, we developed realistic mathematical models of state-dependent Na+ channel block, using a lidocaine model as a prototype, and studied the effects on simulated cholinergic AF in two- and three-dimensional atrial substrates. We then compared the results with in vivo effects of lidocaine on vagotonic AF in dogs. Lidocaine action was modeled with the Hondeghem-Katzung modulated-receptor theory and maximum affinity for activated Na+ channels. Lidocaine produced frequency-dependent Na+ channel blocking and conduction slowing effects and terminated AF in both two- and three-dimensional models with concentration-dependent efficacy (maximum ∼89% at 60 μM). AF termination was not related to increases in wavelength, which tended to decrease with the drug, but rather to decreased source Na+ current in the face of large ACh-sensitive K+ current-related sinks, leading to the destabilization of primary generator rotors and a great reduction in wavebreak, which caused primary rotor annihilations in the absence of secondary rotors to resume generator activity. Lidocaine also reduced the variability and maximum values of the dominant frequency distribution during AF. Qualitatively similar results were obtained in vivo for lidocaine effects on vagal AF in dogs, with an efficacy of 86% at 2 mg/kg iv, as well as with simulations using the guarded-receptor model of lidocaine action. These results provide new insights into the mechanisms by which rapidly unbinding class I antiarrhythmic agents, a class including several novel compounds of considerable promise, terminate AF.

scholarly journals AlphaFold-Predicted Structures of KCTD Proteins Unravel Previously Undetected Relationships among the Members of the Family

Biomolecules ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 1862
Author(s):  
Luciana Esposito ◽  
Nicole Balasco ◽  
Giovanni Smaldone ◽  
Rita Berisio ◽  
Alessia Ruggiero ◽  
...  
Keyword(s):  
Structural Information ◽  
Three Dimensional ◽  
Structural Data ◽  
Protein Family ◽  
General Sequence ◽  
Btb Domain ◽  
The Family ◽  
Three Dimensional Models ◽  
Definition Of ◽  
Sequence Similarities

One of the most striking features of KCTD proteins is their involvement in apparently unrelated yet fundamental physio-pathological processes. Unfortunately, comprehensive structure–function relationships for this protein family have been hampered by the scarcity of the structural data available. This scenario is rapidly changing due to the release of the protein three-dimensional models predicted by AlphaFold (AF). Here, we exploited the structural information contained in the AF database to gain insights into the relationships among the members of the KCTD family with the aim of facilitating the definition of the structural and molecular basis of key roles that these proteins play in many biological processes. The most important finding that emerged from this investigation is the discovery that, in addition to the BTB domain, the vast majority of these proteins also share a structurally similar domain in the C-terminal region despite the absence of general sequence similarities detectable in this region. Using this domain as reference, we generated a novel and comprehensive structure-based pseudo-phylogenetic tree that unraveled previously undetected similarities among the protein family. In particular, we generated a new clustering of the KCTD proteins that will represent a solid ground for interpreting their many functions.

scholarly journals Synthesis, magneto-structural features and thermal behavior of a new MOF

2014 ◽  
Vol 70 (a1) ◽  
pp. C1232-C1232
Author(s):  
Khadidja Aliouane ◽  
Belkacem Benmerad ◽  
Narimène Rahahlia ◽  
Hamza Kherfi ◽  
Achoura Guehria-Laïdoudi ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Structural Data ◽  
Structural Features ◽  
Crystal Structure Analysis ◽  
Magnetic Interactions ◽  
Rare Earth Complex ◽  
Polymeric Compound ◽  
Thermal Stabilities ◽  
Synthesis Conditions ◽  
Weight Losses

By applying soft synthesis conditions, in presence of a mixture that contains malonic acid, (3 mmol), lanthanide salt CeCl3.7H2O (1 mmol), alkaline-earth hydroxide Ca(OH)2 (1 mmol), and at pH below 4, we have obtained single crystals of a novel rare-earth complex which includes the dianion [L2-], {[Ce2(C3H2O4)3(H2O)3].2H2O}.The given formula was deduced from the single crystal structure analysis. The asymmetric unit corresponds to chemical formula (Figure 1). This novel polymeric compound is a three-dimensional MOF, built up from cross-linked infinite chains of one-edge-shared CeO7(H2O)3 and CeO9 polyhedrons embedding solvent molecules. Each crystallographically different metal is connected to two neighbouring ones, through four μ2-oxo bridges, to form repeat four-membered typical Ce/O/Ce/O rings. The three independant ligands show different conformations of their end functional groups: syn-syn, syn-anti, anti-anti, giving explanation for the concomitant magnetic interactions and interesting magneto-structural data. These studies are still in progress. The TG curve shows five successive weight losses. The material begins losing weight directly upon starting the thermal gravimetric experiments. It is one of the more unstable lanthanide malonates known. The correlative thermal behaviour data and X-Ray structural sub-features of the novel Ce(III)-based MOF obtained, brought out the great supramolecular effects: decarboxylation process, crystalline, and thermal stabilities are much more related to the specific infinite hydrogen patterns, than the size, shape, and embedding capability of voids accommodated in the 3D packing.

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