Insight into DEG/ENaC channel gating from genetics and structure.

Science.gov (United States)

Eastwood, Amy L; Goodman, Miriam B

2012-10-01

The founding members of the superfamily of DEG/ENaC ion channel proteins are C. elegans proteins that form mechanosensitive channels in touch and pain receptors. For more than a decade, the research community has used mutagenesis to identify motifs that regulate gating. This review integrates insight derived from unbiased in vivo mutagenesis screens with recent crystal structures to develop new models for activation of mechanically gated DEGs.

Assesment of strength and integrity of fuel channels

International Nuclear Information System (INIS)

2000-01-01

Detailed analysis to base strength and integrity of fuel channels was necessary for the licensing process. Description of tasks performed in this direction in 1999 is presented: fuel channel independent strength calculations, assessment of present fuel channels state, analysis of dynamic processes during partial group distribution header rupture, structural integrity analysis of fuel channels located next to broke channel

KV7 Channels Regulate Firing during Synaptic Integration in GABAergic Striatal Neurons

Directory of Open Access Journals (Sweden)

M. Belén Pérez-Ramírez

2015-01-01

Full Text Available Striatal projection neurons (SPNs process motor and cognitive information. Their activity is affected by Parkinson’s disease, in which dopamine concentration is decreased and acetylcholine concentration is increased. Acetylcholine activates muscarinic receptors in SPNs. Its main source is the cholinergic interneuron that responds with a briefer latency than SPNs during a cortical command. Therefore, an important question is whether muscarinic G-protein coupled receptors and their signaling cascades are fast enough to intervene during synaptic responses to regulate synaptic integration and firing. One of the most known voltage dependent channels regulated by muscarinic receptors is the KV7/KCNQ channel. It is not known whether these channels regulate the integration of suprathreshold corticostriatal responses. Here, we study the impact of cholinergic muscarinic modulation on the synaptic response of SPNs by regulating KV7 channels. We found that KV7 channels regulate corticostriatal synaptic integration and that this modulation occurs in the dendritic/spines compartment. In contrast, it is negligible in the somatic compartment. This modulation occurs on sub- and suprathreshold responses and lasts during the whole duration of the responses, hundreds of milliseconds, greatly altering SPNs firing properties. This modulation affected the behavior of the striatal microcircuit.

Corynebacterium jeikeium jk0268 constitutes for the 40 amino acid long PorACj, which forms a homooligomeric and anion-selective cell wall channel.

Directory of Open Access Journals (Sweden)

Narges Abdali

Full Text Available Corynebacterium jeikeium, a resident of human skin, is often associated with multidrug resistant nosocomial infections in immunodepressed patients. C. jeikeium K411 belongs to mycolic acid-containing actinomycetes, the mycolata and contains a channel-forming protein as judged from reconstitution experiments with artificial lipid bilayer experiments. The channel-forming protein was present in detergent treated cell walls and in extracts of whole cells using organic solvents. A gene coding for a 40 amino acid long polypeptide possibly responsible for the pore-forming activity was identified in the known genome of C. jeikeium by its similar chromosomal localization to known porH and porA genes of other Corynebacterium strains. The gene jk0268 was expressed in a porin deficient Corynebacterium glutamicum strain. For purification temporarily histidine-tailed or with a GST-tag at the N-terminus, the homogeneous protein caused channel-forming activity with an average conductance of 1.25 nS in 1M KCl identical to the channels formed by the detergent extracts. Zero-current membrane potential measurements of the voltage dependent channel implied selectivity for anions. This preference is according to single-channel analysis caused by some excess of cationic charges located in the channel lumen formed by oligomeric alpha-helical wheels. The channel has a suggested diameter of 1.4 nm as judged from the permeability of different sized hydrated anions using the Renkin correction factor. Surprisingly, the genome of C. jeikeium contained only one gene coding for a cell wall channel of the PorA/PorH type found in other Corynebacterium species. The possible evolutionary relationship between the heterooligomeric channels formed by certain Corynebacterium strains and the homooligomeric pore of C. jeikeium is discussed.

Topology of transmembrane channel-like gene 1 protein.

Science.gov (United States)

Labay, Valentina; Weichert, Rachel M; Makishima, Tomoko; Griffith, Andrew J

2010-10-05

Mutations of transmembrane channel-like gene 1 (TMC1) cause hearing loss in humans and mice. TMC1 is the founding member of a family of genes encoding proteins of unknown function that are predicted to contain multiple transmembrane domains. The goal of our study was to define the topology of mouse TMC1 expressed heterologously in tissue culture cells. TMC1 was retained in the endoplasmic reticulum (ER) membrane of five tissue culture cell lines that we tested. We used anti-TMC1 and anti-HA antibodies to probe the topologic orientation of three native epitopes and seven HA epitope tags along full-length TMC1 after selective or complete permeabilization of transfected cells with digitonin or Triton X-100, respectively. TMC1 was present within the ER as an integral membrane protein containing six transmembrane domains and cytosolic N- and C-termini. There is a large cytoplasmic loop, between the fourth and fifth transmembrane domains, with two highly conserved hydrophobic regions that might associate with or penetrate, but do not span, the plasma membrane. Our study is the first to demonstrate that TMC1 is a transmembrane protein. The topologic organization revealed by this study shares some features with that of the shaker-TRP superfamily of ion channels.

Light-activated control of protein channel assembly mediated by membrane mechanics

Science.gov (United States)

Miller, David M.; Findlay, Heather E.; Ces, Oscar; Templer, Richard H.; Booth, Paula J.

2016-12-01

Photochemical processes provide versatile triggers of chemical reactions. Here, we use a photoactivated lipid switch to modulate the folding and assembly of a protein channel within a model biological membrane. In contrast to the information rich field of water-soluble protein folding, there is only a limited understanding of the assembly of proteins that are integral to biological membranes. It is however possible to exploit the foreboding hydrophobic lipid environment and control membrane protein folding via lipid bilayer mechanics. Mechanical properties such as lipid chain lateral pressure influence the insertion and folding of proteins in membranes, with different stages of folding having contrasting sensitivities to the bilayer properties. Studies to date have relied on altering bilayer properties through lipid compositional changes made at equilibrium, and thus can only be made before or after folding. We show that light-activation of photoisomerisable di-(5-[[4-(4-butylphenyl)azo]phenoxy]pentyl)phosphate (4-Azo-5P) lipids influences the folding and assembly of the pentameric bacterial mechanosensitive channel MscL. The use of a photochemical reaction enables the bilayer properties to be altered during folding, which is unprecedented. This mechanical manipulation during folding, allows for optimisation of different stages of the component insertion, folding and assembly steps within the same lipid system. The photochemical approach offers the potential to control channel assembly when generating synthetic devices that exploit the mechanosensitive protein as a nanovalve.

Stapled Voltage-Gated Calcium Channel (CaV) α-Interaction Domain (AID) Peptides Act As Selective Protein-Protein Interaction Inhibitors of CaV Function.

Science.gov (United States)

Findeisen, Felix; Campiglio, Marta; Jo, Hyunil; Abderemane-Ali, Fayal; Rumpf, Christine H; Pope, Lianne; Rossen, Nathan D; Flucher, Bernhard E; DeGrado, William F; Minor, Daniel L

2017-06-21

For many voltage-gated ion channels (VGICs), creation of a properly functioning ion channel requires the formation of specific protein-protein interactions between the transmembrane pore-forming subunits and cystoplasmic accessory subunits. Despite the importance of such protein-protein interactions in VGIC function and assembly, their potential as sites for VGIC modulator development has been largely overlooked. Here, we develop meta-xylyl (m-xylyl) stapled peptides that target a prototypic VGIC high affinity protein-protein interaction, the interaction between the voltage-gated calcium channel (Ca V ) pore-forming subunit α-interaction domain (AID) and cytoplasmic β-subunit (Ca V β). We show using circular dichroism spectroscopy, X-ray crystallography, and isothermal titration calorimetry that the m-xylyl staples enhance AID helix formation are structurally compatible with native-like AID:Ca V β interactions and reduce the entropic penalty associated with AID binding to Ca V β. Importantly, electrophysiological studies reveal that stapled AID peptides act as effective inhibitors of the Ca V α 1 :Ca V β interaction that modulate Ca V function in an Ca V β isoform-selective manner. Together, our studies provide a proof-of-concept demonstration of the use of protein-protein interaction inhibitors to control VGIC function and point to strategies for improved AID-based Ca V modulator design.

Integral equation models for the inverse problem of biological ion channel distributions

International Nuclear Information System (INIS)

French, D A; Groetsch, C W

2007-01-01

Olfactory cilia are thin hair-like filaments that extend from olfactory receptor neurons into the nasal mucus. Transduction of an odor into an electrical signal is accomplished by a depolarizing influx of ions through cyclic-nucleotide-gated channels in the membrane that forms the lateral surface of the cilium. In an experimental procedure developed by S. Kleene, a cilium is detached at its base and drawn into a recording pipette. The cilium base is then immersed in a bath of a channel activating agent (cAMP) which is allowed to diffuse into the cilium interior, opening channels as it goes and initiating a transmembrane current. The total current is recorded as a function of time and serves as data for a nonlinear integral equation of the first kind modeling the spatial distribution of ion channels along the length of the cilium. We discuss some linear Fredholm integral equations that result from simplifications of this model. A numerical procedure is proposed for a class of integral equations suggested by this simplified model and numerical results using simulated and laboratory data are presented

Intra-membrane molecular interactions of K+ channel proteins :

Energy Technology Data Exchange (ETDEWEB)

Moczydlowski, Edward G.

2013-07-01

Ion channel proteins regulate complex patterns of cellular electrical activity and ionic signaling. Certain K+ channels play an important role in immunological biodefense mechanisms of adaptive and innate immunity. Most ion channel proteins are oligomeric complexes with the conductive pore located at the central subunit interface. The long-term activity of many K+ channel proteins is dependent on the concentration of extracellular K+; however, the mechanism is unclear. Thus, this project focused on mechanisms underlying structural stability of tetrameric K+ channels. Using KcsA of Streptomyces lividans as a model K+ channel of known structure, the molecular basis of tetramer stability was investigated by: 1. Bioinformatic analysis of the tetramer interface. 2. Effect of two local anesthetics (lidocaine, tetracaine) on tetramer stability. 3. Molecular simulation of drug docking to the ion conduction pore. The results provide new insights regarding the structural stability of K+ channels and its possible role in cell physiology.

Membrane Incorporation, Channel Formation, and Disruption of Calcium Homeostasis by Alzheimer's β-Amyloid Protein

Directory of Open Access Journals (Sweden)

Masahiro Kawahara

2011-01-01

Full Text Available Oligomerization, conformational changes, and the consequent neurodegeneration of Alzheimer's β-amyloid protein (AβP play crucial roles in the pathogenesis of Alzheimer's disease (AD. Mounting evidence suggests that oligomeric AβPs cause the disruption of calcium homeostasis, eventually leading to neuronal death. We have demonstrated that oligomeric AβPs directly incorporate into neuronal membranes, form cation-sensitive ion channels (“amyloid channels”, and cause the disruption of calcium homeostasis via the amyloid channels. Other disease-related amyloidogenic proteins, such as prion protein in prion diseases or α-synuclein in dementia with Lewy bodies, exhibit similarities in the incorporation into membranes and the formation of calcium-permeable channels. Here, based on our experimental results and those of numerous other studies, we review the current understanding of the direct binding of AβP into membrane surfaces and the formation of calcium-permeable channels. The implication of composition of membrane lipids and the possible development of new drugs by influencing membrane properties and attenuating amyloid channels for the treatment and prevention of AD is also discussed.

A protein interaction mechanism for suppressing the mechanosensitive Piezo channels.

Science.gov (United States)

Zhang, Tingxin; Chi, Shaopeng; Jiang, Fan; Zhao, Qiancheng; Xiao, Bailong

2017-11-27

Piezo proteins are bona fide mammalian mechanotransduction channels for various cell types including endothelial cells. The mouse Piezo1 of 2547 residues forms a three-bladed, propeller-like homo-trimer comprising a central pore-module and three propeller-structures that might serve as mechanotransduction-modules. However, the mechanogating and regulation of Piezo channels remain unclear. Here we identify the sarcoplasmic /endoplasmic-reticulum Ca 2+ ATPase (SERCA), including the widely expressed SERCA2, as Piezo interacting proteins. SERCA2 strategically suppresses Piezo1 via acting on a 14-residue-constituted intracellular linker connecting the pore-module and mechanotransduction-module. Mutating the linker impairs mechanogating and SERCA2-mediated modulation of Piezo1. Furthermore, the synthetic linker-peptide disrupts the modulatory effects of SERCA2, demonstrating the key role of the linker in mechanogating and regulation. Importantly, the SERCA2-mediated regulation affects Piezo1-dependent migration of endothelial cells. Collectively, we identify SERCA-mediated regulation of Piezos and the functional significance of the linker, providing important insights into the mechanogating and regulation mechanisms of Piezo channels.

Mining Protein Evolution for Insights into Mechanisms of Voltage-Dependent Sodium Channel Auxiliary Subunits.

Science.gov (United States)

Molinarolo, Steven; Granata, Daniele; Carnevale, Vincenzo; Ahern, Christopher A

2018-02-21

Voltage-gated sodium channel (VGSC) beta (β) subunits have been called the "overachieving" auxiliary ion channel subunit. Indeed, these subunits regulate the trafficking of the sodium channel complex at the plasma membrane and simultaneously tune the voltage-dependent properties of the pore-forming alpha-subunit. It is now known that VGSC β-subunits are capable of similar modulation of multiple isoforms of related voltage-gated potassium channels, suggesting that their abilities extend into the broader voltage-gated channels. The gene family for these single transmembrane immunoglobulin beta-fold proteins extends well beyond the traditional VGSC β1-β4 subunit designation, with deep roots into the cell adhesion protein family and myelin-related proteins - where inherited mutations result in a myriad of electrical signaling disorders. Yet, very little is known about how VGSC β-subunits support protein trafficking pathways, the basis for their modulation of voltage-dependent gating, and, ultimately, their role in shaping neuronal excitability. An evolutionary approach can be useful in yielding new clues to such functions as it provides an unbiased assessment of protein residues, folds, and functions. An approach is described here which indicates the greater emergence of the modern β-subunits roughly 400 million years ago in the early neurons of Bilateria and bony fish, and the unexpected presence of distant homologues in bacteriophages. Recent structural breakthroughs containing α and β eukaryotic sodium channels containing subunits suggest a novel role for a highly conserved polar contact that occurs within the transmembrane segments. Overall, a mixture of approaches will ultimately advance our understanding of the mechanism for β-subunit interactions with voltage-sensor containing ion channels and membrane proteins.

Continuous internal channels formed in aluminum fusion welds

Science.gov (United States)

Gault, J.; Sabo, W.

1967-01-01

Process produces continuous internal channel systems on a repeatable basis in 2014-T6 aluminum. Standard machining forms the initial channel, which is filled with tungsten carbide powder. TIG machine fusion welding completes formation of the channel. Chem-mill techniques enlarge it to the desired size.

Nanobody mediated crystallization of an archeal mechanosensitive channel.

Directory of Open Access Journals (Sweden)

Christian Löw

Full Text Available Mechanosensitive channels (MS are integral membrane proteins and allow bacteria to survive sudden changes in external osmolarity due to transient opening of their pores. The efflux of cytoplasmic osmolytes reduces the membrane tension and prevents membrane rupture. Therefore these channels serve as emergency valves when experiencing significant environmental stress. The preparation of high quality crystals of integral membrane proteins is a major bottleneck for structure determination by X-ray crystallography. Crystallization chaperones based on various protein scaffolds have emerged as promising tool to increase the crystallization probability of a selected target protein. So far archeal mechanosensitive channels of small conductance have resisted crystallization in our hands. To structurally analyse these channels, we selected nanobodies against an archeal MS channel after immunization of a llama with recombinant expressed, detergent solubilized and purified protein. Here we present the characterization of 23 different binders regarding their interaction with the channel protein using analytical gel filtration, western blotting and surface plasmon resonance. Selected nanobodies bound the target with affinities in the pico- to nanomolar range and some binders had a profound effect on the crystallization of the MS channel. Together with previous data we show that nanobodies are a versatile and valuable tool in structural biology by widening the crystallization space for highly challenging proteins, protein complexes and integral membrane proteins.

ABA signaling in guard cells entails a dynamic protein-protein interaction relay from the PYL-RCAR family receptors to ion channels.

Science.gov (United States)

Lee, Sung Chul; Lim, Chae Woo; Lan, Wenzhi; He, Kai; Luan, Sheng

2013-03-01

Plant hormone abscisic acid (ABA) serves as an integrator of environmental stresses such as drought to trigger stomatal closure by regulating specific ion channels in guard cells. We previously reported that SLAC1, an outward anion channel required for stomatal closure, was regulated via reversible protein phosphorylation events involving ABA signaling components, including protein phosphatase 2C members and a SnRK2-type kinase (OST1). In this study, we reconstituted the ABA signaling pathway as a protein-protein interaction relay from the PYL/RCAR-type receptors, to the PP2C-SnRK2 phosphatase-kinase pairs, to the ion channel SLAC1. The ABA receptors interacted with and inhibited PP2C phosphatase activity against the SnRK2-type kinase, releasing active SnRK2 kinase to phosphorylate, and activate the SLAC1 channel, leading to reduced guard cell turgor and stomatal closure. Both yeast two-hybrid and bimolecular fluorescence complementation assays were used to verify the interactions among the components in the pathway. These biochemical assays demonstrated activity modifications of phosphatases and kinases by their interaction partners. The SLAC1 channel activity was used as an endpoint readout for the strength of the signaling pathway, depending on the presence of different combinations of signaling components. Further study using transgenic plants overexpressing one of the ABA receptors demonstrated that changing the relative level of interacting partners would change ABA sensitivity.

Three-channel gated nanosecond integrator

International Nuclear Information System (INIS)

Tsirkel', B.I.; Martsinovskij, A.M.

1981-01-01

Structure and principle of operation of three-channel gated integrator for investigating the shape of periodical electric and optical signals at high background noise level are described. The integrator consists of an integrating circuit itself for each channel and a circuit of gating pulse formation. If the noise level doesn't exceed the signal, the value of storage capacity can be equal to 22 nF. The value of storage capacity must be increased in the case of a worse signal-to-noise ratio. The gating pulse formation circuit includes a comparator, a sawtooth voltage generator and a reference voltage generator. An integrator flowsheet is given. The time resolution of the system is about 50 ns, time sweep amounts to 5-2000 μs, electric signal sensitivity is about 70 μV. The pulse signal shape recording is performed with manual or automated time sweep at two-coordinate potentiometer. The light signal detection is made on the base of photomultiplier pulse counting rate record by the dynamic capacitor method, sensitivity limit amounts to about 1 pulse/s

Defective channels lead to an impaired skin barrier.

Science.gov (United States)

Blaydon, Diana C; Kelsell, David P

2014-10-15

Channels are integral membrane proteins that form a pore, allowing the passive movement of ions or molecules across a membrane (along a gradient), either between compartments within a cell, between intracellular and extracellular environments or between adjacent cells. The ability of cells to communicate with one another and with their environment is a crucial part of the normal physiology of a tissue that allows it to carry out its function. Cell communication is particularly important during keratinocyte differentiation and formation of the skin barrier. Keratinocytes in the skin epidermis undergo a programme of apoptosis-driven terminal differentiation, whereby proliferating keratinocytes in the basal (deepest) layer of the epidermis stop proliferating, exit the basal layer and move up through the spinous and granular layers of the epidermis to form the stratum corneum, the external barrier. Genes encoding different families of channel proteins have been found to harbour mutations linked to a variety of rare inherited monogenic skin diseases. In this Commentary, we discuss how human genetic findings in aquaporin (AQP) and transient receptor potential (TRP) channels reveal different mechanisms by which these channel proteins function to ensure the proper formation and maintenance of the skin barrier. © 2014. Published by The Company of Biologists Ltd.

Turbine component having surface cooling channels and method of forming same

Science.gov (United States)

Miranda, Carlos Miguel; Trimmer, Andrew Lee; Kottilingam, Srikanth Chandrudu

2017-09-05

A component for a turbine engine includes a substrate that includes a first surface, and an insert coupled to the substrate proximate the substrate first surface. The component also includes a channel. The channel is defined by a first channel wall formed in the substrate and a second channel wall formed by at least one coating disposed on the substrate first surface. The component further includes an inlet opening defined in flow communication with the channel. The inlet opening is defined by a first inlet wall formed in the substrate and a second inlet wall defined by the insert.

Femtosecond Laser Direct Write Integration of Multi-Protein Patterns and 3D Microstructures into 3D Glass Microfluidic Devices

Directory of Open Access Journals (Sweden)

Daniela Serien

2018-01-01

Full Text Available Microfluidic devices and biochips offer miniaturized laboratories for the separation, reaction, and analysis of biochemical materials with high sensitivity and low reagent consumption. The integration of functional or biomimetic elements further functionalizes microfluidic devices for more complex biological studies. The recently proposed ship-in-a-bottle integration based on laser direct writing allows the construction of microcomponents made of photosensitive polymer inside closed microfluidic structures. Here, we expand this technology to integrate proteinaceous two-dimensional (2D and three-dimensional (3D microstructures with the aid of photo-induced cross-linking into glass microchannels. The concept is demonstrated with bovine serum albumin and enhanced green fluorescent protein, each mixed with photoinitiator (Sodium 4-[2-(4-Morpholino benzoyl-2-dimethylamino] butylbenzenesulfonate. Unlike the polymer integration, fabrication over the entire channel cross-section is challenging. Two proteins are integrated into the same channel to demonstrate multi-protein patterning. Using 50% w/w glycerol solvent instead of 100% water achieves almost the same fabrication resolution for in-channel fabrication as on-surface fabrication due to the improved refractive index matching, enabling the fabrication of 3D microstructures. A glycerol-water solvent also reduces the risk of drying samples. We believe this technology can integrate diverse proteins to contribute to the versatility of microfluidics.

Regulation of neuronal excitability by interaction of fragile X mental retardation protein with slack potassium channels.

Science.gov (United States)

Zhang, Yalan; Brown, Maile R; Hyland, Callen; Chen, Yi; Kronengold, Jack; Fleming, Matthew R; Kohn, Andrea B; Moroz, Leonid L; Kaczmarek, Leonard K

2012-10-31

Loss of the RNA-binding protein fragile X mental retardation protein (FMRP) represents the most common form of inherited intellectual disability. Studies with heterologous expression systems indicate that FMRP interacts directly with Slack Na(+)-activated K(+) channels (K(Na)), producing an enhancement of channel activity. We have now used Aplysia bag cell (BC) neurons, which regulate reproductive behaviors, to examine the effects of Slack and FMRP on excitability. FMRP and Slack immunoreactivity were colocalized at the periphery of isolated BC neurons, and the two proteins could be reciprocally coimmunoprecipitated. Intracellular injection of FMRP lacking its mRNA binding domain rapidly induced a biphasic outward current, with an early transient tetrodotoxin-sensitive component followed by a slowly activating sustained component. The properties of this current matched that of the native Slack potassium current, which was identified using an siRNA approach. Addition of FMRP to inside-out patches containing native Aplysia Slack channels increased channel opening and, in current-clamp recordings, produced narrowing of action potentials. Suppression of Slack expression did not alter the ability of BC neurons to undergo a characteristic prolonged discharge in response to synaptic stimulation, but prevented recovery from a prolonged inhibitory period that normally follows the discharge. Recovery from the inhibited period was also inhibited by the protein synthesis inhibitor anisomycin. Our studies indicate that, in BC neurons, Slack channels are required for prolonged changes in neuronal excitability that require new protein synthesis, and raise the possibility that channel-FMRP interactions may link changes in neuronal firing to changes in protein translation.

Integrating Flow, Form, and Function for Improved Environmental Water Management

Science.gov (United States)

Albin Lane, Belize Arela

Rivers are complex, dynamic natural systems. The performance of river ecosystem functions, such as habitat availability and sediment transport, depends on the interplay of hydrologic dynamics (flow) and geomorphic settings (form). However, most river restoration studies evaluate the role of either flow or form without regard for their dynamic interactions. Despite substantial recent interest in quantifying environmental water requirements to support integrated water management efforts, the absence of quantitative, transferable relationships between river flow, form, and ecosystem functions remains a major limitation. This research proposes a novel, process-driven methodology for evaluating river flow-form-function linkages in support of basin-scale environmental water management. This methodology utilizes publically available geospatial and time-series data and targeted field data collection to improve basic understanding of river systems with limited data and resource requirements. First, a hydrologic classification system is developed to characterize natural hydrologic variability across a highly altered, physio-climatically diverse landscape. Next, a statistical analysis is used to characterize reach-scale geomorphic variability and to investigate the utility of topographic variability attributes (TVAs, subreach-scale undulations in channel width and depth), alongside traditional reach-averaged attributes, for distinguishing dominant geomorphic forms and processes across a hydroscape. Finally, the interacting roles of flow (hydrologic regime, water year type, and hydrologic impairment) and form (channel morphology) are quantitatively evaluated with respect to ecosystem functions related to hydrogeomorphic processes, aquatic habitat, and riparian habitat. Synthetic river corridor generation is used to evaluate and isolate the role of distinct geomorphic attributes without the need for intensive topographic surveying. This three-part methodology was successfully

Cooperative endocytosis of the endosomal SNARE protein syntaxin-8 and the potassium channel TASK-1

Science.gov (United States)

Renigunta, Vijay; Fischer, Thomas; Zuzarte, Marylou; Kling, Stefan; Zou, Xinle; Siebert, Kai; Limberg, Maren M.; Rinné, Susanne; Decher, Niels; Schlichthörl, Günter; Daut, Jürgen

2014-01-01

The endosomal SNARE protein syntaxin-8 interacts with the acid-sensitive potassium channel TASK-1. The functional relevance of this interaction was studied by heterologous expression of these proteins (and mutants thereof) in Xenopus oocytes and in mammalian cell lines. Coexpression of syntaxin-8 caused a fourfold reduction in TASK-1 current, a corresponding reduction in the expression of TASK-1 at the cell surface, and a marked increase in the rate of endocytosis of the channel. TASK-1 and syntaxin-8 colocalized in the early endosomal compartment, as indicated by the endosomal markers 2xFYVE and rab5. The stimulatory effect of the SNARE protein on the endocytosis of the channel was abolished when both an endocytosis signal in TASK-1 and an endocytosis signal in syntaxin-8 were mutated. A syntaxin-8 mutant that cannot assemble with other SNARE proteins had virtually the same effect as wild-type syntaxin-8. Total internal reflection fluorescence microscopy showed formation and endocytosis of vesicles containing fluorescence-tagged clathrin, TASK-1, and/or syntaxin-8. Our results suggest that the unassembled form of syntaxin-8 and the potassium channel TASK-1 are internalized via clathrin-mediated endocytosis in a cooperative manner. This implies that syntaxin-8 regulates the endocytosis of TASK-1. Our study supports the idea that endosomal SNARE proteins can have functions unrelated to membrane fusion. PMID:24743596

Monolithically integrated 8-channel WDM reflective modulator

NARCIS (Netherlands)

Stopinski, S.T.; Malinowski, M.; Piramidowicz, R.; Smit, M.K.; Leijtens, X.J.M.

2013-01-01

In this work the design and characterization of a monolithically integrated photonic circuit acting as a reflective modulator for eight WDM channels is presented. The chip was designed and fabricated in a generic integration technology

Continuous production of core-shell protein nanoparticles by antisolvent precipitation using dual-channel microfluidization: Caseinate-coated zein nanoparticles.

Science.gov (United States)

Ebert, Sandra; Koo, Charmaine K W; Weiss, Jochen; McClements, David Julian

2017-02-01

Antisolvent precipitation is commonly used to fabricate protein nanoparticles using a simple batch method that involves injecting a protein-solvent mixture into an antisolvent. In this study, the potential of producing core-shell protein nanoparticles by antisolvent precipitation using a continuous dual-channel microfluidization method was investigated. The solvent phase (zein in ethanol) and antisolvent phase (casein in water) were made to impinge on each other at high velocity, which generates intense shear, turbulent, and cavitation forces that ensure thorough mixing and breakup of the phases. Relatively small core-shell protein nanoparticles (dnanoparticles went from positive at low pH to negative at high pH, with a point of zero charge around pH5. Electron microscopy indicated that the protein particles formed had a roughly spherical shape. The results suggest that the dual-channel microfluidizer could be used to continuously form protein nanoparticles by antisolvent precipitation. Nevertheless, when the microfluidization method was compared with the simple batch method the size of the particles produced under similar conditions were fairly similar. Copyright © 2016 Elsevier Ltd. All rights reserved.

Involvement of TRPV3 and TRPM8 ion channel proteins in induction of mammalian cold-inducible proteins.

Science.gov (United States)

Fujita, Takanori; Liu, Yu; Higashitsuji, Hiroaki; Itoh, Katsuhiko; Shibasaki, Koji; Fujita, Jun; Nishiyama, Hiroyuki

2018-01-01

Cold-inducible RNA-binding protein (CIRP), RNA-binding motif protein 3 (RBM3) and serine and arginine rich splicing factor 5 (SRSF5) are RNA-binding proteins that are transcriptionally upregulated in response to moderately low temperatures and a variety of cellular stresses in mammalian cells. Induction of these cold-inducible proteins (CIPs) is dependent on transient receptor potential (TRP) V4 channel protein, but seems independent of its ion channel activity. We herein report that in addition to TRPV4, TRPV3 and TRPM8 are necessary for the induction of CIPs. We established cell lines from the lung of TRPV4-knockout (KO) mouse, and observed induction of CIPs in them by western blot analysis. A TRPV4 antagonist RN1734 suppressed the induction in wild-type mouse cells, but not in TRPV4-KO cells. A TRPV3 channel blocker S408271 and a TRPM8 channel blocker AMTB as well as siRNAs against TRPV3 and TRPM8 suppressed the CIP induction in mouse TRPV4-KO cells and human U-2 OS cells. A TRPV3 channel agonist 2-APB induced CIP expression, but camphor did not. Neither did a TRPM8 channel agonist WS-12. These results suggest that TRPV4, TRPV3 and TRPM8 proteins, but not their ion channel activities are necessary for the induction of CIPs at 32 °C. Identification of proteins that differentially interact with these TRP channels at 37 °C and 32 °C would help elucidate the underlying mechanisms of CIP induction by hypothermia. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Conformational changes and slow dynamics through microsecond polarized atomistic molecular simulation of an integral Kv1.2 ion channel

DEFF Research Database (Denmark)

Bjelkmar, Pär; Niemelä, Perttu S; Vattulainen, Ilpo

2009-01-01

transitions occur in membrane proteins-not to mention numerous applications in drug design. Here, we present a full 1 micros atomic-detail molecular dynamics simulation of an integral Kv1.2 ion channel, comprising 120,000 atoms. By applying 0.052 V/nm of hyperpolarization, we observe structural rearrangements......Structure and dynamics of voltage-gated ion channels, in particular the motion of the S4 helix, is a highly interesting and hotly debated topic in current membrane protein research. It has critical implications for insertion and stabilization of membrane proteins as well as for finding how...... and significant thinning of the membrane also observed in experiments, this provides additional support for the predictive power of microsecond-scale membrane protein simulations....

Naked mole-rat acid-sensing ion channel 3 forms nonfunctional homomers, but functional heteromers.

Science.gov (United States)

Schuhmacher, Laura-Nadine; Callejo, Gerard; Srivats, Shyam; Smith, Ewan St John

2018-02-02

Acid-sensing ion channels (ASICs) form both homotrimeric and heterotrimeric ion channels that are activated by extracellular protons and are involved in a wide range of physiological and pathophysiological processes, including pain and anxiety. ASIC proteins can form both homotrimeric and heterotrimeric ion channels. The ASIC3 subunit has been shown to be of particular importance in the peripheral nervous system with pharmacological and genetic manipulations demonstrating a role in pain. Naked mole-rats, despite having functional ASICs, are insensitive to acid as a noxious stimulus and show diminished avoidance of acidic fumes, ammonia, and carbon dioxide. Here we cloned naked mole-rat ASIC3 (nmrASIC3) and used a cell-surface biotinylation assay to demonstrate that it traffics to the plasma membrane, but using whole-cell patch clamp electrophysiology we observed that nmrASIC3 is insensitive to both protons and the non-proton ASIC3 agonist 2-guanidine-4-methylquinazoline. However, in line with previous reports of ASIC3 mRNA expression in dorsal root ganglia neurons, we found that the ASIC3 antagonist APETx2 reversibly inhibits ASIC-like currents in naked mole-rat dorsal root ganglia neurons. We further show that like the proton-insensitive ASIC2b and ASIC4, nmrASIC3 forms functional, proton-sensitive heteromers with other ASIC subunits. An amino acid alignment of ASIC3s between 9 relevant rodent species and human identified unique sequence differences that might underlie the proton insensitivity of nmrASIC3. However, introducing nmrASIC3 differences into rat ASIC3 (rASIC3) produced only minor differences in channel function, and replacing the nmrASIC3 sequence with that of rASIC3 did not produce a proton-sensitive ion channel. Our observation that nmrASIC3 forms nonfunctional homomers may reflect a further adaptation of the naked mole-rat to living in an environment with high-carbon dioxide levels. © 2018 by The American Society for Biochemistry and Molecular

The Topographic Design of River Channels for Form-Process Linkages.

Science.gov (United States)

Brown, Rocko A; Pasternack, Gregory B; Lin, Tin

2016-04-01

Scientists and engineers design river topography for a wide variety of uses, such as experimentation, site remediation, dam mitigation, flood management, and river restoration. A recent advancement has been the notion of topographical design to yield specific fluvial mechanisms in conjunction with natural or environmental flow releases. For example, the flow convergence routing mechanism, whereby shear stress and spatially convergent flow migrate or jump from the topographic high (riffle) to the low point (pool) from low to high discharge, is thought to be a key process able to maintain undular relief in gravel bedded rivers. This paper develops an approach to creating riffle-pool topography with a form-process linkage to the flow convergence routing mechanism using an adjustable, quasi equilibrium synthetic channel model. The link from form to process is made through conceptualizing form-process relationships for riffle-pool couplets into geomorphic covariance structures (GCSs) that are then quantitatively embedded in a synthetic channel model. Herein, GCSs were used to parameterize a geometric model to create five straight, synthetic river channels with varying combinations of bed and width undulations. Shear stress and flow direction predictions from 2D hydrodynamic modeling were used to determine if scenarios recreated aspects of the flow convergence routing mechanism. Results show that the creation of riffle-pool couplets that experience flow convergence in straight channels requires GCSs with covarying bed and width undulations in their topography as supported in the literature. This shows that GCSs are a useful way to translate conceptualizations of form-process linkages into quantitative models of channel form.

Mapping of Residues Forming the Voltage Sensor of the Voltage-Dependent Anion-Selective Channel

Science.gov (United States)

Thomas, Lorie; Blachly-Dyson, Elizabeth; Colombini, Marco; Forte, Michael

1993-06-01

Voltage-gated ion-channel proteins contain "voltage-sensing" domains that drive the conformational transitions between open and closed states in response to changes in transmembrane voltage. We have used site-directed mutagenesis to identify residues affecting the voltage sensitivity of a mitochondrial channel, the voltage-dependent anion-selective channel (VDAC). Although charge changes at many sites had no effect, at other sites substitutions that increased positive charge also increased the steepness of voltage dependance and substitutions that decreased positive charge decreased voltage dependance by an appropriate amount. In contrast to the plasma membrane K^+ and Na^+ channels, these residues are distributed over large parts of the VDAC protein. These results have been used to define the conformational transitions that accompany voltage gating of an ion channel. This gating mechanism requires the movement of large portions of the VDAC protein through the membrane.

The small envelope protein of porcine reproductive and respiratory syndrome virus possesses ion channel protein-like properties

International Nuclear Information System (INIS)

Lee, Changhee; Yoo, Dongwan

2006-01-01

The small envelope (E) protein of porcine reproductive and respiratory syndrome virus (PRRSV) is a hydrophobic 73 amino acid protein encoded in the internal open reading frame (ORF) of the bicistronic mRNA2. As a first step towards understanding the biological role of E protein during PRRSV replication, E gene expression was blocked in a full-length infectious clone by mutating the ATG translational initiation to GTG, such that the full-length mutant genomic clone was unable to synthesize the E protein. DNA transfection of PRRSV-susceptible cells with the E gene knocked-out genomic clone showed the absence of virus infectivity. P129-ΔE-transfected cells however produced virion particles in the culture supernatant, and these particles contained viral genomic RNA, demonstrating that the E protein is essential for PRRSV infection but dispensable for virion assembly. Electron microscopy suggests that the P129-ΔE virions assembled in the absence of E had a similar appearance to the wild-type particles. Strand-specific RT-PCR demonstrated that the E protein-negative, non-infectious P129-ΔE virus particles were able to enter cells but further steps of replication were interrupted. The entry of PRRSV has been suggested to be via receptor-mediated endocytosis, and lysomotropic basic compounds and known ion-channel blocking agents both inhibited PRRSV replication effectively during the uncoating process. The expression of E protein in Escherichia coli-mediated cell growth arrests and increased the membrane permeability. Cross-linking experiments in cells infected with PRRSV or transfected with E gene showed that the E protein was able to form homo-oligomers. Taken together, our data suggest that the PRRSV E protein is likely an ion-channel protein embedded in the viral envelope and facilitates uncoating of virus and release of the genome in the cytoplasm

Multi-channel time-division integrator in HL-2A

International Nuclear Information System (INIS)

Yan Ji

2008-01-01

HL-2A is China's first Tokamak device with divertor configuration (magnetic confinement controlled nuclear fusion device). To find out the details of on-going fusion reaction at different times is of important significance in achieving controlled nuclear fusion. We developed a new type multi-channel time-division integrator for HL-2A. It has functions of automatic cutting off negative pulse of the input signals, optional integrating time division spacing 0.2-1 ms, TTL starting trigger signal, automatic regularly work 20 s, and integrating 10 channel at the same time. (authors)

A Lys-Trp cation-π interaction mediates the dimerization and function of the chloride intracellular channel protein 1 transmembrane domain.

Science.gov (United States)

Peter, Bradley; Polyansky, Anton A; Fanucchi, Sylvia; Dirr, Heini W

2014-01-14

Chloride intracellular channel protein 1 (CLIC1) is a dual-state protein that can exist either as a soluble monomer or in an integral membrane form. The oligomerization of the transmembrane domain (TMD) remains speculative despite it being implicated in pore formation. The extent to which electrostatic and van der Waals interactions drive folding and association of the dimorphic TMD is unknown and is complicated by the requirement of interactions favorable in both aqueous and membrane environments. Here we report a putative Lys37-Trp35 cation-π interaction and show that it stabilizes the dimeric form of the CLIC1 TMD in membranes. A synthetic 30-mer peptide comprising a K37M TMD mutant was examined in 2,2,2-trifluoroethanol, sodium dodecyl sulfate micelles, and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine liposomes using far-ultraviolet (UV) circular dichroism, fluorescence, and UV absorbance spectroscopy. Our data suggest that Lys37 is not implicated in the folding, stability, or membrane insertion of the TMD peptide. However, removal of this residue impairs the formation of dimers and higher-order oligomers. This is accompanied by a 30-fold loss of chloride influx activity, suggesting that dimerization modulates the rate of chloride conductance. We propose that, within membranes, individual TMD helices associate via a Lys37-mediated cation-π interaction to form active dimers. The latter findings are also supported by results of modeling a putative TMD dimer conformation in which Lys37 and Trp35 form cation-π pairs at the dimer interface. Dimeric helix bundles may then associate to form fully active ion channels. Thus, within a membrane-like environment, aromatic interactions involving a polar lysine side chain provide a thermodynamic driving force for helix-helix association.

The gap junction protein connexin43 interacts with the second PDZ domain of the zona occludens-1 protein

NARCIS (Netherlands)

Giepmans, B N; Moolenaar, W H

1998-01-01

Gap junctions mediate cell-cell communication in almost all tissues and are composed of channel-forming integral membrane proteins, termed connexins [1-3]. Connexin43 (Cx43) is the most widely expressed and the most well-studied member of this family. Cx43-based cell-cell communication is regulated

Sedimentation equilibrium of a small oligomer-forming membrane protein: effect of histidine protonation on pentameric stability.

Science.gov (United States)

Surya, Wahyu; Torres, Jaume

2015-04-02

Analytical ultracentrifugation (AUC) can be used to study reversible interactions between macromolecules over a wide range of interaction strengths and under physiological conditions. This makes AUC a method of choice to quantitatively assess stoichiometry and thermodynamics of homo- and hetero-association that are transient and reversible in biochemical processes. In the modality of sedimentation equilibrium (SE), a balance between diffusion and sedimentation provides a profile as a function of radial distance that depends on a specific association model. Herein, a detailed SE protocol is described to determine the size and monomer-monomer association energy of a small membrane protein oligomer using an analytical ultracentrifuge. AUC-ES is label-free, only based on physical principles, and can be used on both water soluble and membrane proteins. An example is shown of the latter, the small hydrophobic (SH) protein in the human respiratory syncytial virus (hRSV), a 65-amino acid polypeptide with a single α-helical transmembrane (TM) domain that forms pentameric ion channels. NMR-based structural data shows that SH protein has two protonatable His residues in its transmembrane domain that are oriented facing the lumen of the channel. SE experiments have been designed to determine how pH affects association constant and the oligomeric size of SH protein. While the pentameric form was preserved in all cases, its association constant was reduced at low pH. These data are in agreement with a similar pH dependency observed for SH channel activity, consistent with a lumenal orientation of the two His residues in SH protein. The latter may experience electrostatic repulsion and reduced oligomer stability at low pH. In summary, this method is applicable whenever quantitative information on subtle protein-protein association changes in physiological conditions have to be measured.

General form of genuine multipartite entanglement quantum channels for teleportation

International Nuclear Information System (INIS)

Chen Pingxing; Zhu Shiyao; Guo, Guangcan

2006-01-01

Recently Yeo and Chua [Phys. Rev. Lett. 96, 060502 (2006)] presented an explicit protocol for faithfully teleporting an arbitrary two-qubit state via a genuine four-qubit entanglement channel. Here we generalize completely their results to teleporting an arbitrary N-qubit state via genuine N-qubit entanglement channels. And we present the general form of the genuine multipartite entanglement channels, namely, the sufficient and necessary condition the genuine N-qubit entanglement channels must satisfy to teleport an arbitrary N-qubit state

Magnetic field integral equation analysis of surface plasmon scattering by rectangular dielectric channel discontinuities.

Science.gov (United States)

Chremmos, Ioannis

2010-01-01

The scattering of a surface plasmon polariton (SPP) by a rectangular dielectric channel discontinuity is analyzed through a rigorous magnetic field integral equation method. The scattering phenomenon is formulated by means of the magnetic-type scalar integral equation, which is subsequently treated through an entire-domain Galerkin method of moments (MoM), based on a Fourier-series plane wave expansion of the magnetic field inside the discontinuity. The use of Green's function Fourier transform allows all integrations over the area and along the boundary of the discontinuity to be performed analytically, resulting in a MoM matrix with entries that are expressed as spectral integrals of closed-form expressions. Complex analysis techniques, such as Cauchy's residue theorem and the saddle-point method, are applied to obtain the amplitudes of the transmitted and reflected SPP modes and the radiated field pattern. Through numerical results, we examine the wavelength selectivity of transmission and reflection against the channel dimensions as well as the sensitivity to changes in the refractive index of the discontinuity, which is useful for sensing applications.

Brain-derived neurotrophic factor modulation of Kv1.3 channel is disregulated by adaptor proteins Grb10 and nShc

Directory of Open Access Journals (Sweden)

Marks David R

2009-01-01

Full Text Available Abstract Background Neurotrophins are important regulators of growth and regeneration, and acutely, they can modulate the activity of voltage-gated ion channels. Previously we have shown that acute brain-derived neurotrophic factor (BDNF activation of neurotrophin receptor tyrosine kinase B (TrkB suppresses the Shaker voltage-gated potassium channel (Kv1.3 via phosphorylation of multiple tyrosine residues in the N and C terminal aspects of the channel protein. It is not known how adaptor proteins, which lack catalytic activity, but interact with members of the neurotrophic signaling pathway, might scaffold with ion channels or modulate channel activity. Results We report the co-localization of two adaptor proteins, neuronal Src homology and collagen (nShc and growth factor receptor-binding protein 10 (Grb10, with Kv1.3 channel as demonstrated through immunocytochemical approaches in the olfactory bulb (OB neural lamina. To further explore the specificity and functional ramification of adaptor/channel co-localization, we performed immunoprecipitation and Western analysis of channel, kinase, and adaptor transfected human embryonic kidney 293 cells (HEK 293. nShc formed a direct protein-protein interaction with Kv1.3 that was independent of BDNF-induced phosphorylation of Kv1.3, whereas Grb10 did not complex with Kv1.3 in HEK 293 cells. Both adaptors, however, co-immunoprecipitated with Kv1.3 in native OB. Grb10 was interestingly able to decrease the total expression of Kv1.3, particularly at the membrane surface, and subsequently eliminated the BDNF-induced phosphorylation of Kv1.3. To examine the possibility that the Src homology 2 (SH2 domains of Grb10 were directly binding to basally phosphorylated tyrosines in Kv1.3, we utilized point mutations to substitute multiple tyrosine residues with phenylalanine. Removal of the tyrosines 111–113 and 449 prevented Grb10 from decreasing Kv1.3 expression. In the absence of either adaptor protein

Integrated Analysis of Flow, Form, and Function for River Management and Design Testing

Science.gov (United States)

Lane, B. A. A.; Pasternack, G. B.; Sandoval Solis, S.

2017-12-01

Rivers are highly complex, dynamic systems that support numerous ecosystem functions including transporting sediment, modulating biogeochemical processes, and regulating habitat availability for native species. The extent and timing of these functions is largely controlled by the interplay of hydrologic dynamics (i.e. flow) and the shape and composition of the river corridor (i.e. form). This study applies synthetic channel design to the evaluation of river flow-form-function linkages, with the aim of evaluating these interactions across a range of flows and forms to inform process-driven management efforts with limited data and financial requirements. In an application to California's Mediterranean-montane streams, the interacting roles of channel form, water year type, and hydrologic impairment were evaluated across a suite of ecosystem functions related to hydrogeomorphic processes, aquatic habitat, and riparian habitat. Channel form acted as the dominant control on hydrogeomorphic processes considered, while water year type controlled salmonid habitat functions. Streamflow alteration for hydropower increased redd dewatering risk and altered aquatic habitat availability and riparian recruitment dynamics. Study results highlight critical tradeoffs in ecosystem function performance and emphasize the significance of spatiotemporal diversity of flow and form at multiple scales for maintaining river ecosystem integrity. The approach is broadly applicable and extensible to other systems and ecosystem functions, where findings can be used to characterize complex controls on river ecosystems, assess impacts of proposed flow and form alterations, and inform river restoration strategies.

The Blurred Line between Form and Process: A Comparison of Stream Channel Classification Frameworks.

Directory of Open Access Journals (Sweden)

Alan Kasprak

Full Text Available Stream classification provides a means to understand the diversity and distribution of channels and floodplains that occur across a landscape while identifying links between geomorphic form and process. Accordingly, stream classification is frequently employed as a watershed planning, management, and restoration tool. At the same time, there has been intense debate and criticism of particular frameworks, on the grounds that these frameworks classify stream reaches based largely on their physical form, rather than direct measurements of their component hydrogeomorphic processes. Despite this debate surrounding stream classifications, and their ongoing use in watershed management, direct comparisons of channel classification frameworks are rare. Here we implement four stream classification frameworks and explore the degree to which each make inferences about hydrogeomorphic process from channel form within the Middle Fork John Day Basin, a watershed of high conservation interest within the Columbia River Basin, U.S.A. We compare the results of the River Styles Framework, Natural Channel Classification, Rosgen Classification System, and a channel form-based statistical classification at 33 field-monitored sites. We found that the four frameworks consistently classified reach types into similar groups based on each reach or segment's dominant hydrogeomorphic elements. Where classified channel types diverged, differences could be attributed to the (a spatial scale of input data used, (b the requisite metrics and their order in completing a framework's decision tree and/or, (c whether the framework attempts to classify current or historic channel form. Divergence in framework agreement was also observed at reaches where channel planform was decoupled from valley setting. Overall, the relative agreement between frameworks indicates that criticism of individual classifications for their use of form in grouping stream channels may be overstated. These

TRP channels in kidney disease.

NARCIS (Netherlands)

Hsu, Y.J.; Hoenderop, J.G.J.; Bindels, R.J.M.

2007-01-01

Mammalian TRP channel proteins form six-transmembrane cation-permeable channels that may be grouped into six subfamilies on the basis of amino acid sequence homology (TRPC, TRPV, TRPM, TRPA, TRPP, and TRPML). Recent studies of TRP channels indicate that they are involved in numerous fundamental cell

A structural view of ligand-dependent activation in thermoTRP channels

Directory of Open Access Journals (Sweden)

Ximena eSteinberg

2014-05-01

Full Text Available Transient Receptor Potential (TRP proteins are a large family of ion channels, grouped intoseven sub-families. Although great advances have been made regarding the activation andmodulation of TRP channel activity, detailed molecular mechanisms governing TRPchannel gating are still needed. Sensitive to electric, chemical, mechanical, and thermalcues, TRP channels are tightly associated with the detection and integration of sensoryinput, emerging as a model to study the polymodal activation of ion channel proteins.Among TRP channels, the temperature-activated kind constitute a subgroup by itself,formed by Vanilloid receptors 1-4, Melastatin receptors 2, 4, 5 and 8, TRPC5, and TRPA1.Some of the so-called thermoTRP channels participate in the detection of noxious stimulimaking them an interesting pharmacological target for the treatment of pain. However, thepoor specificity of the compounds available in the market represents an important obstacleto overcome. Understanding the molecular mechanics underlying ligand-dependentmodulation of TRP channels may help with the rational design of novel syntheticanalgesics. The present review focuses on the structural basis of ligand-dependentactivation of TRPV1 and TRPM8 channels. Special attention is drawn to the dissection ofligand-binding sites within TRPV1, PIP 2 -dependent modulation of TRP channels, and thestructure of natural and synthetic ligands.

Molecular Characterization of LRB7 Gene and a Water Channel Protein TIP2 in Chorispora bungeana

Directory of Open Access Journals (Sweden)

Ming Li

2016-01-01

Full Text Available Background. Water channel proteins, also called aquaporins, are integral membrane proteins from major intrinsic protein (MIP family and involved in several pathways including not only water transport but also cell signaling, reproduction, and photosynthesis. The full cDNA and protein sequences of aquaporin in Chorispora bungeana Fisch. & C.A. Mey (C. bungeana are still unknown. Results. In this study, PCR and rapid amplification of cDNA ends approaches were used to clone the full cDNA of LRB7 (GenBank accession number: EU636988 of C. bungeana. Sequence analysis indicated that it was 1235 bp, which had two introns and encoded a protein of 250 amino acids. Structure analysis revealed that the protein had two conserved NPA motifs, one of which is MIP signature sequence (SGxHxNPAVT, six membrane helix regions, and additional membrane-embedded domains. Phylogenetic analysis suggested that the protein was from TIP2 subgroup. Surprisingly, semiquantitative RT-PCR experiment and western blot analysis showed that LRB7 and TIP2 were only detectable in roots, unlike Arabidopsis and Raphanus. Connecting with our previous studies, LRB7 was supported to associate with chilling-tolerance in C. bungeana. Conclusion. This is the first time to characterize the full sequences of LRB7 gene and water channel protein in C. bungeana. Our findings contribute to understanding the water transports in plants under low temperatures.

Beyond voltage-gated ion channels: Voltage-operated membrane proteins and cellular processes.

Science.gov (United States)

Zhang, Jianping; Chen, Xingjuan; Xue, Yucong; Gamper, Nikita; Zhang, Xuan

2018-04-18

Voltage-gated ion channels were believed to be the only voltage-sensitive proteins in excitable (and some non-excitable) cells for a long time. Emerging evidence indicates that the voltage-operated model is shared by some other transmembrane proteins expressed in both excitable and non-excitable cells. In this review, we summarize current knowledge about voltage-operated proteins, which are not classic voltage-gated ion channels as well as the voltage-dependent processes in cells for which single voltage-sensitive proteins have yet to be identified. Particularly, we will focus on the following. (1) Voltage-sensitive phosphoinositide phosphatases (VSP) with four transmembrane segments homologous to the voltage sensor domain (VSD) of voltage-gated ion channels; VSPs are the first family of proteins, other than the voltage-gated ion channels, for which there is sufficient evidence for the existence of the VSD domain; (2) Voltage-gated proton channels comprising of a single voltage-sensing domain and lacking an identified pore domain; (3) G protein coupled receptors (GPCRs) that mediate the depolarization-evoked potentiation of Ca 2+ mobilization; (4) Plasma membrane (PM) depolarization-induced but Ca 2+ -independent exocytosis in neurons. (5) Voltage-dependent metabolism of phosphatidylinositol 4,5-bisphosphate (PtdIns[4,5]P 2 , PIP 2 ) in the PM. These recent discoveries expand our understanding of voltage-operated processes within cellular membranes. © 2018 Wiley Periodicals, Inc.

Fragile X mental retardation protein controls ion channel expression and activity.

Science.gov (United States)

Ferron, Laurent

2016-10-15

Fragile X-associated disorders are a family of genetic conditions resulting from the partial or complete loss of fragile X mental retardation protein (FMRP). Among these disorders is fragile X syndrome, the most common cause of inherited intellectual disability and autism. FMRP is an RNA-binding protein involved in the control of local translation, which has pleiotropic effects, in particular on synaptic function. Analysis of the brain FMRP transcriptome has revealed hundreds of potential mRNA targets encoding postsynaptic and presynaptic proteins, including a number of ion channels. FMRP has been confirmed to bind voltage-gated potassium channels (K v 3.1 and K v 4.2) mRNAs and regulates their expression in somatodendritic compartments of neurons. Recent studies have uncovered a number of additional roles for FMRP besides RNA regulation. FMRP was shown to directly interact with, and modulate, a number of ion channel complexes. The sodium-activated potassium (Slack) channel was the first ion channel shown to directly interact with FMRP; this interaction alters the single-channel properties of the Slack channel. FMRP was also shown to interact with the auxiliary β4 subunit of the calcium-activated potassium (BK) channel; this interaction increases calcium-dependent activation of the BK channel. More recently, FMRP was shown to directly interact with the voltage-gated calcium channel, Ca v 2.2, and reduce its trafficking to the plasma membrane. Studies performed on animal models of fragile X syndrome have revealed links between modifications of ion channel activity and changes in neuronal excitability, suggesting that these modifications could contribute to the phenotypes observed in patients with fragile X-associated disorders. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

Monolithic integration of microfluidic channels and semiconductor lasers

Science.gov (United States)

Cran-McGreehin, Simon J.; Dholakia, Kishan; Krauss, Thomas F.

2006-08-01

We present a fabrication method for the monolithic integration of microfluidic channels into semiconductor laser material. Lasers are designed to couple directly into the microfluidic channel, allowing submerged particles pass through the output beams of the lasers. The interaction between particles in the channel and the lasers, operated in either forward or reverse bias, allows for particle detection, and the optical forces can be used to trap and move particles. Both interrogation and manipulation are made more amenable for lab-on-a-chip applications through monolithic integration. The devices are very small, they require no external optical components, have perfect intrinsic alignment, and can be created with virtually any planar configuration of lasers in order to perform a variety of tasks. Their operation requires no optical expertise and only low electrical power, thus making them suitable for computer interfacing and automation. Insulating the pn junctions from the fluid is the key challenge, which is overcome by using photo-definable SU8-2000 polymer.

Identification and characterization of smallest pore-forming protein in the cell wall of pathogenic Corynebacterium urealyticum DSM 7109.

Science.gov (United States)

Abdali, Narges; Younas, Farhan; Mafakheri, Samaneh; Pothula, Karunakar R; Kleinekathöfer, Ulrich; Tauch, Andreas; Benz, Roland

2018-05-09

Corynebacterium urealyticum, a pathogenic, multidrug resistant member of the mycolata, is known as causative agent of urinary tract infections although it is a bacterium of the skin flora. This pathogenic bacterium shares with the mycolata the property of having an unusual cell envelope composition and architecture, typical for the genus Corynebacterium. The cell wall of members of the mycolata contains channel-forming proteins for the uptake of solutes. In this study, we provide novel information on the identification and characterization of a pore-forming protein in the cell wall of C. urealyticum DSM 7109. Detergent extracts of whole C. urealyticum cultures formed in lipid bilayer membranes slightly cation-selective pores with a single-channel conductance of 1.75 nS in 1 M KCl. Experiments with different salts and non-electrolytes suggested that the cell wall pore of C. urealyticum is wide and water-filled and has a diameter of about 1.8 nm. Molecular modelling and dynamics has been performed to obtain a model of the pore. For the search of the gene coding for the cell wall pore of C. urealyticum we looked in the known genome of C. urealyticum for a similar chromosomal localization of the porin gene to known porH and porA genes of other Corynebacterium strains. Three genes are located between the genes coding for GroEL2 and polyphosphate kinase (PKK2). Two of the genes (cur_1714 and cur_1715) were expressed in different constructs in C. glutamicum ΔporAΔporH and in porin-deficient BL21 DE3 Omp8 E. coli strains. The results suggested that the gene cur_1714 codes alone for the cell wall channel. The cell wall porin of C. urealyticum termed PorACur was purified to homogeneity using different biochemical methods and had an apparent molecular mass of about 4 kDa on tricine-containing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Biophysical characterization of the purified protein (PorACur) suggested indeed that cur_1714 is the gene

Integration of 128 channels for monitoring, acquisition and control with existing LHCD DAC system

International Nuclear Information System (INIS)

Joshi, Ramesh; Virani, Chetan; Wadhwani, Archana; Sharma, P.K.

2015-01-01

Lower Hybrid Current Drive (LHCD) data acquisition system needs to be upgraded for additional channel requirement. The existing VME based DAC has been used since long with 32 analog input channels for data monitoring and control. Additional 128 channels require integrating with existing DAC. There are four layers of waveguides which deliver final output power into tokamak. Each layer requires 32 channels for power measurement. For the same requirement 128 analog input channels have been integrated with the help of carrier board and IP modules. Acromag IP330 modules have been procured and finally integrated with additional carrier board with existing VME hardware. Each module provides 32 analog input channels. Device driver has been developed for each module and integrated with existing program. LHCD DAC system has been upgraded with additional 128 channels requirement. It has been successfully testing with recent SST-1 campaign. (author)

Cardiac sodium channel NaV1.5 distribution in myocytes via interacting proteins: the multiple pool model.

Science.gov (United States)

Shy, Diana; Gillet, Ludovic; Abriel, Hugues

2013-04-01

The cardiac sodium current (INa) is responsible for the rapid depolarization of cardiac cells, thus allowing for their contraction. It is also involved in regulating the duration of the cardiac action potential (AP) and propagation of the impulse throughout the myocardium. Cardiac INa is generated by the voltage-gated Na(+) channel, NaV1.5, a 2016-residue protein which forms the pore of the channel. Over the past years, hundreds of mutations in SCN5A, the human gene coding for NaV1.5, have been linked to many cardiac electrical disorders, including the congenital and acquired long QT syndrome, Brugada syndrome, conduction slowing, sick sinus syndrome, atrial fibrillation, and dilated cardiomyopathy. Similar to many membrane proteins, NaV1.5 has been found to be regulated by several interacting proteins. In some cases, these different proteins, which reside in distinct membrane compartments (i.e. lateral membrane vs. intercalated disks), have been shown to interact with the same regulatory domain of NaV1.5, thus suggesting that several pools of NaV1.5 channels may co-exist in cardiac cells. The aim of this review article is to summarize the recent works that demonstrate its interaction with regulatory proteins and illustrate the model that the sodium channel NaV1.5 resides in distinct and different pools in cardiac cells. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Cardiac Pathways of Differentiation, Metabolism and Contraction. Copyright © 2012 Elsevier B.V. All rights reserved.

Intrinsically disordered proteins aggregate at fungal cell-to-cell channels and regulate intercellular connectivity.

Science.gov (United States)

Lai, Julian; Koh, Chuan Hock; Tjota, Monika; Pieuchot, Laurent; Raman, Vignesh; Chandrababu, Karthik Balakrishna; Yang, Daiwen; Wong, Limsoon; Jedd, Gregory

2012-09-25

Like animals and plants, multicellular fungi possess cell-to-cell channels (septal pores) that allow intercellular communication and transport. Here, using a combination of MS of Woronin body-associated proteins and a bioinformatics approach that identifies related proteins based on composition and character, we identify 17 septal pore-associated (SPA) proteins that localize to the septal pore in rings and pore-centered foci. SPA proteins are not homologous at the primary sequence level but share overall physical properties with intrinsically disordered proteins. Some SPA proteins form aggregates at the septal pore, and in vitro assembly assays suggest aggregation through a nonamyloidal mechanism involving mainly α-helical and disordered structures. SPA loss-of-function phenotypes include excessive septation, septal pore degeneration, and uncontrolled Woronin body activation. Together, our data identify the septal pore as a complex subcellular compartment and focal point for the assembly of unstructured proteins controlling diverse aspects of intercellular connectivity.

Two forms of Vibrio cholerae O1 El Tor hemolysin derived from identical precursor protein.

Science.gov (United States)

Ikigai, H; Ono, T; Nakae, T; Otsuru, H; Shimamura, T

1999-01-08

Vibrio cholerae O1 grown in heart infusion broth produces two forms of El Tor hemolysin (ETH) monomers of 65 and 50 kDa. These monomers form several different sizes of mixed oligomers ranging from 180 to 280 kDa in the liposomal membranes. We found that the N-terminal amino acid sequences, NH2-Trp-Pro-Ala-Pro-Ala-Asn-Ser-Glu, of both the 65- and 50-kDa toxins were identical. We assumed, therefore, that the 65- and 50-kDa toxins were derivatives of the identical precursor protein and the 50-kDa protein was a truncated derivative of 65-kDa ETH. To substantiate this assumption, we treated the 260-kDa oligomer with trypsin and obtained a 190-kDa oligomer. This 190-kDa oligomer consisted of only the 50-kDa subunits. Both 260- and 190-kDa oligomers formed ion channels indistinguishable from each other in planar lipid bilayers. These results suggest that the essential part of the ETH in forming the membrane-damaging aggregate is a 50-kDa protein.

Laboratory Modeling of Self-Formed Leveed Channels From Sediment-Laden Flows Entering Still Water

Science.gov (United States)

Rowland, J. C.; Dietrich, W. E.

2004-12-01

Self-formed leveed channels constructed by deposition of suspended sediment from sediment-laden flows entering still water are common features in nature. Such channels drive delta progradation, develop at tidal inlets and occur where mainstem river flows empty into oxbows and blocked valley lakes. Presently there is no theory for the formation of such channels. This lack of theory is partly due to a lack of field or laboratory studies that provide insight about the mechanism controlling these self-formed, propagating channels. The creation of such features in the laboratory, have proved illusive to date. Our ongoing experiments aimed at modeling the formation of floodplain tie channels provide insight into the necessary conditions for levee formation and channel growth. Under conditions of steady water discharge, constant sediment feed rate, unimodal sediment distribution and invariant basin stage we are able to create subaqueous lateral bars (submerged levees) along the margins of a sediment laden jet. Our results highlight the sensitivity of channel formation to issues of scaling and experimental design. In the laboratory, levee formation has only been possible with the use of plastic particles (specific gravity ~1.5); complete bed alluviation and dune formation results from the use of particles with specific gravities of ~ 2.65 across a range grain diameters and shapes. We hypothesize this effect is related to high entrainment thresholds relative to suspension thresholds of small (< 100 mm) natural particles under conditions of reduced turbulence in laboratory scaled flows. Additionally, both the width to depth ratio and the form of the outlet channel introducing the sediment laden flow into the experimental basin exert a strong control on sedimentation pattern and levee growth. Continuing experiments are focused on generating emergent channel levees and a basin ward propagation of the channel by adjusting the form of the feed channel, varying basin stage, and

Dysfunctional HCN ion channels in neurological diseases

Directory of Open Access Journals (Sweden)

Jacopo C. DiFrancesco

2015-03-01

Full Text Available Hyperpolarization-activated cyclic nucleotide-gated (HCN channels are expressed as four different isoforms (HCN1-4 in the heart and in the central and peripheral nervous systems. HCN channels are activated by membrane hyperpolarization at voltages close to resting membrane potentials and carry the hyperpolarization-activated current, dubbed If (funny current in heart and Ih in neurons. HCN channels contribute in several ways to neuronal activity and are responsible for many important cellular functions, including cellular excitability, generation and modulation of rhythmic activity, dendritic integration, transmission of synaptic potentials and plasticity phenomena. Because of their role, defective HCN channels are natural candidates in the search for potential causes of neurological disorders in humans. Several data, including growing evidence that some forms of epilepsy are associated with HCN mutations, support the notion of an involvement of dysfunctional HCN channels in different experimental models of the disease. Additionally, some anti-epileptic drugs are known to modify the activity of the Ih current. HCN channels are widely expressed in the peripheral nervous system and recent evidence has highlighted the importance of the HCN2 isoform in the transmission of pain. HCN channels are also present in the midbrain system, where they finely regulate the activity of dopaminergic neurons, and a potential role of these channels in the pathogenesis of Parkinson’s disease has recently emerged. The function of HCN channels is regulated by specific accessory proteins, which control the correct expression and modulation of the neuronal Ih current. Alteration of these proteins can severely interfere with the physiological channel function, potentially predisposing to pathological conditions. In this review we address the present knowledge of the association between HCN dysfunctions and neurological diseases, including clinical, genetic and

PIEZO channel protein naturally expressed in human breast cancer cell MDA-MB-231 as probed by atomic force microscopy

Science.gov (United States)

Weng, Yuanqi; Yan, Fei; Chen, Runkang; Qian, Ming; Ou, Yun; Xie, Shuhong; Zheng, Hairong; Li, Jiangyu

2018-05-01

Mechanical stimuli drives many physiological processes through mechanically activated channels, and the recent discovery of PIEZO channel has generated great interests in its mechanotransduction. Many previous researches investigated PIEZO proteins by transcribing them in cells that originally have no response to mechanical stimulation, or by forming PIEZO-combined complexes in vitro, and few studied PIEZO protein's natural characteristics in cells. In this study we show that MDA-MB-231, a malignant cell in human breast cancer cell line, expresses the mechanosensitive behavior of PIEZO in nature without extra treatment, and we report its characteristics in response to localized mechanical stimulation under an atomic force microscope, wherein a correlation between the force magnitude applied and the channel opening probability is observed. The results on PIEZO of MDA-MB-231 can help establish a basis of preventing and controlling of human breast cancer cell via mechanical forces.

Experimental Study of Axially Tension Cold Formed Steel Channel Members

Science.gov (United States)

Apriani, Widya; Lubis, Fadrizal; Angraini, Muthia

2017-12-01

Experimental testing is commonly used as one of the steps to determine the cause of the collapse of a building structure. The collapse of structures can be due to low quality materials. Although material samples have passed laboratory tests and the existing technical specifications have been met but there may be undetected defects and known material after failure. In this paper will be presented Experimental Testing of Axially Tension Cold Formed Steel Channel Members to determine the cause of the collapse of a building roof truss x in Pekanbaru. Test of tensile strength material cold formed channel sections was performed to obtain the main characteristics of Cold Formed steel material, namely ultimate tensile strength loads that can be held by members and the yield stress possessed by channel sections used in construction. Analysis of axially tension cold formed steel channel section presents in this paper was conducted through experimental study based on specificationsAnnualBook of ASTM Standards: Metal Test methods and Analitical Procedures, Section 3 (1991). The result of capacity loads experimental test was compared with design based on SNI 03-7971-2013standard of Indonesia for the design of cold formed steel structural members. The results of the yield stress of the material will be seen against the minimum allowable allowable stress range. After the test, the percentace of ultimate axial tension capacity theory has a result that is 16.46% larger than the ultimate axial tension capacity experimental. When compared with the load that must be borne 5.673 kN/m it can be concluded that 2 specimens do not meet. Yield stress of member has fulfilled requirement that wass bigger than 550 MPa. Based on the curve obtained ultimate axial tension capacity theory, results greater than experimental. The greatest voltage value (fu) is achieved under the same conditions as its yield stress. For this specimen with a melting voltage value fy = 571.5068 MPa has fulfilled the

Channel crossing: how are proteins shipped across the bacterial plasma membrane?

Science.gov (United States)

Collinson, Ian; Corey, Robin A; Allen, William J

2015-10-05

The structure of the first protein-conducting channel was determined more than a decade ago. Today, we are still puzzled by the outstanding problem of protein translocation--the dynamic mechanism underlying the consignment of proteins across and into membranes. This review is an attempt to summarize and understand the energy transducing capabilities of protein-translocating machines, with emphasis on bacterial systems: how polypeptides make headway against the lipid bilayer and how the process is coupled to the free energy associated with ATP hydrolysis and the transmembrane protein motive force. In order to explore how cargo is driven across the membrane, the known structures of the protein-translocation machines are set out against the background of the historic literature, and in the light of experiments conducted in their wake. The paper will focus on the bacterial general secretory (Sec) pathway (SecY-complex), and its eukaryotic counterpart (Sec61-complex), which ferry proteins across the membrane in an unfolded state, as well as the unrelated Tat system that assembles bespoke channels for the export of folded proteins. © 2015 The Authors.

Micro- and nanofabrication methods for ion channel reconstitution in bilayer lipid membranes

Science.gov (United States)

Tadaki, Daisuke; Yamaura, Daichi; Arata, Kohei; Ohori, Takeshi; Ma, Teng; Yamamoto, Hideaki; Niwano, Michio; Hirano-Iwata, Ayumi

2018-03-01

The self-assembled bilayer lipid membrane (BLM) forms the basic structure of the cell membrane and serves as a major barrier against ion movement. Ion channel proteins function as gated pores that permit ion permeation across the BLM. The reconstitution of ion channel proteins in artificially formed BLMs represents a well-defined system for investigating channel functions and screening drug effects on ion channels. In this review, we will discuss our recent microfabrication approaches to the formation of stable BLMs containing ion channel proteins as a potential platform for next-generation drug screening systems. BLMs formed in a microaperture having a tapered edge exhibited highly stable properties, such as a lifetime of ∼65 h and tolerance to solution changes even after the incorporation of the human ether-a-go-go-related gene (hERG) channel. We also explore a new method of efficiently incorporating human ion channels into BLMs by centrifugation. Our approaches to the formation of stable BLMs and efficient channel incorporation markedly improve the experimental efficiency of BLM reconstitution systems, leading to the realization of a BLM-based high-throughput platform for functional assays of various ion channels.

Fragile X mental retardation protein controls synaptic vesicle exocytosis by modulating N-type calcium channel density

Science.gov (United States)

Ferron, Laurent; Nieto-Rostro, Manuela; Cassidy, John S.; Dolphin, Annette C.

2014-04-01

Fragile X syndrome (FXS), the most common heritable form of mental retardation, is characterized by synaptic dysfunction. Synaptic transmission depends critically on presynaptic calcium entry via voltage-gated calcium (CaV) channels. Here we show that the functional expression of neuronal N-type CaV channels (CaV2.2) is regulated by fragile X mental retardation protein (FMRP). We find that FMRP knockdown in dorsal root ganglion neurons increases CaV channel density in somata and in presynaptic terminals. We then show that FMRP controls CaV2.2 surface expression by targeting the channels to the proteasome for degradation. The interaction between FMRP and CaV2.2 occurs between the carboxy-terminal domain of FMRP and domains of CaV2.2 known to interact with the neurotransmitter release machinery. Finally, we show that FMRP controls synaptic exocytosis via CaV2.2 channels. Our data indicate that FMRP is a potent regulator of presynaptic activity, and its loss is likely to contribute to synaptic dysfunction in FXS.

The human TRPV6 channel protein is associated with cyclophilin B in human placenta.

Science.gov (United States)

Stumpf, Tobias; Zhang, Qi; Hirnet, Daniela; Lewandrowski, Urs; Sickmann, Albert; Wissenbach, Ulrich; Dörr, Janka; Lohr, Christian; Deitmer, Joachim W; Fecher-Trost, Claudia

2008-06-27

Transcellular calcium transport in the kidney, pancreas, small intestine, and placenta is partly mediated by transient receptor potential (TRP) channels. The highly selective TRPV6 calcium channel protein is most likely important for the calcium transfer in different specialized epithelial cells. In the human placenta the protein is expressed in trophoblast tissue, where it is implicated in the transepithelial calcium transfer from mother to the fetus. We enriched the TRPV6 channel protein endogenously expressed in placenta together with annexin A2 and cyclophilin B (CypB), which is a member of the huge immunophilin family. In the human placenta TRPV6 and CypB are mainly located intracellularly in the syncytiotrophoblast layer, but a small amount of the mature glycosylated TRPV6 channel protein and CypB is also expressed in microvilli apical membranes, the fetomaternal barrier. To understand the role of CypB on the TRPV6 channel function, we evaluated the effect of CypB co-expression on TRPV6-mediated calcium uptake into Xenopus laevis oocytes expressing TRPV6. A significant increase of TRPV6-mediated calcium uptake was observed after CypB/TRPV6 co-expression. This stimulatory effect of CypB was reversed by the immunosuppressive drug cyclosporin A, which inhibits the enzymatic activity of CypB. Cyclosporin A had no significant effect on TRPV6 and CypB protein expression levels in the oocytes. In summary, our results establish CypB as a new TRPV6 accessory protein with potential involvement in TRPV6 channel activation through its peptidyl-prolyl cis/trans isomerase activity.

On the Role of the Annihilation Channel in Front Form Positronium

OpenAIRE

Trittmann, Uwe

1997-01-01

The annihilation channel is implemented into the front form calculations of the positronium spectrum presented in a previous publication. The effective Hamiltonian is calculated analytically. Its eigensolutions are obtained numerically. A complete separation of the dynamical and instantaneous part of the annihilation interaction is observed. We find the remarkable effect that the annihilation channel stabilizes the cutoff behavior of the spectrum.

RING finger protein 121 facilitates the degradation and membrane localization of voltage-gated sodium channels

Science.gov (United States)

Ogino, Kazutoyo; Low, Sean E.; Yamada, Kenta; Saint-Amant, Louis; Zhou, Weibin; Muto, Akira; Asakawa, Kazuhide; Nakai, Junichi; Kawakami, Koichi; Kuwada, John Y.; Hirata, Hiromi

2015-01-01

Following their synthesis in the endoplasmic reticulum (ER), voltage-gated sodium channels (NaV) are transported to the membranes of excitable cells, where they often cluster, such as at the axon initial segment of neurons. Although the mechanisms by which NaV channels form and maintain clusters have been extensively examined, the processes that govern their transport and degradation have received less attention. Our entry into the study of these processes began with the isolation of a new allele of the zebrafish mutant alligator, which we found to be caused by mutations in the gene encoding really interesting new gene (RING) finger protein 121 (RNF121), an E3-ubiquitin ligase present in the ER and cis-Golgi compartments. Here we demonstrate that RNF121 facilitates two opposing fates of NaV channels: (i) ubiquitin-mediated proteasome degradation and (ii) membrane localization when coexpressed with auxiliary NaVβ subunits. Collectively, these results indicate that RNF121 participates in the quality control of NaV channels during their synthesis and subsequent transport to the membrane. PMID:25691753

Mechanically Gated Ion Channels in Mammalian Hair Cells

Directory of Open Access Journals (Sweden)

Xufeng Qiu

2018-04-01

Full Text Available Hair cells in the inner ear convert mechanical stimuli provided by sound waves and head movements into electrical signal. Several mechanically evoked ionic currents with different properties have been recorded in hair cells. The search for the proteins that form the underlying ion channels is still in progress. The mechanoelectrical transduction (MET channel near the tips of stereociliary in hair cells, which is responsible for sensory transduction, has been studied most extensively. Several components of the sensory mechanotransduction machinery in stereocilia have been identified, including the multi-transmembrane proteins tetraspan membrane protein in hair cell stereocilia (TMHS/LHFPL5, transmembrane inner ear (TMIE and transmembrane channel-like proteins 1 and 2 (TMC1/2. However, there remains considerable uncertainty regarding the molecules that form the channel pore. In addition to the sensory MET channel, hair cells express the mechanically gated ion channel PIEZO2, which is localized near the base of stereocilia and not essential for sensory transduction. The function of PIEZO2 in hair cells is not entirely clear but it might have a role in damage sensing and repair processes. Additional stretch-activated channels of unknown molecular identity and function have been found to localize at the basolateral membrane of hair cells. Here, we review current knowledge regarding the different mechanically gated ion channels in hair cells and discuss open questions concerning their molecular composition and function.

Activation of purified calcium channels by stoichiometric protein phosphorylation

Energy Technology Data Exchange (ETDEWEB)

Nunoki, K.; Florio, V.; Catterall, W.A. (Univ. of Washington, Seattle (USA))

1989-09-01

Purified dihydropyridine-sensitive calcium channels from rabbit skeletal muscle were reconstituted into phosphatidylcholine vesicles to evaluate the effect of phosphorylation by cyclic AMP-dependent protein kinase (PK-A) on their function. Both the rate and extent of {sup 45}Ca{sup 2+} uptake into vesicles containing reconstituted calcium channels were increased severalfold after incubation with ATP and PK-A. The degree of stimulation of {sup 45}Ca{sup 2+} uptake was linearly proportional to the extent of phosphorylation of the alpha 1 and beta subunits of the calcium channel up to a stoichiometry of approximately 1 mol of phosphate incorporated into each subunit. The calcium channels activated by phosphorylation were determined to be incorporated into the reconstituted vesicles in the inside-out orientation and were completely inhibited by low concentrations of dihydropyridines, phenylalkylamines, Cd{sup 2+}, Ni{sup 2+}, and Mg{sup 2+}. The results demonstrate a direct relationship between PK-A-catalyzed phosphorylation of the alpha 1 and beta subunits of the purified calcium channel and activation of the ion conductance activity of the dihydropyridine-sensitive calcium channels.

Activation of purified calcium channels by stoichiometric protein phosphorylation

International Nuclear Information System (INIS)

Nunoki, K.; Florio, V.; Catterall, W.A.

1989-01-01

Purified dihydropyridine-sensitive calcium channels from rabbit skeletal muscle were reconstituted into phosphatidylcholine vesicles to evaluate the effect of phosphorylation by cyclic AMP-dependent protein kinase (PK-A) on their function. Both the rate and extent of 45 Ca 2+ uptake into vesicles containing reconstituted calcium channels were increased severalfold after incubation with ATP and PK-A. The degree of stimulation of 45 Ca 2+ uptake was linearly proportional to the extent of phosphorylation of the alpha 1 and beta subunits of the calcium channel up to a stoichiometry of approximately 1 mol of phosphate incorporated into each subunit. The calcium channels activated by phosphorylation were determined to be incorporated into the reconstituted vesicles in the inside-out orientation and were completely inhibited by low concentrations of dihydropyridines, phenylalkylamines, Cd 2+ , Ni 2+ , and Mg 2+ . The results demonstrate a direct relationship between PK-A-catalyzed phosphorylation of the alpha 1 and beta subunits of the purified calcium channel and activation of the ion conductance activity of the dihydropyridine-sensitive calcium channels

Regulation of Connexin-Based Channels by Fatty Acids

Science.gov (United States)

Puebla, Carlos; Retamal, Mauricio A.; Acuña, Rodrigo; Sáez, Juan C.

2017-01-01

In this mini-review, we briefly summarize the current knowledge about the effects of fatty acids (FAs) on connexin-based channels, as well as discuss the limited information about the impact FAs may have on pannexins (Panxs). FAs regulate diverse cellular functions, some of which are explained by changes in the activity of channels constituted by connexins (Cxs) or Panxs, which are known to play critical roles in maintaining the functional integrity of diverse organs and tissues. Cxs are transmembrane proteins that oligomerize into hexamers to form hemichannels (HCs), which in turn can assemble into dodecamers to form gap junction channels (GJCs). While GJCs communicate the cytoplasm of contacting cells, HCs serve as pathways for the exchange of ions and small molecules between the intra and extracellular milieu. Panxs, as well as Cx HCs, form channels at the plasma membrane that enable the interchange of molecules between the intra and extracellular spaces. Both Cx- and Panx-based channels are controlled by several post-translational modifications. However, the mechanism of action of FAs on these channels has not been described in detail. It has been shown however that FAs frequently decrease GJC-mediated cell-cell communication. The opposite effect also has been described for HC or Panx-dependent intercellular communication, where, the acute FA effect can be reversed upon washout. Additionally, changes in GJCs mediated by FAs have been associated with post-translational modifications (e.g., phosphorylation), and seem to be directly related to chemical properties of FAs (e.g., length of carbon chain and/or degree of saturation), but this possible link remains poorly understood. PMID:28174541

Gap junctions and connexin-interacting proteins

NARCIS (Netherlands)

Giepmans, Ben N G

2004-01-01

Gap junctions form channels between adjacent cells. The core proteins of these channels are the connexins. Regulation of gap junction communication (GJC) can be modulated by connexin-associating proteins, such as regulatory protein phosphatases and protein kinases, of which c-Src is the

Integration of G protein α (Gα) signaling by the regulator of G protein signaling 14 (RGS14).

Science.gov (United States)

Brown, Nicole E; Goswami, Devrishi; Branch, Mary Rose; Ramineni, Suneela; Ortlund, Eric A; Griffin, Patrick R; Hepler, John R

2015-04-03

RGS14 contains distinct binding sites for both active (GTP-bound) and inactive (GDP-bound) forms of Gα subunits. The N-terminal regulator of G protein signaling (RGS) domain binds active Gαi/o-GTP, whereas the C-terminal G protein regulatory (GPR) motif binds inactive Gαi1/3-GDP. The molecular basis for how RGS14 binds different activation states of Gα proteins to integrate G protein signaling is unknown. Here we explored the intramolecular communication between the GPR motif and the RGS domain upon G protein binding and examined whether RGS14 can functionally interact with two distinct forms of Gα subunits simultaneously. Using complementary cellular and biochemical approaches, we demonstrate that RGS14 forms a stable complex with inactive Gαi1-GDP at the plasma membrane and that free cytosolic RGS14 is recruited to the plasma membrane by activated Gαo-AlF4(-). Bioluminescence resonance energy transfer studies showed that RGS14 adopts different conformations in live cells when bound to Gα in different activation states. Hydrogen/deuterium exchange mass spectrometry revealed that RGS14 is a very dynamic protein that undergoes allosteric conformational changes when inactive Gαi1-GDP binds the GPR motif. Pure RGS14 forms a ternary complex with Gαo-AlF4(-) and an AlF4(-)-insensitive mutant (G42R) of Gαi1-GDP, as observed by size exclusion chromatography and differential hydrogen/deuterium exchange. Finally, a preformed RGS14·Gαi1-GDP complex exhibits full capacity to stimulate the GTPase activity of Gαo-GTP, demonstrating that RGS14 can functionally engage two distinct forms of Gα subunits simultaneously. Based on these findings, we propose a working model for how RGS14 integrates multiple G protein signals in host CA2 hippocampal neurons to modulate synaptic plasticity. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

VKCDB: Voltage-gated potassium channel database

Directory of Open Access Journals (Sweden)

Gallin Warren J

2004-01-01

Full Text Available Abstract Background The family of voltage-gated potassium channels comprises a functionally diverse group of membrane proteins. They help maintain and regulate the potassium ion-based component of the membrane potential and are thus central to many critical physiological processes. VKCDB (Voltage-gated potassium [K] Channel DataBase is a database of structural and functional data on these channels. It is designed as a resource for research on the molecular basis of voltage-gated potassium channel function. Description Voltage-gated potassium channel sequences were identified by using BLASTP to search GENBANK and SWISSPROT. Annotations for all voltage-gated potassium channels were selectively parsed and integrated into VKCDB. Electrophysiological and pharmacological data for the channels were collected from published journal articles. Transmembrane domain predictions by TMHMM and PHD are included for each VKCDB entry. Multiple sequence alignments of conserved domains of channels of the four Kv families and the KCNQ family are also included. Currently VKCDB contains 346 channel entries. It can be browsed and searched using a set of functionally relevant categories. Protein sequences can also be searched using a local BLAST engine. Conclusions VKCDB is a resource for comparative studies of voltage-gated potassium channels. The methods used to construct VKCDB are general; they can be used to create specialized databases for other protein families. VKCDB is accessible at http://vkcdb.biology.ualberta.ca.

Study on density wave oscillation in parallel channel by section form

International Nuclear Information System (INIS)

Huang Jun; Huang Yanping; Wang Yanlin

2013-01-01

Based on 170 density wave oscillation experimental data from parallel round tube and narrow rectangular channel, the experiment method, identification method of oscillation and analysis method of experimental data have be uniformed, and the oscillation boundary of round tube and narrow rectangular channel have be analyzed. The investigation results show that the oscillation boundary is not affected by the channel section forms with identical equivalent diameter with pressure l.0∼19.2 MPa, mass flux 101.9∼1200.0 kg·m-2·s -1 and inlet sub cooling 18.0∼85.2℃. (authors)

Protein translocation channel of mitochondrial inner membrane and matrix-exposed import motor communicate via two-domain coupling protein.

Science.gov (United States)

Banerjee, Rupa; Gladkova, Christina; Mapa, Koyeli; Witte, Gregor; Mokranjac, Dejana

2015-12-29

The majority of mitochondrial proteins are targeted to mitochondria by N-terminal presequences and use the TIM23 complex for their translocation across the mitochondrial inner membrane. During import, translocation through the channel in the inner membrane is coupled to the ATP-dependent action of an Hsp70-based import motor at the matrix face. How these two processes are coordinated remained unclear. We show here that the two domain structure of Tim44 plays a central role in this process. The N-terminal domain of Tim44 interacts with the components of the import motor, whereas its C-terminal domain interacts with the translocation channel and is in contact with translocating proteins. Our data suggest that the translocation channel and the import motor of the TIM23 complex communicate through rearrangements of the two domains of Tim44 that are stimulated by translocating proteins.

Calcium channel agonists and antagonists regulate protein phosphorylation in intact synaptosomes

International Nuclear Information System (INIS)

Robinson, P.J.; Lovenberg, Walter

1986-01-01

Protein phosphorylation in intact synaptosomes is highly sensitive to alterations in calcium fluxes and was used to probe the possible mechanism of action of the calcium channel agonist BAY K 8644 and antagonists verapamil and nifedipine. These agents (at 1μM) all increased the basal phosphorylation of a specific set of 4 synaptosomal phosphoproteins termed P139, P124, P96 and P60, but did not alter depolarization-dependent protein phosphorylation. The increases could not be explained by a direct stimulation of protein kinases and appears unrelated to the known effects of these + drugs on K + -stimulated neuro-transmitter release. This finding may reveal a possible new mechanism of action for drugs which interact with calcium channels. (Author)

Retroviral DNA Integration Directed by HIV Integration Protein in Vitro

Science.gov (United States)

Bushman, Frederic D.; Fujiwara, Tamio; Craigie, Robert

1990-09-01

Efficient retroviral growth requires integration of a DNA copy of the viral RNA genome into a chromosome of the host. As a first step in analyzing the mechanism of integration of human immunodeficiency virus (HIV) DNA, a cell-free system was established that models the integration reaction. The in vitro system depends on the HIV integration (IN) protein, which was partially purified from insect cells engineered to express IN protein in large quantities. Integration was detected in a biological assay that scores the insertion of a linear DNA containing HIV terminal sequences into a λ DNA target. Some integration products generated in this assay contained five-base pair duplications of the target DNA at the recombination junctions, a characteristic of HIV integration in vivo; the remaining products contained aberrant junctional sequences that may have been produced in a variation of the normal reaction. These results indicate that HIV IN protein is the only viral protein required to insert model HIV DNA sequences into a target DNA in vitro.

Ion channel electrophysiology via integrated planar patch-clamp chip with on-demand drug exchange.

Science.gov (United States)

Chen, Chang-Yu; Tu, Ting-Yuan; Jong, De-Shien; Wo, Andrew M

2011-06-01

Planar patch clamp has revolutionized characterization of ion channel behavior in drug discovery primarily via advancement in high throughput. Lab use of planar technology, however, addresses different requirements and suffers from inflexibility to enable wide range of interrogation via a single cell. This work presents integration of planar patch clamp with microfluidics, achieving multiple solution exchanges for tailor-specific measurement and allowing rapid replacement of the cell-contacting aperture. Studies via endogenously expressed ion channels in HEK 293T cells were commenced to characterize the device. Results reveal the microfluidic concentration generator produces distinct solution/drug combination/concentrations on-demand. Volume-regulated chloride channel and voltage-gated potassium channels in HEK 293T cells immersed in generated solutions under various osmolarities or drug concentrations show unique channel signature under specific condition. Excitation and blockage of ion channels in a single cell was demonstrated via serial solution exchange. Robustness of the reversible bonding and ease of glass substrate replacement were proven via repeated usage of the integrated device. The present approach reveals the capability and flexibility of integrated microfluidic planar patch-clamp system for ion channel assays. Copyright © 2011 Wiley Periodicals, Inc.

A Finite Element Solution of Lateral Periodic Poisson–Boltzmann Model for Membrane Channel Proteins

Science.gov (United States)

Xu, Jingjie; Lu, Benzhuo

2018-01-01

Membrane channel proteins control the diffusion of ions across biological membranes. They are closely related to the processes of various organizational mechanisms, such as: cardiac impulse, muscle contraction and hormone secretion. Introducing a membrane region into implicit solvation models extends the ability of the Poisson–Boltzmann (PB) equation to handle membrane proteins. The use of lateral periodic boundary conditions can properly simulate the discrete distribution of membrane proteins on the membrane plane and avoid boundary effects, which are caused by the finite box size in the traditional PB calculations. In this work, we: (1) develop a first finite element solver (FEPB) to solve the PB equation with a two-dimensional periodicity for membrane channel proteins, with different numerical treatments of the singular charges distributions in the channel protein; (2) add the membrane as a dielectric slab in the PB model, and use an improved mesh construction method to automatically identify the membrane channel/pore region even with a tilt angle relative to the z-axis; and (3) add a non-polar solvation energy term to complete the estimation of the total solvation energy of a membrane protein. A mesh resolution of about 0.25 Å (cubic grid space)/0.36 Å (tetrahedron edge length) is found to be most accurate in linear finite element calculation of the PB solvation energy. Computational studies are performed on a few exemplary molecules. The results indicate that all factors, the membrane thickness, the length of periodic box, membrane dielectric constant, pore region dielectric constant, and ionic strength, have individually considerable influence on the solvation energy of a channel protein. This demonstrates the necessity to treat all of those effects in the PB model for membrane protein simulations. PMID:29495644

Investigation of InP/InGaAs metamorphic co-integrated complementary doping-channel field-effect transistors for logic application

Science.gov (United States)

Tsai, Jung-Hui

2014-01-01

DC performance of InP/InGaAs metamorphic co-integrated complementary doping-channel field-effect transistors (DCFETs) grown on a low-cost GaAs substrate is first demonstrated. In the complementary DCFETs, the n-channel device was fabricated on the InxGa1-xP metamorphic linearly graded buffer layer and the p-channel field-effect transistor was stacked on the top of the n-channel device. Particularly, the saturation voltage of the n-channel device is substantially reduced to decrease the VOL and VIH values attributed that two-dimensional electron gas is formed and could be modulated in the n-InGaAs channel. Experimentally, a maximum extrinsic transconductance of 215 (17) mS/mm and a maximum saturation current density of 43 (-27) mA/mm are obtained in the n-channel (p-channel) device. Furthermore, the noise margins NMH and NML are up to 0.842 and 0.330 V at a supply voltage of 1.5 V in the complementary logic inverter application.

Some continual integrals from gaussian forms

International Nuclear Information System (INIS)

Mazmanishvili, A.S.

1985-01-01

The result summary of continual integration of gaussian functional type is given. The summary contains 124 continual integrals which are the mathematical expectation of the corresponding gaussian form by the continuum of random trajectories of four types: real-valued Ornstein-Uhlenbeck process, Wiener process, complex-valued Ornstein-Uhlenbeck process and the stochastic harmonic one. The summary includes both the known continual integrals and the unpublished before integrals. Mathematical results of the continual integration carried in the work may be applied in the problem of the theory of stochastic process, approaching to the finding of mean from gaussian forms by measures generated by the pointed stochastic processes

Imaging and structural studies of DNA–protein complexes and membrane ion channels

KAUST Repository

Marini, Monica; Limongi, Tania; Falqui, Andrea; Genovese, Alessandro; Allione, Marco; Moretti, Manola; Lopatin, Sergei; Tirinato, Luca; Das, Gobind; Torre, Bruno; Giugni, Andrea; Cesca, F.; Benfenati, F.; Di Fabrizio, Enzo M.

2017-01-01

In bio-imaging by electron microscopy, damage of the sample and limited contrast are the two main hurdles for reaching high image quality. We extend a new preparation method based on nanofabrication and super-hydrophobicity to the imaging and structural studies of nucleic acids, nucleic acid-protein complexes (DNA/Rad51 repair protein complex) and neuronal ion channels (gap-junction, K+ and GABA(A) channels) as paradigms of biological significance and increasing complexity. The preparation method is based on the liquid phase and is compatible with physiological conditions. Only in the very last stage, samples are dried for TEM analysis. Conventional TEM and high-resolution TEM (HRTEM) were used to achieve a resolution of 3.3 and 1.5 angstrom, respectively. The EM dataset quality allows the determination of relevant structural and metrological information on the DNA structure, DNA-protein interactions and ion channels, allowing the identification of specific macromolecules and their structure.

Imaging and structural studies of DNA–protein complexes and membrane ion channels

KAUST Repository

Marini, Monica

2017-01-17

In bio-imaging by electron microscopy, damage of the sample and limited contrast are the two main hurdles for reaching high image quality. We extend a new preparation method based on nanofabrication and super-hydrophobicity to the imaging and structural studies of nucleic acids, nucleic acid-protein complexes (DNA/Rad51 repair protein complex) and neuronal ion channels (gap-junction, K+ and GABA(A) channels) as paradigms of biological significance and increasing complexity. The preparation method is based on the liquid phase and is compatible with physiological conditions. Only in the very last stage, samples are dried for TEM analysis. Conventional TEM and high-resolution TEM (HRTEM) were used to achieve a resolution of 3.3 and 1.5 angstrom, respectively. The EM dataset quality allows the determination of relevant structural and metrological information on the DNA structure, DNA-protein interactions and ion channels, allowing the identification of specific macromolecules and their structure.

Integrated 1 GHz 4-channel InP phasar based WDM-receiver with Si bipolar frontend array

NARCIS (Netherlands)

Steenbergen, C.A.M.; Vreede, de L.C.N.; Dam, van C.; Scholtes, T.L.M.; Smit, M.K.; Tauritz, J.L.; Pedersen, J.W.; Moerman, I.; Verbeek, B.H.; Baets, R.G.F.

1995-01-01

An integrated 4-channel WDM-receiver frontend with 1 GHz channel bandwidth is described. The receiver consists of an integrated wavelength demultiplexer with photodiodes in InP technology connected through bond wires with a 4 channel Si bipolar transimpedance amplifier mounted on an epoxy board. The

Enhanced biocompatibility of neural probes by integrating microstructures and delivering anti-inflammatory agents via microfluidic channels

Science.gov (United States)

Liu, Bin; Kim, Eric; Meggo, Anika; Gandhi, Sachin; Luo, Hao; Kallakuri, Srinivas; Xu, Yong; Zhang, Jinsheng

2017-04-01

Objective. Biocompatibility is a major issue for chronic neural implants, involving inflammatory and wound healing responses of neurons and glial cells. To enhance biocompatibility, we developed silicon-parylene hybrid neural probes with open architecture electrodes, microfluidic channels and a reservoir for drug delivery to suppress tissue responses. Approach. We chronically implanted our neural probes in the rat auditory cortex and investigated (1) whether open architecture electrode reduces inflammatory reaction by measuring glial responses; and (2) whether delivery of antibiotic minocycline reduces inflammatory and tissue reaction. Four weeks after implantation, immunostaining for glial fibrillary acid protein (astrocyte marker) and ionizing calcium-binding adaptor molecule 1 (macrophages/microglia cell marker) were conducted to identify immunoreactive astrocyte and microglial cells, and to determine the extent of astrocytes and microglial cell reaction/activation. A comparison was made between using traditional solid-surface electrodes and newly-designed electrodes with open architecture, as well as between deliveries of minocycline and artificial cerebral-spinal fluid diffused through microfluidic channels. Main results. The new probes with integrated micro-structures induced minimal tissue reaction compared to traditional electrodes at 4 weeks after implantation. Microcycline delivered through integrated microfluidic channels reduced tissue response as indicated by decreased microglial reaction around the neural probes implanted. Significance. The new design will help enhance the long-term stability of the implantable devices.

Activation of protein kinase C alters the intracellular distribution and mobility of cardiac Na+ channels.

Science.gov (United States)

Hallaq, Haifa; Wang, Dao W; Kunic, Jennifer D; George, Alfred L; Wells, K Sam; Murray, Katherine T

2012-02-01

Na(+) current derived from expression of the cardiac isoform SCN5A is reduced by receptor-mediated or direct activation of protein kinase C (PKC). Previous work has suggested a possible role for loss of Na(+) channels at the plasma membrane in this effect, but the results are controversial. In this study, we tested the hypothesis that PKC activation acutely modulates the intracellular distribution of SCN5A channels and that this effect can be visualized in living cells. In human embryonic kidney cells that stably expressed SCN5A with green fluorescent protein (GFP) fused to the channel COOH-terminus (SCN5A-GFP), Na(+) currents were suppressed by an exposure to PKC activation. Using confocal microscopy, colocalization of SCN5A-GFP channels with the plasma membrane under control and stimulated conditions was quantified. A separate population of SCN5A channels containing an extracellular epitope was immunolabeled to permit temporally stable labeling of the plasma membrane. Our results demonstrated that Na(+) channels were preferentially trafficked away from the plasma membrane by PKC activation, with a major contribution by Ca(2+)-sensitive or conventional PKC isoforms, whereas stimulation of protein kinase A (PKA) had the opposite effect. Removal of the conserved PKC site Ser(1503) or exposure to the NADPH oxidase inhibitor apocynin eliminated the PKC-mediated effect to alter channel trafficking, indicating that both channel phosphorylation and ROS were required. Experiments using fluorescence recovery after photobleaching demonstrated that both PKC and PKA also modified channel mobility in a manner consistent with the dynamics of channel distribution. These results demonstrate that the activation of protein kinases can acutely regulate the intracellular distribution and molecular mobility of cardiac Na(+) channels in living cells.

Structural domains required for channel function of the mouse transient receptor potential protein homologue TRP1beta.

Science.gov (United States)

Engelke, Michael; Friedrich, Olaf; Budde, Petra; Schäfer, Christina; Niemann, Ursula; Zitt, Christof; Jüngling, Eberhard; Rocks, Oliver; Lückhoff, Andreas; Frey, Jürgen

2002-07-17

Transient receptor potential proteins (TRP) are supposed to participate in the formation of store-operated Ca(2+) influx channels by co-assembly. However, little is known which domains facilitate the interaction of subunits. Contribution of the N-terminal coiled-coil domain and ankyrin-like repeats and the putative pore region of the mouse TRP1beta (mTRP1beta) variant to the formation of functional cation channels were analyzed following overexpression in HEK293 (human embryonic kidney) cells. MTRP1beta expressing cells exhibited enhanced Ca(2+) influx and enhanced whole-cell membrane currents compared to mTRP1beta deletion mutants. Using a yeast two-hybrid assay only the coiled-coil domain facilitated homodimerization of the N-terminus. These results suggest that the N-terminus of mTRP1beta is required for structural organization thus forming functional channels.

G-protein mediates voltage regulation of agonist binding to muscarinic receptors: effects on receptor-Na+ channel interaction

International Nuclear Information System (INIS)

Cohen-Armon, M.; Garty, H.; Sokolovsky, M.

1988-01-01

The authors previous experiments in membranes prepared from rat heart and brain led them to suggest that the binding of agonist to the muscarinic receptors and to the Na + channels is a coupled event mediated by guanine nucleotide binding protein(s) [G-protein(s)]. These in vitro findings prompted us to employ synaptoneurosomes from brain stem tissue to examine (i) the binding properties of [ 3 H] acetylcholine at resting potential and under depolarization conditions in the absence and presence of pertussis toxin; (ii) the binding of [ 3 H]batrachotoxin to Na + channel(s) in the presence of the muscarinic agonists; and (iii) muscarinically induced 22 Na + uptake in the presence and absence of tetrodotoxin, which blocks Na + channels. The findings indicate that agonist binding to muscarinic receptors is voltage dependent, that this process is mediated by G-protein(s), and that muscarinic agonists induce opening of Na + channels. The latter process persists even after pertussis toxin treatment, indicating that it is not likely to be mediated by pertussis toxin sensitive G-protein(s). The system with its three interacting components-receptor, G-protein, and Na + channel-is such that at resting potential the muscarinic receptor induces opening of Na + channels; this property may provide a possible physiological mechanism for the depolarization stimulus necessary for autoexcitation or repetitive firing in heart or brain tissues

Multi-channel integrated circuits for the detection and measurement of ionizing radiation

International Nuclear Information System (INIS)

Engel, G.L.; Duggireddi, N.; Vangapally, V.; Elson, J.M.; Sobotka, L.G.; Charity, R.J.

2011-01-01

The Integrated Circuits (IC) Design Research Laboratory at Southern Illinois University Edwardsville (SIUE) has collaborated with the Nuclear Reactions Group at Washington University (WU) to develop a family of multi-channel integrated circuits. To date, the collaboration has successfully produced two micro-chips. The first was an analog shaped and peak sensing chip with on-board constant-fraction discriminators and sparsified readout. This chip is known as Heavy-Ion Nuclear Physics-16 Channel (HINP16C). The second chip, christened PSD8C, was designed to logically complement (in terms of detector types) the HINP16C chip. Pulse Shape Discrimination-8 Channel (PSD8C), featuring three settable charge integration windows per channel, performs pulse shape discrimination (PSD). This paper summarizes the design, capabilities, and features of the HINP16C and PSD8C ICs. It proceeds to discuss the modifications, made to the ICs and their associated systems, which have attempted to improve ease of use, increase performance, and extend capabilities. The paper concludes with a brief discussion of what may be the next chip (employing a multi-sampling scheme) to be added to our CMOS ASIC 'tool box' for radiation detection instrumentation.

KCNQ1 channel modulation by KCNE proteins via the voltage-sensing domain.

Science.gov (United States)

Nakajo, Koichi; Kubo, Yoshihiro

2015-06-15

The gating of the KCNQ1 potassium channel is drastically regulated by auxiliary subunit KCNE proteins. KCNE1, for example, slows the activation kinetics of KCNQ1 by two orders of magnitude. Like other voltage-gated ion channels, the opening of KCNQ1 is regulated by the voltage-sensing domain (VSD; S1-S4 segments). Although it has been known that KCNE proteins interact with KCNQ1 via the pore domain, some recent reports suggest that the VSD movement may be altered by KCNE. The altered VSD movement of KCNQ1 by KCNE proteins has been examined by site-directed mutagenesis, the scanning cysteine accessibility method (SCAM), voltage clamp fluorometry (VCF) and gating charge measurements. These accumulated data support the idea that KCNE proteins interact with the VSDs of KCNQ1 and modulate the gating of the KCNQ1 channel. In this review, we will summarize recent findings and current views of the KCNQ1 modulation by KCNE via the VSD. In this context, we discuss our recent findings that KCNE1 may alter physical interactions between the S4 segment (VSD) and the S5 segment (pore domain) of KCNQ1. Based on these findings from ourselves and others, we propose a hypothetical mechanism for how KCNE1 binding alters the VSD movement and the gating of the channel. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Control of Excitation/Inhibition Balance in a Hippocampal Circuit by Calcium Sensor Protein Regulation of Presynaptic Calcium Channels.

Science.gov (United States)

Nanou, Evanthia; Lee, Amy; Catterall, William A

2018-05-02

Activity-dependent regulation controls the balance of synaptic excitation to inhibition in neural circuits, and disruption of this regulation impairs learning and memory and causes many neurological disorders. The molecular mechanisms underlying short-term synaptic plasticity are incompletely understood, and their role in inhibitory synapses remains uncertain. Here we show that regulation of voltage-gated calcium (Ca 2+ ) channel type 2.1 (Ca V 2.1) by neuronal Ca 2+ sensor (CaS) proteins controls synaptic plasticity and excitation/inhibition balance in a hippocampal circuit. Prevention of CaS protein regulation by introducing the IM-AA mutation in Ca V 2.1 channels in male and female mice impairs short-term synaptic facilitation at excitatory synapses of CA3 pyramidal neurons onto parvalbumin (PV)-expressing basket cells. In sharp contrast, the IM-AA mutation abolishes rapid synaptic depression in the inhibitory synapses of PV basket cells onto CA1 pyramidal neurons. These results show that CaS protein regulation of facilitation and inactivation of Ca V 2.1 channels controls the direction of short-term plasticity at these two synapses. Deletion of the CaS protein CaBP1/caldendrin also blocks rapid depression at PV-CA1 synapses, implicating its upregulation of inactivation of Ca V 2.1 channels in control of short-term synaptic plasticity at this inhibitory synapse. Studies of local-circuit function revealed reduced inhibition of CA1 pyramidal neurons by the disynaptic pathway from CA3 pyramidal cells via PV basket cells and greatly increased excitation/inhibition ratio of the direct excitatory input versus indirect inhibitory input from CA3 pyramidal neurons to CA1 pyramidal neurons. This striking defect in local-circuit function may contribute to the dramatic impairment of spatial learning and memory in IM-AA mice. SIGNIFICANCE STATEMENT Many forms of short-term synaptic plasticity in neuronal circuits rely on regulation of presynaptic voltage-gated Ca 2+ (Ca V

Design and performance of a multi-channel, multi-sampling, PSD-enabling integrated circuit

Energy Technology Data Exchange (ETDEWEB)

Engel, G.L., E-mail: gengel@siue.ed [Department of Electrical and Computer Engineering, VLSI Design Research Laboratory, Southern Illinois University Edwardsville, Engineering Building, Room 3043 Edwardsville, IL 62026 1081 (United States); Hall, M.J.; Proctor, J.M. [Department of Electrical and Computer Engineering, VLSI Design Research Laboratory, Southern Illinois University Edwardsville, Engineering Building, Room 3043 Edwardsville, IL 62026 1081 (United States); Elson, J.M.; Sobotka, L.G.; Shane, R.; Charity, R.J. [Departments of Chemistry and Physics, Washington University, Saint Louis, MO 63130 (United States)

2009-12-21

This paper presents the design and test results of an eight-channel prototype integrated circuit chip intended to greatly simplify the pulse-processing electronics needed for large arrays of scintillation detectors. Because the chip design employs (user-controlled) multi-region charge integration, particle identification is incorporated into the basic design. Each channel on the chip also contains a time-to-voltage converter which provides relative time information. The pulse-height integrals and the relative time are all stored on capacitors and are either reset, after a user controlled time, or sequentially read out if acquisition of the event is desired. Each of the three pulse-height sub-channels consists of a gated integrator with eight programmable charging rates and an externally programmable gate generator that defines the start (with four time ranges) and width (with four time ranges) of the gate relative to an external discriminator signal. The chip supports three triggering modes, two time ranges, two power modes, and produces four sparsified analog pulse trains (three for the integrators and another for the time) with synchronized addresses for off-chip digitization with a pipelined ADC. The eight-channel prototype chip occupies an area of 2.8 mmx5.7 mm, dissipates 60 mW (low-power mode), and was fabricated in the AMI 0.5-mum process (C5N).

Design and performance of a multi-channel, multi-sampling, PSD-enabling integrated circuit

International Nuclear Information System (INIS)

Engel, G.L.; Hall, M.J.; Proctor, J.M.; Elson, J.M.; Sobotka, L.G.; Shane, R.; Charity, R.J.

2009-01-01

This paper presents the design and test results of an eight-channel prototype integrated circuit chip intended to greatly simplify the pulse-processing electronics needed for large arrays of scintillation detectors. Because the chip design employs (user-controlled) multi-region charge integration, particle identification is incorporated into the basic design. Each channel on the chip also contains a time-to-voltage converter which provides relative time information. The pulse-height integrals and the relative time are all stored on capacitors and are either reset, after a user controlled time, or sequentially read out if acquisition of the event is desired. Each of the three pulse-height sub-channels consists of a gated integrator with eight programmable charging rates and an externally programmable gate generator that defines the start (with four time ranges) and width (with four time ranges) of the gate relative to an external discriminator signal. The chip supports three triggering modes, two time ranges, two power modes, and produces four sparsified analog pulse trains (three for the integrators and another for the time) with synchronized addresses for off-chip digitization with a pipelined ADC. The eight-channel prototype chip occupies an area of 2.8 mmx5.7 mm, dissipates 60 mW (low-power mode), and was fabricated in the AMI 0.5-μm process (C5N).

Fluoride export (FEX) proteins from fungi, plants and animals are 'single barreled' channels containing one functional and one vestigial ion pore

Science.gov (United States)

Berbasova, Tetyana; Nallur, Sunitha; Sells, Taylor; Smith, Kathryn D.; Gordon, Patricia B.; Tausta, Susan Lori

2017-01-01

The fluoride export protein (FEX) in yeast and other fungi provides tolerance to fluoride (F-), an environmentally ubiquitous anion. FEX efficiently eliminates intracellular fluoride that otherwise would accumulate at toxic concentrations. The FEX homolog in bacteria, Fluc, is a ‘double-barreled’ channel formed by dimerization of two identical or similar subunits. FEX in yeast and other eukaryotes is a monomer resulting from covalent fusion of the two subunits. As a result, both potential fluoride pores are created from different parts of the same protein. Here we identify FEX proteins from two multicellular eukaryotes, a plant Arabidopsis thaliana and an animal Amphimedon queenslandica, by demonstrating significant fluoride tolerance when these proteins are heterologously expressed in the yeast Saccharomyces cerevisiae. Residues important for eukaryotic FEX function were determined by phylogenetic sequence alignment and functional analysis using a yeast growth assay. Key residues of the fluoride channel are conserved in only one of the two potential fluoride-transporting pores. FEX activity is abolished upon mutation of residues in this conserved pore, suggesting that only one of the pores is functional. The same topology is conserved for the newly identified FEX proteins from plant and animal. These data suggest that FEX family of fluoride channels in eukaryotes are ‘single-barreled’ transporters containing one functional pore and a second non-functional vestigial remnant of a homologous gene fusion event. PMID:28472134

Clear Channel Assessment in Integrated Medical Environments

Directory of Open Access Journals (Sweden)

Hara Shinsuke

2008-01-01

Full Text Available Abstract Complementary WLAN and WPAN technologies as well as other wireless technologies will play a fundamental role in the medical environments to support ubiquitous healthcare delivery. This paper investigates clear channel assessment (CCA and its impact on the coexistence of WLAN (IEEE 802.11 high rate direct sequence spread spectrum (HR/DSSS PHY and WPAN (IEEE 802.15.4b in the 2.4 GHz industrial, scientific, and medical (ISM band. We derived closed-form expressions of both energy-based CCA and feature-based CCA. We qualified unequal sensing abilities between them and termed this inequality asymmetric CCA, which is different from the traditional "hidden node" or "exposed node" issues in the homogeneous network. The energy-based CCA was considered in the considered integrated medical environment because the 2.4 GHz ISM band is too crowded to apply feature-based CCA. The WPAN is oversensitive to the 802.11 HR/DSSS signals and the WLAN is insensitive to the 802.15.4b signals. Choosing an optimal CCA threshold requires some prior knowledge of the underlying signals. In the integrated medical environment we considered here, energy-based CCA can effectively avoid possible packet collisions when they are close within the "heterogeneous exclusive CCA range" (HECR. However, when they are separated beyond the HECR, WPAN can still sense the 802.11 HR/DSSS signals, but WLAN loses its sense to the 802.15.4b signals. The asymmetric CCA leads to WPAN traffic in a position secondary to WLAN traffic.

Integrating channel form and processes in the Gangetic plains rivers: Implications for geomorphic diversity

Science.gov (United States)

Roy, N. G.; Sinha, R.

2018-02-01

Geomorphic diversity at a variety of spatial and temporal scales has been studied in the western Ganga plains (WGP), India, to isolate the dominating factors at each scale that have the potential to cause major geomorphic change. The Ganga River and its major tributaries draining the WGP have been investigated in terms of longitudinal, cross-sectional, and planform morphology to assess the influence of potential controls such as climate, geology, topography, land use, hydrology, and sediment transport. These data were then compared with those from the rivers draining the eastern Ganga plains (EGP) to understand the geomorphic diversity across the Ganga plains and the causal factors. Our investigations suggest that in-channel geomorphic diversity over decadal scale in rivers with low width-to-depth (W/D) ratio is caused by periodic incision/aggradation, but it is driven by channel avulsion in rivers characterized by high W/D ratio. Similarly, planform (reach-scale) parameters such as sinuosity and braid-channel-ratio are influenced by intrinsic factors such as changes in hydrological conditions and morphodynamics (cutoffs, small-scale avulsion) that are in turn impacted by natural and human-induced factors. Finally, we have isolated the climatic and hydrologic effects on the longitudinal profile concavity of alluvial trunk channels in tectonically stable and unstable landscapes. We demonstrate that the rivers flowing through a tectonically stable landscape are graded in nature where higher discharge tends to create more concave longitudinal profiles compared to those in tectonically unstable landscape at 103-year scale.

Protein structure and ionic selectivity in calcium channels: selectivity filter size, not shape, matters.

Science.gov (United States)

Malasics, Attila; Gillespie, Dirk; Nonner, Wolfgang; Henderson, Douglas; Eisenberg, Bob; Boda, Dezso

2009-12-01

Calcium channels have highly charged selectivity filters (4 COO(-) groups) that attract cations in to balance this charge and minimize free energy, forcing the cations (Na(+) and Ca(2+)) to compete for space in the filter. A reduced model was developed to better understand the mechanism of ion selectivity in calcium channels. The charge/space competition (CSC) mechanism implies that Ca(2+) is more efficient in balancing the charge of the filter because it provides twice the charge as Na(+) while occupying the same space. The CSC mechanism further implies that the main determinant of Ca(2+) versus Na(+) selectivity is the density of charged particles in the selectivity filter, i.e., the volume of the filter (after fixing the number of charged groups in the filter). In this paper we test this hypothesis by changing filter length and/or radius (shape) of the cylindrical selectivity filter of our reduced model. We show that varying volume and shape together has substantially stronger effects than varying shape alone with volume fixed. Our simulations show the importance of depletion zones of ions in determining channel conductance calculated with the integrated Nernst-Planck equation. We show that confining the protein side chains with soft or hard walls does not influence selectivity.

Mechanism underlying selective regulation of G protein-gated inwardly rectifying potassium channels by the psychostimulant-sensitive sorting nexin 27

Science.gov (United States)

Balana, Bartosz; Maslennikov, Innokentiy; Kwiatkowski, Witek; Stern, Kalyn M.; Bahima, Laia; Choe, Senyon; Slesinger, Paul A.

2011-01-01

G protein-gated inwardly rectifying potassium (GIRK) channels are important gatekeepers of neuronal excitability. The surface expression of neuronal GIRK channels is regulated by the psychostimulant-sensitive sorting nexin 27 (SNX27) protein through a class I (-X-Ser/Thr-X-Φ, where X is any residue and Φ is a hydrophobic amino acid) PDZ-binding interaction. The G protein-insensitive inward rectifier channel (IRK1) contains the same class I PDZ-binding motif but associates with a different synaptic PDZ protein, postsynaptic density protein 95 (PSD95). The mechanism by which SNX27 and PSD95 discriminate these channels was previously unclear. Using high-resolution structures coupled with biochemical and functional analyses, we identified key amino acids upstream of the channel's canonical PDZ-binding motif that associate electrostatically with a unique structural pocket in the SNX27-PDZ domain. Changing specific charged residues in the channel's carboxyl terminus or in the PDZ domain converts the selective association and functional regulation by SNX27. Elucidation of this unique interaction site between ion channels and PDZ-containing proteins could provide a therapeutic target for treating brain diseases. PMID:21422294

Carbon nanotubes integrated in electrically insulated channels for lab-on-a-chip applications

International Nuclear Information System (INIS)

Mogensen, K B; Boggild, P; Kutter, J P; Gangloff, L; Teo, K B K; Milne, W I

2009-01-01

A fabrication process for monolithic integration of vertically aligned carbon nanotubes in electrically insulated microfluidic channels is presented. A 150 nm thick amorphous silicon layer could be used both for anodic bonding of a glass lid to hermetically seal the microfluidic glass channels and for de-charging of the wafer during plasma enhanced chemical vapor deposition of the carbon nanotubes. The possibility of operating the device with electroosmotic flow was shown by performing standard electrophoretic separations of 50 μM fluorescein and 50 μM 5-carboxyfluorescein in a 25 mm long column containing vertical aligned carbon nanotubes. This is the first demonstration of electroosmotic pumping and electrokinetic separations in microfluidic channels with a monolithically integrated carbon nanotube forest.

Carbon nanotubes integrated in electrically insulated channels for lab-on-a-chip applications

DEFF Research Database (Denmark)

Mogensen, Klaus Bo; Gangloff, L.; Bøggild, Peter

2009-01-01

A fabrication process for monolithic integration of vertically aligned carbon nanotubes in electrically insulated microfluidic channels is presented. A 150 nm thick amorphous silicon layer could be used both for anodic bonding of a glass lid to hermetically seal the microfluidic glass channels...... and for de-charging of the wafer during plasma enhanced chemical vapor deposition of the carbon nanotubes. The possibility of operating the device with electroosmotic flow was shown by performing standard electrophoretic separations of 50 mu M fluorescein and 50 mu M 5-carboxyfluorescein in a 25 mm long...... column containing vertical aligned carbon nanotubes. This is the first demonstration of electroosmotic pumping and electrokinetic separations in microfluidic channels with a monolithically integrated carbon nanotube forest....

Lipid-protein nanodiscs for cell-free production of integral membrane proteins in a soluble and folded state: comparison with detergent micelles, bicelles and liposomes.

Science.gov (United States)

Lyukmanova, E N; Shenkarev, Z O; Khabibullina, N F; Kopeina, G S; Shulepko, M A; Paramonov, A S; Mineev, K S; Tikhonov, R V; Shingarova, L N; Petrovskaya, L E; Dolgikh, D A; Arseniev, A S; Kirpichnikov, M P

2012-03-01

Production of integral membrane proteins (IMPs) in a folded state is a key prerequisite for their functional and structural studies. In cell-free (CF) expression systems membrane mimicking components could be added to the reaction mixture that promotes IMP production in a soluble form. Here lipid-protein nanodiscs (LPNs) of different lipid compositions (DMPC, DMPG, POPC, POPC/DOPG) have been compared with classical membrane mimicking media such as detergent micelles, lipid/detergent bicelles and liposomes by their ability to support CF synthesis of IMPs in a folded and soluble state. Three model membrane proteins of different topology were used: homodimeric transmembrane (TM) domain of human receptor tyrosine kinase ErbB3 (TM-ErbB3, 1TM); voltage-sensing domain of K(+) channel KvAP (VSD, 4TM); and bacteriorhodopsin from Exiguobacterium sibiricum (ESR, 7TM). Structural and/or functional properties of the synthesized proteins were analyzed. LPNs significantly enhanced synthesis of the IMPs in a soluble form regardless of the lipid composition. A partial disintegration of LPNs composed of unsaturated lipids was observed upon co-translational IMP incorporation. Contrary to detergents the nanodiscs resulted in the synthesis of ~80% active ESR and promoted correct folding of the TM-ErbB3. None of the tested membrane mimetics supported CF synthesis of correctly folded VSD, and the protocol of the domain refolding was developed. The use of LPNs appears to be the most promising approach to CF production of IMPs in a folded state. NMR analysis of (15)N-Ile-TM-ErbB3 co-translationally incorporated into LPNs shows the great prospects of this membrane mimetics for structural studies of IMPs produced by CF systems. Copyright © 2011 Elsevier B.V. All rights reserved.

Channel integration in governmental service delivery: the effects on citizen behavior and perceptions

NARCIS (Netherlands)

Pieterson, Willem Jan; Teerling, Marije; Wimmer, Maria A.; Scholl, Hans J.; Janssen, Marijn; Traunmüller, Roland

2009-01-01

Governmental agencies continuously work on the improvement of their service delivery through an array of channels. To improve service satisfaction and to reduce the cost of service delivery, channel integration gets more popular with governmental agencies. In a quasi experimental longitudinal field

The transient receptor potential, TRP4, cation channel is a novel member of the family of calmodulin binding proteins.

OpenAIRE

Trost, C; Bergs, C; Himmerkus, N; Flockerzi, V

2001-01-01

The mammalian gene products, transient receptor potential (trp)1 to trp7, are related to the Drosophila TRP and TRP-like ion channels, and are candidate proteins underlying agonist-activated Ca(2+)-permeable ion channels. Recently, the TRP4 protein has been shown to be part of native store-operated Ca(2+)-permeable channels. These channels, most likely, are composed of other proteins in addition to TRP4. In the present paper we report the direct interaction of TRP4 and calmodulin (CaM) by: (1...

Characterization of canine mitochondrial protein expression in natural and induced forms of idiopathic dilated cardiomyopathy.

Science.gov (United States)

Lopes, Rosana; Solter, Philip F; Sisson, D David; Oyama, Mark A; Prosek, Robert

2006-06-01

To map canine mitochondrial proteins and identify qualitative and quantitative differences in heart mitochondrial protein expression between healthy dogs and dogs with naturally occurring and induced dilated cardiomyopathy (DCM). Left ventricle samples were obtained from 7 healthy dogs, 7 Doberman Pinschers with naturally occurring DCM, and 7 dogs with induced DCM. Fresh and frozen mitochondrial fractions were isolated from the left ventricular free wall and analyzed by 2-dimensional electrophoresis. Protein spots that increased or decreased in density by >or= 2-fold between groups were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry or quadrupole selecting, quadrupole collision cell, time-of-flight mass spectrometry. Within narrow pH gradients of control canine heart mitochondrial samples, a total of 1,528 protein spots were revealed. Forty subunits of heart mitochondrial proteins that differ significantly from control tissues were altered in tissue specimens from dogs with naturally occurring and induced forms of DCM. The most affected heart mitochondrial proteins in both groups were those of oxidative phosphorylation (55%). Upregulation of manganese superoxide dismutase was suggestive of heart oxidative injury in tissue specimens from dogs with both forms of DCM. Evidence of apoptosis was associated with overexpression of the heart mitochondrial voltage-dependent anion channel-2 protein and endonuclease G in tissue specimens from dogs with induced DCM. Alterations of heart mitochondrial proteins related to oxidative phosphorylation dysfunction were more prevalent in tissue specimens from dogs with induced or naturally occurring DCM, compared with those of control dogs.

Fabrication of resonant micro cantilevers with integrated transparent fluidic channel

DEFF Research Database (Denmark)

Khan, Faheem; Schmid, Silvan; Davis, Zachary James

2011-01-01

Microfabricated cantilevers are proving their potential as excellent tools for analysis applications. In this paper, we describe the design, fabrication and testing of resonant micro cantilevers with integrated transparent fluidic channels. The cantilevers have been devised to measure the density...

Electrostatics promotes molecular crowding and selects the aggregation pathway in fibril-forming protein solutions

International Nuclear Information System (INIS)

Raccosta, S.; Martorana, V.; Manno, M.; Blanco, M.; Roberts, C.J.

2016-01-01

The role of intermolecular interaction in fibril-forming protein solutions and its relation with molecular conformation are crucial aspects for the control and inhibition of amyloid structures. Here, we study the fibril formation and the protein-protein interactions for two proteins at acidic ph, lysozyme and α-chymotrypsinogen. By using light scattering experiments and the Kirkwood-Buff integral approach, we show how concentration fluctuations are damped even at moderate protein concentrations by the dominant long-ranged electrostatic repulsion, which determines an effective crowded environment. In denaturing conditions, electrostatic repulsion keeps the monomeric solution in a thermodynamically metastable state, which is escaped through kinetically populated conformational sub-states. This explains how electrostatics acts as a gatekeeper in selecting a specific aggregation pathway.

Channel Capacity Calculation at Large SNR and Small Dispersion within Path-Integral Approach

Science.gov (United States)

Reznichenko, A. V.; Terekhov, I. S.

2018-04-01

We consider the optical fiber channel modelled by the nonlinear Shrödinger equation with additive white Gaussian noise. Using Feynman path-integral approach for the model with small dispersion we find the first nonzero corrections to the conditional probability density function and the channel capacity estimations at large signal-to-noise ratio. We demonstrate that the correction to the channel capacity in small dimensionless dispersion parameter is quadratic and positive therefore increasing the earlier calculated capacity for a nondispersive nonlinear optical fiber channel in the intermediate power region. Also for small dispersion case we find the analytical expressions for simple correlators of the output signals in our noisy channel.

TRPP2 and TRPV4 form an EGF-activated calcium permeable channel at the apical membrane of renal collecting duct cells.

Directory of Open Access Journals (Sweden)

Zhi-Ren Zhang

Full Text Available Regulation of apical calcium entry is important for the function of principal cells of the collecting duct. However, the molecular identity and the regulators of the transporter/channel, which is responsible for apical calcium entry and what factors regulate the calcium conduction remain unclear.We report that endogenous TRPP2 and TRPV4 assemble to form a 23-pS divalent cation-permeable non-selective ion channel at the apical membrane of renal principal cells of the collecting duct. TRPP2\\TRPV4 channel complex was identified by patch-clamp, immunofluorescence and co-immunprecipitation studies in both principal cells that either possess normal cilia (cilia (+ or in which cilia are absent (cilia (-. This channel has distinct biophysical and pharmacological and regulatory profiles compared to either TRPP2 or TRPV4 channels. The rate of occurrence detected by patch clamp was higher in cilia (- compared to cilia (+ cells. In addition, shRNA knockdown of TRPP2 increased the prevalence of TRPV4 channel activity while knockdown of TRPV4 resulted in TRPP2 activity and knockdown of both proteins vastly decreased the 23-pS channel activity. Epidermal growth factor (EGF stimulated TRPP2\\TRPV4 channel through the EGF receptor (EGFR tyrosine kinase-dependent signaling. With loss of cilia, apical EGF treatment resulted in 64-fold increase in channel activity in cilia (- but not cilia (+ cells. In addition EGF increased cell proliferation in cilia (- cell that was dependent upon TRPP2\\TRPV4 channel mediated increase in intracellular calcium.We conclude that in the absence of cilia, an EGF activated TRPP2\\TRPV4 channel may play an important role in increased cell proliferation and cystogenesis.

Characterization of auxin-binding proteins from zucchini plasma membrane

Science.gov (United States)

Hicks, G. R.; Rice, M. S.; Lomax, T. L.

1993-01-01

We have previously identified two auxin-binding polypeptides in plasma membrane (PM) preparations from zucchini (Cucurbita pepo L.) (Hicks et al. 1989, Proc. Natl. Acad. Sci. USA 86, 4948-4952). These polypeptides have molecular weights of 40 kDa and 42 kDa and label specifically with the photoaffinity auxin analog 5-N3-7-3H-IAA (azido-IAA). Azido-IAA permits both the covalent and radioactive tagging of auxin-binding proteins and has allowed us to characterize further the 40-kDa and 42-kDa polypeptides, including the nature of their attachment to the PM, their relationship to each other, and their potential function. The azido-IAA-labeled polypeptides remain in the pelleted membrane fraction following high-salt and detergent washes, which indicates a tight and possibly integral association with the PM. Two-dimensional electrophoresis of partially purified azido-IAA-labeled protein demonstrates that, in addition to the major isoforms of the 40-kDa and 42-kDa polypeptides, which possess isoelectric points (pIs) of 8.2 and 7.2, respectively, several less abundant isoforms that display unique pIs are apparent at both molecular masses. Tryptic and chymotryptic digestion of the auxin-binding proteins indicates that the 40-kDa and 42-kDa polypeptides are closely related or are modifications of the same polypeptide. Phase extraction with the nonionic detergent Triton X-114 results in partitioning of the azido-IAA-labeled polypeptides into the aqueous (hydrophilic) phase. This apparently paradoxical behavior is also exhibited by certain integral membrane proteins that aggregate to form channels. The results of gel filtration indicate that the auxin-binding proteins do indeed aggregate strongly and that the polypeptides associate to form a dimer or multimeric complex in vivo. These characteristics are consistent with the hypothesis that the 40-kDa and 42-kDa polypeptides are subunits of a multimeric integral membrane protein which has an auxin-binding site, and which may

Structures of pseudechetoxin and pseudecin, two snake-venom cysteine-rich secretory proteins that target cyclic nucleotide-gated ion channels: implications for movement of the C-terminal cysteine-rich domain

International Nuclear Information System (INIS)

Suzuki, Nobuhiro; Yamazaki, Yasuo; Brown, R. Lane; Fujimoto, Zui; Morita, Takashi; Mizuno, Hiroshi

2008-01-01

The structures of pseudechetoxin and pseudecin suggest that both proteins bind to cyclic nucleotide-gated ion channels in a manner in which the concave surface occludes the pore entrance. Cyclic nucleotide-gated (CNG) ion channels play pivotal roles in sensory transduction by retinal photoreceptors and olfactory neurons. The elapid snake toxins pseudechetoxin (PsTx) and pseudecin (Pdc) are the only known protein blockers of CNG channels. These toxins belong to a cysteine-rich secretory protein (CRISP) family containing an N-terminal pathogenesis-related proteins of group 1 (PR-1) domain and a C-terminal cysteine-rich domain (CRD). PsTx and Pdc are highly homologous proteins, but their blocking affinities on CNG channels are different: PsTx blocks both the olfactory and retinal channels with ∼15–30-fold higher affinity than Pdc. To gain further insights into their structure and function, the crystal structures of PsTx, Pdc and Zn 2+ -bound Pdc were determined. The structures revealed that most of the amino-acid-residue differences between PsTx and Pdc are located around the concave surface formed between the PR-1 domain and the CRD, suggesting that the concave surface is functionally important for CNG-channel binding and inhibition. A structural comparison in the presence and absence of Zn 2+ ion demonstrated that the concave surface can open and close owing to movement of the CRD upon Zn 2+ binding. The data suggest that PsTx and Pdc occlude the pore entrance and that the dynamic motion of the concave surface facilitates interaction with the CNG channels

Structures of pseudechetoxin and pseudecin, two snake-venom cysteine-rich secretory proteins that target cyclic nucleotide-gated ion channels: implications for movement of the C-terminal cysteine-rich domain

Energy Technology Data Exchange (ETDEWEB)

Suzuki, Nobuhiro [Department of Applied Biochemistry, University of Tsukuba, Tsukuba, Ibaraki 305-8572 (Japan); Department of Biochemistry, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8602 (Japan); Yamazaki, Yasuo [Department of Biochemistry, Meiji Pharmaceutical University, Kiyose, Tokyo 204-8588 (Japan); Brown, R. Lane [Neurological Science Institute, Oregon Health and Science University, Beaverton, Oregon 97006 (United States); Fujimoto, Zui [Department of Biochemistry, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8602 (Japan); Morita, Takashi, E-mail: tmorita@my-pharm.ac.jp [Department of Biochemistry, Meiji Pharmaceutical University, Kiyose, Tokyo 204-8588 (Japan); Mizuno, Hiroshi, E-mail: tmorita@my-pharm.ac.jp [Department of Biochemistry, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki 305-8602 (Japan); VALWAY Technology Center, NEC Soft Ltd, Koto-ku, Tokyo 136-8627 (Japan); Institute for Biological Resources and Functions, National Institute of Advanced Industrial Science and Technology, Central 6, Tsukuba, Ibaraki 305-8566 (Japan); Department of Applied Biochemistry, University of Tsukuba, Tsukuba, Ibaraki 305-8572 (Japan)

2008-10-01

The structures of pseudechetoxin and pseudecin suggest that both proteins bind to cyclic nucleotide-gated ion channels in a manner in which the concave surface occludes the pore entrance. Cyclic nucleotide-gated (CNG) ion channels play pivotal roles in sensory transduction by retinal photoreceptors and olfactory neurons. The elapid snake toxins pseudechetoxin (PsTx) and pseudecin (Pdc) are the only known protein blockers of CNG channels. These toxins belong to a cysteine-rich secretory protein (CRISP) family containing an N-terminal pathogenesis-related proteins of group 1 (PR-1) domain and a C-terminal cysteine-rich domain (CRD). PsTx and Pdc are highly homologous proteins, but their blocking affinities on CNG channels are different: PsTx blocks both the olfactory and retinal channels with ∼15–30-fold higher affinity than Pdc. To gain further insights into their structure and function, the crystal structures of PsTx, Pdc and Zn{sup 2+}-bound Pdc were determined. The structures revealed that most of the amino-acid-residue differences between PsTx and Pdc are located around the concave surface formed between the PR-1 domain and the CRD, suggesting that the concave surface is functionally important for CNG-channel binding and inhibition. A structural comparison in the presence and absence of Zn{sup 2+} ion demonstrated that the concave surface can open and close owing to movement of the CRD upon Zn{sup 2+} binding. The data suggest that PsTx and Pdc occlude the pore entrance and that the dynamic motion of the concave surface facilitates interaction with the CNG channels.

Monolithic integration of microfluidic channels and optical waveguides in silica on silicon

DEFF Research Database (Denmark)

Friis, Peter; Hoppe, Karsten; Leistiko, Otto

2001-01-01

-back technique are possible, but troublesome. We present a simple but efficient alternative: By means of changing the waveguide layout, bonding pads are formed along the microfluidic channels. With the same height as the waveguide, they effectively prevent leakage and hermetically seal the channels during...

Fatty acids and related Kv2 channel blockers: electrophysiology and toxicity on mosquitoes

Science.gov (United States)

Ligand-gated ion channels form an important superfamily of proteins involved in many biological processes. Among them, the potassium channels constitute a very diverse group involved in neural signaling, neuronal activity and action potential. Among the different types of channel activation, voltage...

Clear Channel Assessment in Integrated Medical Environments

Directory of Open Access Journals (Sweden)

Bin Zhen

2008-03-01

Full Text Available Complementary WLAN and WPAN technologies as well as other wireless technologies will play a fundamental role in the medical environments to support ubiquitous healthcare delivery. This paper investigates clear channel assessment (CCA and its impact on the coexistence of WLAN (IEEE 802.11 high rate direct sequence spread spectrum (HR/DSSS PHY and WPAN (IEEE 802.15.4b in the 2.4 GHz industrial, scientific, and medical (ISM band. We derived closed-form expressions of both energy-based CCA and feature-based CCA. We qualified unequal sensing abilities between them and termed this inequality asymmetric CCA, which is different from the traditional “hidden node” or “exposed node” issues in the homogeneous network. The energy-based CCA was considered in the considered integrated medical environment because the 2.4 GHz ISM band is too crowded to apply feature-based CCA. The WPAN is oversensitive to the 802.11 HR/DSSS signals and the WLAN is insensitive to the 802.15.4b signals. Choosing an optimal CCA threshold requires some prior knowledge of the underlying signals. In the integrated medical environment we considered here, energy-based CCA can effectively avoid possible packet collisions when they are close within the “heterogeneous exclusive CCA range” (HECR. However, when they are separated beyond the HECR, WPAN can still sense the 802.11 HR/DSSS signals, but WLAN loses its sense to the 802.15.4b signals. The asymmetric CCA leads to WPAN traffic in a position secondary to WLAN traffic.

Engineering of an E. coli outer membrane protein FhuA with increased channel diameter

Directory of Open Access Journals (Sweden)

Dworeck Tamara

2011-08-01

Full Text Available Abstract Background Channel proteins like FhuA can be an alternative to artificial chemically synthesized nanopores. To reach such goals, channel proteins must be flexible enough to be modified in their geometry, i.e. length and diameter. As continuation of a previous study in which we addressed the lengthening of the channel, here we report the increasing of the channel diameter by genetic engineering. Results The FhuA Δ1-159 diameter increase has been obtained by doubling the amino acid sequence of the first two N-terminal β-strands, resulting in variant FhuA Δ1-159 Exp. The total number of β-strands increased from 22 to 24 and the channel surface area is expected to increase by ~16%. The secondary structure analysis by circular dichroism (CD spectroscopy shows a high β-sheet content, suggesting the correct folding of FhuA Δ1-159 Exp. To further prove the FhuA Δ1-159 Exp channel functionality, kinetic measurement using the HRP-TMB assay (HRP = Horse Radish Peroxidase, TMB = 3,3',5,5'-tetramethylbenzidine were conducted. The results indicated a 17% faster diffusion kinetic for FhuA Δ1-159 Exp as compared to FhuA Δ1-159, well correlated to the expected channel surface area increase of ~16%. Conclusion In this study using a simple "semi rational" approach the FhuA Δ1-159 diameter was enlarged. By combining the actual results with the previous ones on the FhuA Δ1-159 lengthening a new set of synthetic nanochannels with desired lengths and diameters can be produced, broadening the FhuA Δ1-159 applications. As large scale protein production is possible our approach can give a contribution to nanochannel industrial applications.

Protein kinase A-induced internalization of Slack channels from the neuronal membrane occurs by adaptor protein-2/clathrin-mediated endocytosis.

Science.gov (United States)

Gururaj, Sushmitha; Evely, Katherine M; Pryce, Kerri D; Li, Jun; Qu, Jun; Bhattacharjee, Arin

2017-11-24

The sodium-activated potassium (K Na ) channel Kcnt1 (Slack) is abundantly expressed in nociceptor (pain-sensing) neurons of the dorsal root ganglion (DRG), where they transmit the large outward conductance I KNa and arbitrate membrane excitability. Slack channel expression at the DRG membrane is necessary for their characteristic firing accommodation during maintained stimulation, and reduced membrane channel density causes hyperexcitability. We have previously shown that in a pro-inflammatory state, a decrease in membrane channel expression leading to reduced Slack-mediated I KNa expression underlies DRG neuronal sensitization. An important component of the inflammatory milieu, PKA internalizes Slack channels from the DRG membrane, reduces I KNa , and produces DRG neuronal hyperexcitability when activated in cultured primary DRG neurons. Here, we show that this PKA-induced retrograde trafficking of Slack channels also occurs in intact spinal cord slices and that it is carried out by adaptor protein-2 (AP-2) via clathrin-mediated endocytosis. We provide mass spectrometric and biochemical evidence of an association of native neuronal AP-2 adaptor proteins with Slack channels, facilitated by a dileucine motif housed in the cytoplasmic Slack C terminus that binds AP-2. By creating a competitive peptide blocker of AP-2-Slack binding, we demonstrated that this interaction is essential for clathrin recruitment to the DRG membrane, Slack channel endocytosis, and DRG neuronal hyperexcitability after PKA activation. Together, these findings uncover AP-2 and clathrin as players in Slack channel regulation. Given the significant role of Slack in nociceptive neuronal excitability, the AP-2 clathrin-mediated endocytosis trafficking mechanism may enable targeting of peripheral and possibly, central neuronal sensitization. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Lipid bilayer-bound conformation of an integral membrane beta barrel protein by multidimensional MAS NMR

International Nuclear Information System (INIS)

Eddy, Matthew T.; Su, Yongchao; Silvers, Robert; Andreas, Loren; Clark, Lindsay; Wagner, Gerhard; Pintacuda, Guido; Emsley, Lyndon; Griffin, Robert G.

2015-01-01

The human voltage dependent anion channel 1 (VDAC) is a 32 kDa β-barrel integral membrane protein that controls the transport of ions across the outer mitochondrial membrane. Despite the determination of VDAC solution and diffraction structures, a structural basis for the mechanism of its function is not yet fully understood. Biophysical studies suggest VDAC requires a lipid bilayer to achieve full function, motivating the need for atomic resolution structural information of VDAC in a membrane environment. Here we report an essential step toward that goal: extensive assignments of backbone and side chain resonances for VDAC in DMPC lipid bilayers via magic angle spinning nuclear magnetic resonance (MAS NMR). VDAC reconstituted into DMPC lipid bilayers spontaneously forms two-dimensional lipid crystals, showing remarkable spectral resolution (0.5–0.3 ppm for 13 C line widths and <0.5 ppm 15 N line widths at 750 MHz). In addition to the benefits of working in a lipid bilayer, several distinct advantages are observed with the lipid crystalline preparation. First, the strong signals and sharp line widths facilitated extensive NMR resonance assignments for an integral membrane β-barrel protein in lipid bilayers by MAS NMR. Second, a large number of residues in loop regions were readily observed and assigned, which can be challenging in detergent-solubilized membrane proteins where loop regions are often not detected due to line broadening from conformational exchange. Third, complete backbone and side chain chemical shift assignments could be obtained for the first 25 residues, which comprise the functionally important N-terminus. The reported assignments allow us to compare predicted torsion angles for VDAC prepared in DMPC 2D lipid crystals, DMPC liposomes, and LDAO-solubilized samples to address the possible effects of the membrane mimetic environment on the conformation of the protein. Concluding, we discuss the strengths and weaknesses of the reported

G-protein-coupled inward rectifier potassium channels involved in corticostriatal presynaptic modulation.

Science.gov (United States)

Meneses, David; Mateos, Verónica; Islas, Gustavo; Barral, Jaime

2015-09-01

Presynaptic modulation has been associated mainly with calcium channels but recent data suggests that inward rectifier potassium channels (K(IR)) also play a role. In this work we set to characterize the role of presynaptic K(IR) channels in corticostriatal synaptic transmission. We elicited synaptic potentials in striatum by stimulating cortical areas and then determined the synaptic responses of corticostriatal synapsis by using paired pulse ratio (PPR) in the presence and absence of several potassium channel blockers. Unspecific potassium channels blockers Ba(2+) and Cs(+) reduced the PPR, suggesting that these channels are presynaptically located. Further pharmacological characterization showed that application of tertiapin-Q, a specific K(IR)3 channel family blocker, also induced a reduction of PPR, suggesting that K(IR)3 channels are present at corticostriatal terminals. In contrast, exposure to Lq2, a specific K(IR)1.1 inward rectifier potassium channel, did not induce any change in PPR suggesting the absence of these channels in the presynaptic corticostriatal terminals. Our results indicate that K(IR)3 channels are functionally expressed at the corticostriatal synapses, since blockage of these channels result in PPR decrease. Our results also help to explain how synaptic activity may become sensitive to extracellular signals mediated by G-protein coupled receptors. A vast repertoire of receptors may influence neurotransmitter release in an indirect manner through regulation of K(IR)3 channels. © 2015 Wiley Periodicals, Inc.

Inferior Olive HCN1 Channels Coordinate Synaptic Integration and Complex Spike Timing

Directory of Open Access Journals (Sweden)

Derek L.F. Garden

2018-02-01

Full Text Available Cerebellar climbing-fiber-mediated complex spikes originate from neurons in the inferior olive (IO, are critical for motor coordination, and are central to theories of cerebellar learning. Hyperpolarization-activated cyclic-nucleotide-gated (HCN channels expressed by IO neurons have been considered as pacemaker currents important for oscillatory and resonant dynamics. Here, we demonstrate that in vitro, network actions of HCN1 channels enable bidirectional glutamatergic synaptic responses, while local actions of HCN1 channels determine the timing and waveform of synaptically driven action potentials. These roles are distinct from, and may complement, proposed pacemaker functions of HCN channels. We find that in behaving animals HCN1 channels reduce variability in the timing of cerebellar complex spikes, which serve as a readout of IO spiking. Our results suggest that spatially distributed actions of HCN1 channels enable the IO to implement network-wide rules for synaptic integration that modulate the timing of cerebellar climbing fiber signals.

Differential and integral forms in supergauge theories and supergravity

International Nuclear Information System (INIS)

Zupnik, B.M.; Pak, D.G.

1989-01-01

D = 3, 4, N = 1 supergauge theories and D = 3, N = 1 supergravity are considered in the superfield formalism by using differential and integral forms. A special map of the space of differential forms into the space of integral forms is proposed. By means of this map we find the superfield Chern-Simons terms in D = 3, N = 1 Yang-Mills theory and supergravity. The integral forms corresponding to superfield invariants of D = 4, N = 1 supergauge theory have also been constructed. (Author)

Identification of a probable pore-forming domain in the multimeric vacuolar anion channel AtALMT9.

Science.gov (United States)

Zhang, Jingbo; Baetz, Ulrike; Krügel, Undine; Martinoia, Enrico; De Angeli, Alexis

2013-10-01

Aluminum-activated malate transporters (ALMTs) form an important family of anion channels involved in fundamental physiological processes in plants. Because of their importance, the role of ALMTs in plant physiology is studied extensively. In contrast, the structural basis of their functional properties is largely unknown. This lack of information limits the understanding of the functional and physiological differences between ALMTs and their impact on anion transport in plants. This study aimed at investigating the structural organization of the transmembrane domain of the Arabidopsis (Arabidopsis thaliana) vacuolar channel AtALMT9. For that purpose, we performed a large-scale mutagenesis analysis and found two residues that form a salt bridge between the first and second putative transmembrane α-helices (TMα1 and TMα2). Furthermore, using a combination of pharmacological and mutagenesis approaches, we identified citrate as an "open channel blocker" of AtALMT9 and used this tool to examine the inhibition sensitivity of different point mutants of highly conserved amino acid residues. By this means, we found a stretch within the cytosolic moiety of the TMα5 that is a probable pore-forming domain. Moreover, using a citrate-insensitive AtALMT9 mutant and biochemical approaches, we could demonstrate that AtALMT9 forms a multimeric complex that is supposedly composed of four subunits. In summary, our data provide, to our knowledge, the first evidence about the structural organization of an ion channel of the ALMT family. We suggest that AtALMT9 is a tetramer and that the TMα5 domains of the subunits contribute to form the pore of this anion channel.

Super-Quantum Mechanics in the Integral Form Formalism

Science.gov (United States)

Castellani, L.; Catenacci, R.; Grassi, P. A.

2018-05-01

We reformulate Super Quantum Mechanics in the context of integral forms. This framework allows to interpolate between different actions for the same theory, connected by different choices of Picture Changing Operators (PCO). In this way we retrieve component and superspace actions, and prove their equivalence. The PCO are closed integral forms, and can be interpreted as super Poincar\\'e duals of bosonic submanifolds embedded into a supermanifold.. We use them to construct Lagrangians that are top integral forms, and therefore can be integrated on the whole supermanifold. The $D=1, ~N=1$ and the $D=1,~ N=2$ cases are studied, in a flat and in a curved supermanifold. In this formalism we also consider coupling with gauge fields, Hilbert space of quantum states and observables.

Effect of ceramic membrane channel diameter on limiting retentate protein concentration during skim milk microfiltration.

Science.gov (United States)

Adams, Michael C; Barbano, David M

2016-01-01

Our objective was to determine the effect of retentate flow channel diameter (4 or 6mm) of nongraded permeability 100-nm pore size ceramic membranes operated in nonuniform transmembrane pressure mode on the limiting retentate protein concentration (LRPC) while microfiltering (MF) skim milk at a temperature of 50°C, a flux of 55 kg · m(-2) · h(-1), and an average cross-flow velocity of 7 m · s(-1). At the above conditions, the retentate true protein concentration was incrementally increased from 7 to 11.5%. When temperature, flux, and average cross-flow velocity were controlled, ceramic membrane retentate flow channel diameter did not affect the LRPC. This indicates that LRPC is not a function of the Reynolds number. Computational fluid dynamics data, which indicated that both membranes had similar radial velocity profiles within their retentate flow channels, supported this finding. Membranes with 6-mm flow channels can be operated at a lower pressure decrease from membrane inlet to membrane outlet (ΔP) or at a higher cross-flow velocity, depending on which is controlled, than membranes with 4-mm flow channels. This implies that 6-mm membranes could achieve a higher LRPC than 4-mm membranes at the same ΔP due to an increase in cross-flow velocity. In theory, the higher LRPC of the 6-mm membranes could facilitate 95% serum protein removal in 2 MF stages with diafiltration between stages if no serum protein were rejected by the membrane. At the same flux, retentate protein concentration, and average cross-flow velocity, 4-mm membranes require 21% more energy to remove a given amount of permeate than 6-mm membranes, despite the lower surface area of the 6-mm membranes. Equations to predict skim milk MF retentate viscosity as a function of protein concentration and temperature are provided. Retentate viscosity, retentate recirculation pump frequency required to maintain a given cross-flow velocity at a given retentate viscosity, and retentate protein

Martian outflow channels: How did their source aquifers form, and why did they drain so rapidly?

Science.gov (United States)

Rodriguez, J Alexis P; Kargel, Jeffrey S; Baker, Victor R; Gulick, Virginia C; Berman, Daniel C; Fairén, Alberto G; Linares, Rogelio; Zarroca, Mario; Yan, Jianguo; Miyamoto, Hideaki; Glines, Natalie

2015-09-08

Catastrophic floods generated ~3.2 Ga by rapid groundwater evacuation scoured the Solar System's most voluminous channels, the southern circum-Chryse outflow channels. Based on Viking Orbiter data analysis, it was hypothesized that these outflows emanated from a global Hesperian cryosphere-confined aquifer that was infused by south polar meltwater infiltration into the planet's upper crust. In this model, the outflow channels formed along zones of superlithostatic pressure generated by pronounced elevation differences around the Highland-Lowland Dichotomy Boundary. However, the restricted geographic location of the channels indicates that these conditions were not uniform. Furthermore, some outflow channel sources are too high to have been fed by south polar basal melting. Using more recent mission data, we argue that during the Late Noachian fluvial and glacial sediments were deposited into a clastic wedge within a paleo-basin located in the southern circum-Chryse region, which at the time was completely submerged under a primordial northern plains ocean [corrected]. Subsequent Late Hesperian outflow channels were sourced from within these geologic materials and formed by gigantic groundwater outbursts driven by an elevated hydraulic head from the Valles Marineris region. Thus, our findings link the formation of the southern circum-Chryse outflow channels to ancient marine, glacial, and fluvial erosion and sedimentation.

Modulation of CaV1.2 calcium channel by neuropeptide W regulates vascular myogenic tone via G protein-coupled receptor 7.

Science.gov (United States)

Ji, Li; Zhu, Huayuan; Chen, Hong; Fan, Wenyong; Chen, Junjie; Chen, Jing; Zhu, Guoqing; Wang, Juejin

2015-12-01

Neuropeptide W (NPW), an endogenous ligand for the G protein-coupled receptor 7 (GPR7), was first found to make important roles in central nerve system. In periphery, NPW was also present and regulated intracellular calcium homeostasis by L-type calcium channels. This study was designed to discover the effects of NPW-GPR7 on the function of CaV1.2 calcium channels in the vascular smooth muscle cells (VSMCs) and vasotone of arterial vessels. By whole-cell patch clamp, we studied the effects of NPW-23, the active form of NPW, on the CaV1.2 channels in the heterologously transfected human embryonic kidney 293 cells and VSMCs isolated from rat. Living system was used to explore the physiological function of NPW-23 in arterial myogenic tone. To investigate the pathological relevance, NPW mRNA level of mesenteric arteries was measured in the hypertensive and normotensive rats. NPW's receptor GPR7 was coexpressed with CaV1.2 channels in arterial smooth muscle. NPW-23 increased the ICa,L in transfected human embryonic kidney 293 cells and VSMCs via GPR7, which could be abrogated by phospholipase C (PLC)/protein kinase C (PKC) inhibitors, not protein kinase A or protein kinase G inhibitor. After NPW-23 application, the expression of pan phospho-PKC was increased; moreover, intracellular diacylglycerol level, the second messenger catalyzed by PLC, was increased 1.5-2-fold. Application with NPW-23 increased pressure-induced vasotone of the rat mesenteric arteries. Importantly, the expression of NPW was decreased in the hypertensive rats. NPW-23 regulates ICa,L via GPR7, which is mediated by PLC/PKC signaling, and such a mechanism plays a role in modulating vascular myogenic tone, which may involve in the development of vascular hypertension.

Do cysteine residues regulate transient receptor potential canonical type 6 (TRPC6) channel protein expression?

DEFF Research Database (Denmark)

Thilo, Florian; Liu, Ying; Krueger, Katharina

2012-01-01

The regulation of calcium influx through transient receptor potential canonical type 6 channel is mandatory for the activity of human monocytes. We submit the first evidence that cysteine residues of homocysteine or acetylcysteine affect TRPC6 expression in human monocytes. We observed that patie......The regulation of calcium influx through transient receptor potential canonical type 6 channel is mandatory for the activity of human monocytes. We submit the first evidence that cysteine residues of homocysteine or acetylcysteine affect TRPC6 expression in human monocytes. We observed...... that patients with chronic renal failure had significantly elevated homocysteine levels and TRPC6 mRNA expression levels in monocytes compared to control subjects. We further observed that administration of homocysteine or acetylcysteine significantly increased TRPC6 channel protein expression compared...... to control conditions. We therefore hypothesize that cysteine residues increase TRPC6 channel protein expression in humans....

The use of dansyl-calmodulin to study interactions with channels and other proteins.

Science.gov (United States)

Alaimo, Alessandro; Malo, Covadonga; Areso, Pilar; Aloria, Kerman; Millet, Oscar; Villarroel, Alvaro

2013-01-01

Steady-state fluorescence spectroscopy is a biophysical technique widely employed to characterize -interactions between proteins in vitro. Only a few proteins naturally fluoresce in cells, but by covalently attaching fluorophores virtually all proteins can be monitored. One of the first extrinsic fluorescent probes to be developed, and that is still in use, is dansyl chloride. We have used this method to monitor the interaction of a variety of proteins, including ion channels, with the Ca(2+)-dependent regulatory protein calmodulin. Here we describe the preparation and use of dansyl-calmodulin (D-CaM).

Development of multi-channel gated integrator and PXI-DAQ system for nuclear detector arrays

International Nuclear Information System (INIS)

Kong Jie; Su Hong; Chen Zhiqiang; Dong Chengfu; Qian Yi; Gao Shanshan; Zhou Chaoyang; Lu Wan; Ye Ruiping; Ma Junbing

2010-01-01

A multi-channel gated integrator and PXI based data acquisition system have been developed for nuclear detector arrays with hundreds of detector units. The multi-channel gated integrator can be controlled by a programmable GI controller. The PXI-DAQ system consists of NI PXI-1033 chassis with several PXI-DAQ cards. The system software has a user-friendly GUI which is written in C language using LabWindows/CVI under Windows XP operating system. The performance of the PXI-DAQ system is very reliable and capable of handling event rate up to 40 kHz.

The integral form of supergravity

Energy Technology Data Exchange (ETDEWEB)

Castellani, L. [Dipartimento di Scienze e Innovazione Tecnologica, Università del Piemonte Orientale,Viale T. Michel, 11, 15121 Alessandria (Italy); INFN - Sezione di Torino,via P. Giuria 1, 10125 Torino (Italy); Catenacci, R. [Dipartimento di Scienze e Innovazione Tecnologica, Università del Piemonte Orientale,Viale T. Michel, 11, 15121 Alessandria (Italy); Gruppo Nazionale di Fisica Matematica, INdAM,P.le Aldo Moro 5, 00185 Roma (Italy); Grassi, P.A. [Dipartimento di Scienze e Innovazione Tecnologica, Università del Piemonte Orientale,Viale T. Michel, 11, 15121 Alessandria (Italy); INFN - Sezione di Torino,via P. Giuria 1, 10125 Torino (Italy)

2016-10-11

By using integral forms we derive the superspace action of D=3,N=1 supergravity as an integral on a supermanifold. The construction is based on target space picture changing operators, here playing the rôle of Poincaré duals to the lower-dimensional spacetime surfaces embedded into the supermanifold. We show how the group geometrical action based on the group manifold approach interpolates between the superspace and the component supergravity actions, thus providing another proof of their equivalence.

Molecular properties of mammalian proteins that interact with cGMP: protein kinases, cation channels, phosphodiesterases, and multi-drug anion transporters.

Science.gov (United States)

Francis, Sharron H; Blount, Mitsi A; Zoraghi, Roya; Corbin, Jackie D

2005-09-01

Cyclic GMP is a critical second messenger signaling molecule in many mammalian cell types. It is synthesized by a family of guanylyl cyclases that is activated in response to stimuli from hormones such as natriuretic peptides, members of the guanylin family, and chemical stimuli including nitric oxide and carbon monoxide. The resulting elevation of cGMP modulates myriad physiological processes. Three major groups of cellular proteins bind cGMP specifically at allosteric sites; interaction of cGMP with these sites modulates the activities and functions of other domains within these protein groups to bring about physiological effects. These proteins include the cyclic nucleotide (cN)-dependent protein kinases, cN-gated cation channels, and cGMP-binding phosphodiesterases (PDE). Cyclic GMP also interacts with the catalytic sites of many cN PDEs and with some members of the multi-drug anion transporter family (MRPs) which can extrude nucleotides from cells. The allosteric cN-binding sites in the kinases and the cN-gated channels are evolutionarily and biochemically related, whereas the allosteric cGMP-binding sites in PDEs (also known as GAF domains), the catalytic sites of PDEs , and the ligand-binding sites in the MRPs are evolutionarily and biochemically distinct from each other and from those in the kinase and channel families. The sites that interact with cGMP within each of these groups of proteins have unique properties that provide for cGMP binding. Within a given cell, cGMP can potentially interact with members of all these groups of proteins if they are present. The relative abundance and affinities of these various cGMP-binding sites in conjunction with their subcellular compartmentation, proximity to cyclases and PDEs, and post-translational modification contribute importantly in determining the impact of these respective proteins to cGMP signaling within a particular cell.

Point mutations in the transmembrane region of the clic1 ion channel selectively modify its biophysical properties.

Directory of Open Access Journals (Sweden)

Stefania Averaimo

Full Text Available Chloride intracellular Channel 1 (CLIC1 is a metamorphic protein that changes from a soluble cytoplasmic protein into a transmembrane protein. Once inserted into membranes, CLIC1 multimerises and is able to form chloride selective ion channels. Whilst CLIC1 behaves as an ion channel both in cells and in artificial lipid bilayers, its structure in the soluble form has led to some uncertainty as to whether it really is an ion channel protein. CLIC1 has a single putative transmembrane region that contains only two charged residues: arginine 29 (Arg29 and lysine 37 (Lys37. As charged residues are likely to have a key role in ion channel function, we hypothesized that mutating them to neutral alanine to generate K37A and R29A CLIC1 would alter the electrophysiological characteristics of CLIC1. By using three different electrophysiological approaches: i single channel Tip-Dip in artificial bilayers using soluble recombinant CLIC1, ii cell-attached and iii whole-cell patch clamp recordings in transiently transfected HEK cells, we determined that the K37A mutation altered the single-channel conductance while the R29A mutation affected the single-channel open probability in response to variation in membrane potential. Our results show that mutation of the two charged amino acids (K37 and R29 in the putative transmembrane region of CLIC1 alters the biophysical properties of the ion channel in both artificial bilayers and cells. Hence these charged residues are directly involved in regulating its ion channel activity. This strongly suggests that, despite its unusual structure, CLIC1 itself is able to form a chloride ion channel.

N-(2-methoxyphenyl) benzenesulfonamide, a novel regulator of neuronal G protein-gated inward rectifier K+ channels.

Science.gov (United States)

Walsh, Kenneth B; Gay, Elaine A; Blough, Bruce E; Geurkink, David W

2017-11-15

G protein-gated inward rectifier K + (GIRK) channels are members of the super-family of proteins known as inward rectifier K + (Kir) channels and are expressed throughout the peripheral and central nervous systems. Neuronal GIRK channels are the downstream targets of a number of neuromodulators including opioids, somatostatin, dopamine and cannabinoids. Previous studies have demonstrated that the ATP-sensitive K + channel, another member of the Kir channel family, is regulated by sulfonamide drugs. Therefore, to determine if sulfonamides also modulate GIRK channels, we screened a library of arylsulfonamide compounds using a GIRK channel fluorescent assay that utilized pituitary AtT20 cells expressing GIRK channels along with the somatostatin type-2 and -5 receptors. Enhancement of the GIRK channel fluorescent signal by one compound, N-(2-methoxyphenyl) benzenesulfonamide (MPBS), was dependent on the activation of the channel by somatostatin. In whole-cell patch clamp experiments, application of MPBS both shifted the somatostatin concentration-response curve (EC 50 = 3.5nM [control] vs.1.0nM [MPBS]) for GIRK channel activation and increased the maximum GIRK current measured with 100nM somatostatin. However, GIRK channel activation was not observed when MPBS was applied to the cells in the absence of somatostatin. While the MPBS structural analog 4-fluoro-N-(2-methoxyphenyl) benzenesulfonamide also augmented the somatostatin-induced GIRK fluorescent signal, no increase in the signal was observed with the sulfonamides tolbutamide, sulfapyridine and celecoxib. In conclusion, MPBS represents a novel prototypic GPCR-dependent regulator of neuronal GIRK channels. Copyright © 2017 Elsevier B.V. All rights reserved.

The Business Cluster's Distribution e-Channels

OpenAIRE

Milan Davidovic

2011-01-01

The business cluster cooperative potential and business capability improvement are dependent on e-business implementation and business model change dynamics in cluster and his members based in new and existing distribution channels, customer relationships management and supplychain integration. In this work analyse cluster’s e-business models, e-commerce forms and distribution e-channels for three business cases: when cluster members are oriented on own business, on cooperative’s project or c...

Spontaneous formation of structurally diverse membrane channel architectures from a single antimicrobial peptide

Science.gov (United States)

Wang, Yukun; Chen, Charles H.; Hu, Dan; Ulmschneider, Martin B.; Ulmschneider, Jakob P.

2016-11-01

Many antimicrobial peptides (AMPs) selectively target and form pores in microbial membranes. However, the mechanisms of membrane targeting, pore formation and function remain elusive. Here we report an experimentally guided unbiased simulation methodology that yields the mechanism of spontaneous pore assembly for the AMP maculatin at atomic resolution. Rather than a single pore, maculatin forms an ensemble of structurally diverse temporarily functional low-oligomeric pores, which mimic integral membrane protein channels in structure. These pores continuously form and dissociate in the membrane. Membrane permeabilization is dominated by hexa-, hepta- and octamers, which conduct water, ions and small dyes. Pores form by consecutive addition of individual helices to a transmembrane helix or helix bundle, in contrast to current poration models. The diversity of the pore architectures--formed by a single sequence--may be a key feature in preventing bacterial resistance and could explain why sequence-function relationships in AMPs remain elusive.

G Protein Regulation of Neuronal Calcium Channels: Back to the Future

Czech Academy of Sciences Publication Activity Database

Proft, Juliane; Weiss, Norbert

2015-01-01

Roč. 87, č. 6 (2015), s. 890-906 ISSN 0026-895X R&D Projects: GA ČR GA15-13556S Institutional support: RVO:61388963 Keywords : voltage gated calcium channels Cav * G proteins * GPCR Subject RIV: CE - Biochemistry Impact factor: 3.931, year: 2015

Crystal Structure of the Mammalian GIRK2 K+ Channel and Gating Regulation by G-Proteins, PIP2 and Sodium

Science.gov (United States)

Whorton, Matthew R.; MacKinnon, Roderick

2011-01-01

Summary G-protein-gated K+ channels (Kir3.1–Kir3.4) control electrical excitability in many different cells. Among their functions relevant to human physiology and disease, they regulate the heart rate and govern a wide range of neuronal activities. Here we present the first crystal structures of a G-protein-gated K+ channel. By comparing the wild-type structure to that of a constitutively active mutant, we identify a global conformational change through which G-proteins could open a G-loop gate in the cytoplasmic domain. The structures of both channels in the absence and presence of PIP2 show that G-proteins open only the G-loop gate in the absence of PIP2, but in the presence of PIP2 the G-loop gate and a second inner helix gate become coupled, so that both gates open. We also identify a strategically located Na+ ion-binding site, which would allow intracellular Na+ to modulate GIRK channel activity. These data provide a mechanistic description of multi-ligand regulation of GIRK channel gating. PMID:21962516

Polycystin-1 Is a Cardiomyocyte Mechanosensor That Governs L-Type Ca2+ Channel Protein Stability.

Science.gov (United States)

Pedrozo, Zully; Criollo, Alfredo; Battiprolu, Pavan K; Morales, Cyndi R; Contreras-Ferrat, Ariel; Fernández, Carolina; Jiang, Nan; Luo, Xiang; Caplan, Michael J; Somlo, Stefan; Rothermel, Beverly A; Gillette, Thomas G; Lavandero, Sergio; Hill, Joseph A

2015-06-16

L-type calcium channel activity is critical to afterload-induced hypertrophic growth of the heart. However, the mechanisms governing mechanical stress-induced activation of L-type calcium channel activity are obscure. Polycystin-1 (PC-1) is a G protein-coupled receptor-like protein that functions as a mechanosensor in a variety of cell types and is present in cardiomyocytes. We subjected neonatal rat ventricular myocytes to mechanical stretch by exposing them to hypo-osmotic medium or cyclic mechanical stretch, triggering cell growth in a manner dependent on L-type calcium channel activity. RNAi-dependent knockdown of PC-1 blocked this hypertrophy. Overexpression of a C-terminal fragment of PC-1 was sufficient to trigger neonatal rat ventricular myocyte hypertrophy. Exposing neonatal rat ventricular myocytes to hypo-osmotic medium resulted in an increase in α1C protein levels, a response that was prevented by PC-1 knockdown. MG132, a proteasomal inhibitor, rescued PC-1 knockdown-dependent declines in α1C protein. To test this in vivo, we engineered mice harboring conditional silencing of PC-1 selectively in cardiomyocytes (PC-1 knockout) and subjected them to mechanical stress in vivo (transverse aortic constriction). At baseline, PC-1 knockout mice manifested decreased cardiac function relative to littermate controls, and α1C L-type calcium channel protein levels were significantly lower in PC-1 knockout hearts. Whereas control mice manifested robust transverse aortic constriction-induced increases in cardiac mass, PC-1 knockout mice showed no significant growth. Likewise, transverse aortic constriction-elicited increases in hypertrophic markers and interstitial fibrosis were blunted in the knockout animals PC-1 is a cardiomyocyte mechanosensor that is required for cardiac hypertrophy through a mechanism that involves stabilization of α1C protein. © 2015 American Heart Association, Inc.

Application of amphipols for structure-functional analysis of TRP channels.

Science.gov (United States)

Huynh, Kevin W; Cohen, Matthew R; Moiseenkova-Bell, Vera Y

2014-10-01

Amphipathic polymers (amphipols), such as A8-35 and SApol, are a new tool for stabilizing integral membrane proteins in detergent-free conditions for structural and functional studies. Transient receptor potential (TRP) ion channels function as tetrameric protein complexes in a diverse range of cellular processes including sensory transduction. Mammalian TRP channels share ~20 % sequence similarity and are categorized into six subfamilies: TRPC (canonical), TRPV (vanilloid), TRPA (ankyrin), TRPM (melastatin), TRPP (polycystin), and TRPML (mucolipin). Due to the inherent difficulties in purifying eukaryotic membrane proteins, structural studies of TRP channels have been limited. Recently, A8-35 was essential in resolving the molecular architecture of the nociceptor TRPA1 and led to the determination of a high-resolution structure of the thermosensitive TRPV1 channel by cryo-EM. Newly developed maltose-neopentyl glycol (MNG) detergents have also proven to be useful in stabilizing TRP channels for structural analysis. In this review, we will discuss the impacts of amphipols and MNG detergents on structural studies of TRP channels by cryo-EM. We will compare how A8-35 and MNG detergents interact with the hydrophobic transmembrane domains of TRP channels. In addition, we will discuss what these cryo-EM studies reveal on the importance of screening different types of surfactants toward determining high-resolution structures of TRP channels.

Peptidoglycan-associated outer membrane protein Mep45 of rumen anaerobe Selenomonas ruminantium forms a non-specific diffusion pore via its C-terminal transmembrane domain.

Science.gov (United States)

Kojima, Seiji; Hayashi, Kanako; Tochigi, Saeko; Kusano, Tomonobu; Kaneko, Jun; Kamio, Yoshiyuki

2016-10-01

The major outer membrane protein Mep45 of Selenomonas ruminantium, an anaerobic Gram-negative bacterium, comprises two distinct domains: the N-terminal S-layer homologous (SLH) domain that protrudes into the periplasm and binds to peptidoglycan, and the remaining C-terminal transmembrane domain, whose function has been unknown. Here, we solubilized and purified Mep45 and characterized its function using proteoliposomes reconstituted with Mep45. We found that Mep45 forms a nonspecific diffusion channel via its C-terminal region. The channel was permeable to solutes smaller than a molecular weight of roughly 600, and the estimated pore radius was 0.58 nm. Truncation of the SLH domain did not affect the channel property. On the basis of the fact that Mep45 is the most abundant outer membrane protein in S. ruminantium, we conclude that Mep45 serves as a main pathway through which small solutes diffuse across the outer membrane of this bacterium.

DC biased low-frequency insulating constriction dielectrophoresis for protein biomolecules concentration.

Science.gov (United States)

Zhang, Peng; Liu, Yuxin

2017-09-01

Sample enrichment or molecules concentration is considered an essential step in sample processing of miniaturized devices aimed at biosensing and bioanalysis. Among all the means involved to achieve this aim, dielectrophoresis (DEP) is increasingly employed in molecules manipulation and concentration because it is non-destructive and high efficiency. This paper presents a methodology to achieve protein concentration utilizing the combination effects of electrokinetics and low frequency insulating dielectrophoresis (iDEP) generated within a microfluidic device, in which a submicron constricted channel was fabricated using DNA molecular combing and replica molding. This fabrication technique avoids using e-beam lithography or other complicated nanochannel fabrication methods, and provides an easy and low cost approach with the flexibility of controlling channel dimensions to create highly constricted channels embedded in a microfluidic device. With theoretical analysis and experiments, we demonstrated that fluorescein isothiocyanate conjugated bovine serum albumin (FITC-BSA) protein molecules can be significantly concentrated to form an arc-shaped band near the constricted channel under the effects of a negative dielectrophoretic force and DC electrokinetic forces within a short period of time. It was also observed that the amplitudes of the applied DC and AC electric fields, the AC frequencies as well as the suspending medium conductivities had strong effects on the concentration responses of the FITC-BSA molecules, including the concentrated area and position, intensities of the focused molecules, and concentration speed. Our method provides a simple and flexible approach for quickly concentrating protein molecules by controlling the applied electric field parameters. The iDEP device reported in this paper can be used as a stand-alone sensor or worked as a pre-concentration module integrated with biosensors for protein biomarker detection. Furthermore, low

Tuning the allosteric regulation of artificial muscarinic and dopaminergic ligand-gated potassium channels by protein engineering of G protein-coupled receptors

Science.gov (United States)

Moreau, Christophe J.; Revilloud, Jean; Caro, Lydia N.; Dupuis, Julien P.; Trouchet, Amandine; Estrada-Mondragón, Argel; Nieścierowicz, Katarzyna; Sapay, Nicolas; Crouzy, Serge; Vivaudou, Michel

2017-01-01

Ligand-gated ion channels enable intercellular transmission of action potential through synapses by transducing biochemical messengers into electrical signal. We designed artificial ligand-gated ion channels by coupling G protein-coupled receptors to the Kir6.2 potassium channel. These artificial channels called ion channel-coupled receptors offer complementary properties to natural channels by extending the repertoire of ligands to those recognized by the fused receptors, by generating more sustained signals and by conferring potassium selectivity. The first artificial channels based on the muscarinic M2 and the dopaminergic D2L receptors were opened and closed by acetylcholine and dopamine, respectively. We find here that this opposite regulation of the gating is linked to the length of the receptor C-termini, and that C-terminus engineering can precisely control the extent and direction of ligand gating. These findings establish the design rules to produce customized ligand-gated channels for synthetic biology applications. PMID:28145461

[G-protein potentiates the activation of TNF-alpha on calcium-activated potassium channel in ECV304].

Science.gov (United States)

Lin, L; Zheng, Y; Qu, J; Bao, G

2000-06-01

Observe the effect of tumor necrosis factor-alpha (TNF-alpha) on calcium-activated potassium channel in ECV304 and the possible involvement of G-protein mediation in the action of TNF-alpha. Using the cell-attached configuration of patch clamp technique. (1) the activity of high-conductance calcium-activated potassium channel (BKca) was recorded. Its conductance is (202.54 +/- 16.62) pS; (2) the activity of BKca was potentiated by 200 U/ml TNF-alpha; (3) G-protein would intensify this TNF-alpha activation. TNF-alpha acted on vascular endothelial cell ECV304 could rapidly activate the activity of BKca. Opening of BKca resulted in membrane hyper-polarization which could increase electro-chemical gradient for the resting Ca2+ influx and open leakage calcium channel, thus resting cytoplasmic free Ca2+ concentration could be elevated. G-protein may exert an important regulation in this process.

Surface expression, single-channel analysis and membrane topology of recombinant Chlamydia trachomatis Major Outer Membrane Protein

Directory of Open Access Journals (Sweden)

McClafferty Heather

2005-01-01

Full Text Available Abstract Background Chlamydial bacteria are obligate intracellular pathogens containing a cysteine-rich porin (Major Outer Membrane Protein, MOMP with important structural and, in many species, immunity-related roles. MOMP forms extensive disulphide bonds with other chlamydial proteins, and is difficult to purify. Leaderless, recombinant MOMPs expressed in E. coli have yet to be refolded from inclusion bodies, and although leadered MOMP can be expressed in E. coli cells, it often misfolds and aggregates. We aimed to improve the surface expression of correctly folded MOMP to investigate the membrane topology of the protein, and provide a system to display native and modified MOMP epitopes. Results C. trachomatis MOMP was expressed on the surface of E. coli cells (including "porin knockout" cells after optimizing leader sequence, temperature and medium composition, and the protein was functionally reconstituted at the single-channel level to confirm it was folded correctly. Recombinant MOMP formed oligomers even in the absence of its 9 cysteine residues, and the unmodified protein also formed inter- and intra-subunit disulphide bonds. Its topology was modeled as a (16-stranded β-barrel, and specific structural predictions were tested by removing each of the four putative surface-exposed loops corresponding to highly immunogenic variable sequence (VS domains, and one or two of the putative transmembrane strands. The deletion of predicted external loops did not prevent folding and incorporation of MOMP into the E. coli outer membrane, in contrast to the removal of predicted transmembrane strands. Conclusions C. trachomatis MOMP was functionally expressed on the surface of E. coli cells under newly optimized conditions. Tests of its predicted membrane topology were consistent with β-barrel oligomers in which major immunogenic regions are displayed on surface-exposed loops. Functional surface expression, coupled with improved understanding of MOMP

Ion Transport in Confined Geometries below the Nanoscale: Access Resistance Dominates Protein Channel Conductance in Diluted Solutions.

Science.gov (United States)

Alcaraz, Antonio; López, M Lidón; Queralt-Martín, María; Aguilella, Vicente M

2017-10-24

Synthetic nanopores and mesoscopic protein channels have common traits like the importance of electrostatic interactions between the permeating ions and the nanochannel. Ion transport at the nanoscale occurs under confinement conditions so that the usual assumptions made in microfluidics are challenged, among others, by interfacial effects such as access resistance (AR). Here, we show that a sound interpretation of electrophysiological measurements in terms of channel ion selective properties requires the consideration of interfacial effects, up to the point that they dominate protein channel conductance in diluted solutions. We measure AR in a large ion channel, the bacterial porin OmpF, by means of single-channel conductance measurements in electrolyte solutions containing varying concentrations of high molecular weight PEG, sterically excluded from the pore. Comparison of experiments performed in charged and neutral planar membranes shows that lipid surface charges modify the ion distribution and determine the value of AR, indicating that lipid molecules are more than passive scaffolds even in the case of large transmembrane proteins. We also found that AR may reach up to 80% of the total channel conductance in diluted solutions, where electrophysiological recordings register essentially the AR of the system and depend marginally on the pore characteristics. These findings may have implications for several low aspect ratio biological channels that perform their physiological function in a low ionic strength and macromolecule crowded environment, just the two conditions enhancing the AR contribution.

Integrated microchip incorporating atomic magnetometer and microfluidic channel for NMR and MRI

Science.gov (United States)

Ledbetter, Micah P [Oakland, CA; Savukov, Igor M [Los Alamos, NM; Budker, Dmitry [El Cerrito, CA; Shah, Vishal K [Plainsboro, NJ; Knappe, Svenja [Boulder, CO; Kitching, John [Boulder, CO; Michalak, David J [Berkeley, CA; Xu, Shoujun [Houston, TX; Pines, Alexander [Berkeley, CA

2011-08-09

An integral microfluidic device includes an alkali vapor cell and microfluidic channel, which can be used to detect magnetism for nuclear magnetic resonance (NMR) and magnetic resonance imaging (MRI). Small magnetic fields in the vicinity of the vapor cell can be measured by optically polarizing and probing the spin precession in the small magnetic field. This can then be used to detect the magnetic field of in encoded analyte in the adjacent microfluidic channel. The magnetism in the microfluidic channel can be modulated by applying an appropriate series of radio or audio frequency pulses upstream from the microfluidic chip (the remote detection modality) to yield a sensitive means of detecting NMR and MRI.

An electronic channel switching-based aptasensor for ultrasensitive protein detection

Energy Technology Data Exchange (ETDEWEB)

Li Hongbo; Wang Cui [State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082 (China); Wu Zaisheng, E-mail: wuzaisheng@163.com [State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082 (China); Lu Limin; Qiu Liping; Zhou Hui; Shen Guoli [State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082 (China); Yu Ruqin, E-mail: rqyu@hnu.edu.cn [State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082 (China)

2013-01-03

Highlights: Black-Right-Pointing-Pointer Target IgE is successfully designed to serve as a barrier to separate enzyme from its substrate. Black-Right-Pointing-Pointer This sensing platform of electronic channel switching-based aptasensor can be simply manipulated. Black-Right-Pointing-Pointer The stable hairpin structure of anti-IgE aptamer is utilized to detect target IgE. Black-Right-Pointing-Pointer The sensor is ultrasensitive sensitivity, excellent selectivity and small volume of sample. Black-Right-Pointing-Pointer It is a powerful platform to be further expanded to detect more kinds of proteins and even cells. - Abstract: Due to the ubiquity and essential of the proteins in all living organisms, the identification and quantification of disease-specific proteins are particularly important. Because the conformational change of aptamer upon its target or probe/target/probe sandwich often is the primary prerequisite for the design of an electrochemical aptameric assay system, it is extremely difficult to construct the electrochemical aptasensor for protein assay because the corresponding aptamers cannot often meet the requirement. To circumvent the obstacles mentioned, an electronic channel switching-based (ECS) aptasensor for ultrasensitive protein detection is developed. The essential achievement made is that an innovative sensing concept is proposed: the hairpin structure of aptamer is designed to pull electroactive species toward electrode surface and makes the surface-immobilized IgE serve as a barrier that separates enzyme from its substrate. It seemingly ensures that the ECS aptasensor exhibits most excellent assay features, such as, a detection limit of 4.44 Multiplication-Sign 10{sup -6} {mu}g mL{sup -1} (22.7 fM, 220 zmol in 10-{mu}L sample) (demonstrating a 5 orders of magnitude improvement in detection sensitivity compared with classical electronic aptasensors) and dynamic response range from 4.44 Multiplication-Sign 10{sup -6} to 4.44 Multiplication

Multi-Channel Capacitive Sensor Arrays

Directory of Open Access Journals (Sweden)

Bingnan Wang

2016-01-01

Full Text Available In this paper, multi-channel capacitive sensor arrays based on microstrip band-stop filters are studied. The sensor arrays can be used to detect the proximity of objects at different positions and directions. Each capacitive sensing structure in the array is connected to an inductive element to form resonance at different frequencies. The resonances are designed to be isolated in the frequency spectrum, such that the change in one channel does not affect resonances at other channels. The inductive element associated with each capacitive sensor can be surface-mounted inductors, integrated microstrip inductors or metamaterial-inspired structures. We show that by using metamaterial split-ring structures coupled to a microstrip line, the quality factor of each resonance can be greatly improved compared to conventional surface-mounted or microstrip meander inductors. With such a microstrip-coupled split-ring design, more sensing elements can be integrated in the same frequency spectrum, and the sensitivity can be greatly improved.

Modulating secretory pathway pH by proton channel co-expression can increase recombinant protein stability in plants.

Science.gov (United States)

Jutras, Philippe V; D'Aoust, Marc-André; Couture, Manon M-J; Vézina, Louis-Philippe; Goulet, Marie-Claire; Michaud, Dominique; Sainsbury, Frank

2015-09-01

Eukaryotic expression systems are used for the production of complex secreted proteins. However, recombinant proteins face considerable biochemical challenges along the secretory pathway, including proteolysis and pH variation between organelles. As the use of synthetic biology matures into solutions for protein production, various host-cell engineering approaches are being developed to ameliorate host-cell factors that can limit recombinant protein quality and yield. We report the potential of the influenza M2 ion channel as a novel tool to neutralize the pH in acidic subcellular compartments. Using transient expression in the plant host, Nicotiana benthamiana, we show that ion channel expression can significantly raise pH in the Golgi apparatus and that this can have a strong stabilizing effect on a fusion protein separated by an acid-susceptible linker peptide. We exemplify the utility of this effect in recombinant protein production using influenza hemagglutinin subtypes differentially stable at low pH; the expression of hemagglutinins prone to conformational change in mildly acidic conditions is considerably enhanced by M2 co-expression. The co-expression of a heterologous ion channel to stabilize acid-labile proteins and peptides represents a novel approach to increasing the yield and quality of secreted recombinant proteins in plants and, possibly, in other eukaryotic expression hosts. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

InSilico Proteomics System: Integration and Application of Protein and Protein-Protein Interaction Data using Microsoft .NET

Directory of Open Access Journals (Sweden)

Straßer Wolfgang

2006-12-01

Full Text Available In the last decades, biological databases became the major knowledge resource for researchers in the field of molecular biology. The distribution of information among these databases is one of the major problems. An overview about the subject area of data access and representation of protein and protein-protein interaction data within public biological databases is described. For a comprehensive and consistent way of searching and analysing integrated protein and protein-protein interaction data, the InSilico Proteomics (ISP project has been initiated. Its three main objectives are (1 to provide an integrated knowledge pool for data investigation and global network analysis functions for a better understanding of a cell’s interactome, (2 employment of public data for plausibility analysis and validation of in-house experimental data and (3 testing the applicability of Microsoft’s .NET architecture for bioinformatics applications. Data integrated into the ISP database can be queried through the Web portal PRIMOS (PRotein Interaction and MOlecule Search which is freely available at http://biomis.fh-hagenberg.at/isp/primos.

DEVELOPMENT OF MODEL FOR QUANTITATIVE EVALUATION OF DYNAMICALLY STABLE FORMS OF RIVER CHANNELS

Directory of Open Access Journals (Sweden)

O. V. Zenkin

2017-01-01

Full Text Available The article highlights the method of calculating the optimum curvature of the river channels using the kinematic model of the flow structure based on the concept of discrete nature of the channel process. It offers the analytic form of the equation of motion of river flow, which can be used simulation modeling for searching dynamically stable form of the river channel, and which can control water level in rivers. The source data for the illustrations of given in the article modeling methods have been served the images received from MODIS on the Terra satellite, for the lower reaches of the river Kur, which merges with the river Urmi, forming the Tunguska river – the left tributary of the Amur.The modified geometric method can be used to calculate obliquity of tangent to the curve and normal in those situations when observed on satellite imagery points are located on the coordinate of the network irregularly and when three points lying on the curve of the riverbed do not form isosceles triangle.The model assembles tangential and radial components of the forces acting on the water flow (centrifugal, friction and gravity. Curvature radius is explicitly expressed in the model through the parameter  – gradient angle relative to the axis X. As solution for the value of the angle  is searched, when the correlation function reaches its maximum. It is assumed that the riverbed shape “wrong” and could be modified so that the resulting curve better correlated with calculated curve. Morphometric dependences for macroforms allow creating series of morphological methods for the calculation of deformations and displacement of the shore in any section of meander scroll.The proposed technique has been tested also on satellite imagery of high resolution. The presented methods of calculation are used as the basis for hydrological projects of geoinformation systems oriented at prediction of morphodynamic processes and morphological evolution of river

Protein-Protein Interactions of Viroporins in Coronaviruses and Paramyxoviruses: New Targets for Antivirals?

Directory of Open Access Journals (Sweden)

Jaume Torres

2015-06-01

Full Text Available Viroporins are members of a rapidly growing family of channel-forming small polypeptides found in viruses. The present review will be focused on recent structural and protein-protein interaction information involving two viroporins found in enveloped viruses that target the respiratory tract; (i the envelope protein in coronaviruses and (ii the small hydrophobic protein in paramyxoviruses. Deletion of these two viroporins leads to viral attenuation in vivo, whereas data from cell culture shows involvement in the regulation of stress and inflammation. The channel activity and structure of some representative members of these viroporins have been recently characterized in some detail. In addition, searches for protein-protein interactions using yeast-two hybrid techniques have shed light on possible functional roles for their exposed cytoplasmic domains. A deeper analysis of these interactions should not only provide a more complete overview of the multiple functions of these viroporins, but also suggest novel strategies that target protein-protein interactions as much needed antivirals. These should complement current efforts to block viroporin channel activity.

A 64-channel integrated circuit for signal readout from coordinate detectors

International Nuclear Information System (INIS)

Aulchenko, V.; Shekhtman, L.; Zhulanov, V.

2017-01-01

A specialized integrated circuit was developed for the readout of signal from coordinate detectors of different types, including gas micro-pattern detectors and silicon microstrip detectors. The ASIC includes 64 channels, each containing a low-noise charge-sensitive amplifier with a connectable feedback capacitor and resistor, and fast reset of the feedback capacitor. Each channel of the ASIC also contains 100 cells of analogue memory where the signal can be stored at a rate of 10 MHz. The pitch of input pads is 50 μm and the chip size is 5× 5 mm 2 . The equivalent noise charge of the ASIC channel is about 2000 electrons with 10 pF capacitance at the input and maximal signal before saturation corresponds to 2× 10 6 electrons. The first application for this ASIC is the detector for imaging of explosions at a synchrotron radiation beam (DIMEX), where it has to substitute the old and slower APC128 ASIC. The full-size electronics including 8 ASICs for 512 channels was assembled and tested.

Mechanisms of Rose Bengal inhibition on SecA ATPase and ion channel activities.

Science.gov (United States)

Hsieh, Ying-Hsin; Huang, Ying-Ju; Jin, Jin-Shan; Yu, Liyan; Yang, Hsiuchin; Jiang, Chun; Wang, Binghe; Tai, Phang C

2014-11-14

SecA is an essential protein possessing ATPase activity in bacterial protein translocation for which Rose Bengal (RB) is the first reported sub-micromolar inhibitor in ATPase activity and protein translocation. Here, we examined the mechanisms of inhibition on various forms of SecA ATPase by conventional enzymatic assays, and by monitoring the SecA-dependent channel activity in the semi-physiological system in cells. We build on the previous observation that SecA with liposomes form active protein-conducting channels in the oocytes. Such ion channel activity is enhanced by purified Escherichia coli SecYEG-SecDF·YajC liposome complexes. Inhibition by RB could be monitored, providing correlation of in vitro activity and intact cell functionality. In this work, we found the intrinsic SecA ATPase is inhibited by RB competitively at low ATP concentration, and non-competitively at high ATP concentrations while the translocation ATPase with precursors and SecYEG is inhibited non-competitively by RB. The Inhibition by RB on SecA channel activity in the oocytes with exogenous ATP-Mg(2+), mimicking translocation ATPase activity, is also non-competitive. The non-competitive inhibition on channel activity has also been observed with SecA from other bacteria which otherwise would be difficult to examine without the cognate precursors and membranes. Copyright © 2014 Elsevier Inc. All rights reserved.

Development of supported biomimetic membranes for insertion of aquaporin protein water channels for novel water filtration applications

DEFF Research Database (Denmark)

Hansen, Jesper Søndergaard

). This constitutes a new methodology to correctly and functionally reconstitute membrane proteins in controllable amounts into giant vesicles. The method for formation of giant protein vesicles subsequently led to the first functional prototype of an aquaporin-membrane water filtration device.......Aquaporins represent a class of membrane protein channels found in all living organisms that selectively transport water molecules across biological membranes. The work presented in this thesis was motivated by the conceptual idea of incorporating aquaporin water channels into biomimetic membranes...... to develop novel water separation technologies. To accomplish this, it is necessary to construct an efficient platform to handle biomimetic membranes. Moreover, general methods are required to reliable and controllable reconstitute membrane proteins into artificially made model membranes...

Exact Symbol Error Probability of Cross-QAM in AWGN and Fading Channels

Directory of Open Access Journals (Sweden)

Zhang Xi-chun

2010-01-01

Full Text Available The exact symbol error probability (SEP performance of -ary cross quadrature amplitude modulation (QAM in additive white Gaussian noise (AWGN channel and fading channels, including Rayleigh, Nakagami-m, Rice, and Nakagami-q (Hoyt channels, is analyzed. The obtained closed-form SEP expressions contain a finite (in proportion to sum of single integrals with finite limits and an integrand composed of elementary (exponential, trigonometric, and/or power functions, thus readily enabling numerical evaluation. Particularly, Gaussian -function is a special case of these integrals and is included in the SEP expressions. Simple and very precise approximations, which contain only Gaussian -function for AWGN channel and contain three terms of the single integrals mentioned above for fading channels, respectively, are also given. The analytical expressions show excellent agreement with the simulation results, and numerical evaluation with the proposed expressions reveals that cross QAM can obtain at least 1.1 dB gain compared to rectangular QAM when SEP < 0.3 in all the considered channels.

Structure of a second crystal form of Bence-Jones protein Loc: Strikingly different domain associations in two crystal forms of a single protein

International Nuclear Information System (INIS)

Schiffer, M.; Ainsworth, C.; Xu, Z.B.; Carperos, W.; Olsen, K.; Solomon, A.; Stevens, F.J.; Chang, C.H.

1989-01-01

The authors have determined the structure of the immunoglobulin light-chain dimer Loc in a second crystal form that was grown from distilled water. The crystal structure was determined to 2.8-angstrom resolution; the R factor is 0.22. The two variable domains are related by local 2-fold axes and form an antigen binding pocket. The variable domain-variable domain interaction observed in this crystal form differs from the one exhibited by the protein when crystallized from ammonium sulfate in which the two variable domains formed a protrusion. The structure attained in the distilled water crystals is similar to, but not identical with, the one observed for the Mcg light-chain dimer in crystals grown from ammonium sulfate. Thus, two strikingly different structures were attained by this multisubunit protein in crystals grown under two different, commonly used, crystallization techniques. The quaternary interactions exhibited by the protein in the two crystal forms are sufficiently different to suggest fundamentally different interpretations of the structural basis for the function of this protein. This observation may have general implications regarding the use of single crystallographic determinations for detailed identification of structural and functional relationships. On the other hand, proteins whose structures can be altered by manipulation of crystallization conditions may provide useful systems for study of fundamental structural chemistry

Behavioural processes in marketing channel relationships: Review and integration of empirical evidence

DEFF Research Database (Denmark)

Jensen, Nils Bøgelund; Skytte, Hans

1997-01-01

This paper reviews the empirical research on behavioural processes in marketing channel relationships. Systematically examining nine international journals, we find 49 papers on behavioural processes. On the basis of the hypothesis tests in the papers, we discuss the results and integrate...

A ligand channel through the G protein coupled receptor opsin.

Directory of Open Access Journals (Sweden)

Peter W Hildebrand

Full Text Available The G protein coupled receptor rhodopsin contains a pocket within its seven-transmembrane helix (TM structure, which bears the inactivating 11-cis-retinal bound by a protonated Schiff-base to Lys296 in TM7. Light-induced 11-cis-/all-trans-isomerization leads to the Schiff-base deprotonated active Meta II intermediate. With Meta II decay, the Schiff-base bond is hydrolyzed, all-trans-retinal is released from the pocket, and the apoprotein opsin reloaded with new 11-cis-retinal. The crystal structure of opsin in its active Ops* conformation provides the basis for computational modeling of retinal release and uptake. The ligand-free 7TM bundle of opsin opens into the hydrophobic membrane layer through openings A (between TM1 and 7, and B (between TM5 and 6, respectively. Using skeleton search and molecular docking, we find a continuous channel through the protein that connects these two openings and comprises in its central part the retinal binding pocket. The channel traverses the receptor over a distance of ca. 70 A and is between 11.6 and 3.2 A wide. Both openings are lined with aromatic residues, while the central part is highly polar. Four constrictions within the channel are so narrow that they must stretch to allow passage of the retinal beta-ionone-ring. Constrictions are at openings A and B, respectively, and at Trp265 and Lys296 within the retinal pocket. The lysine enforces a 90 degrees elbow-like kink in the channel which limits retinal passage. With a favorable Lys side chain conformation, 11-cis-retinal can take the turn, whereas passage of the all-trans isomer would require more global conformational changes. We discuss possible scenarios for the uptake of 11-cis- and release of all-trans-retinal. If the uptake gate of 11-cis-retinal is assigned to opening B, all-trans is likely to leave through the same gate. The unidirectional passage proposed previously requires uptake of 11-cis-retinal through A and release of photolyzed all

A multi-channel integrated circuit for the readout of a microstrip gas chamber

Energy Technology Data Exchange (ETDEWEB)

Krummenacher, F.; Enz, C. (Smart Silicon Systems S.A., Lausanne (Switzerland)); Bellazzini, R. (Dipt. di Fisica, Pisa (Italy) INFN, Pisa (Italy))

1992-03-15

The design and test of an 8 channel integrated circuit for the readout of the microstrip gas chamber and other multielectrode detectors are described. The circuit is composed of 8 identical channels, each providing the amplification and the shaping of the signal delivered by the detector. The peaking time of the shaper is 25 ns and the overall amplifier gain is 8 mV/1000 e{sup -}. In addition to the analog output, each channel provides a TTL compatible digital output. The equivalent input noise is less than 700 e{sup -} rms and the total dc power consumption is about 5 mW/channel. To avoid a baseline shift due to the tail of the current issued from the detector, an adjustable pole-zero cancellation circuit has been included. (orig.).

KCNE regulation of K+ channel trafficking – a Sisyphean task?

Directory of Open Access Journals (Sweden)

Vikram Anmol Kanda

2012-06-01

Full Text Available Voltage-gated potassium (Kv channels shape the action potentials of excitable cells and regulate membrane potential and ion homeostasis in excitable and nonexcitable cells. With forty known members in the human genome and a variety of homomeric and heteromeric pore-forming alpha subunit interactions, post-translational modifications, cellular locations and expression patterns, the functional repertoire of the Kv alpha subunit family is monumental. This versatility is amplified by a host of interacting proteins, including the single membrane-spanning KCNE ancillary subunits. Here, examining both the secretory and the endocytic pathways, we review recent findings illustrating the surprising virtuosity of the KCNE proteins in orchestrating not just the function, but also the composition, diaspora and retrieval of channels formed by their Kv alpha subunit partners.

Pore size matters for potassium channel conductance

Science.gov (United States)

Moldenhauer, Hans; Pincuntureo, Matías

2016-01-01

Ion channels are membrane proteins that mediate efficient ion transport across the hydrophobic core of cell membranes, an unlikely process in their absence. K+ channels discriminate K+ over cations with similar radii with extraordinary selectivity and display a wide diversity of ion transport rates, covering differences of two orders of magnitude in unitary conductance. The pore domains of large- and small-conductance K+ channels share a general architectural design comprising a conserved narrow selectivity filter, which forms intimate interactions with permeant ions, flanked by two wider vestibules toward the internal and external openings. In large-conductance K+ channels, the inner vestibule is wide, whereas in small-conductance channels it is narrow. Here we raise the idea that the physical dimensions of the hydrophobic internal vestibule limit ion transport in K+ channels, accounting for their diversity in unitary conductance. PMID:27619418

Paramecium BBS genes are key to presence of channels in Cilia

Directory of Open Access Journals (Sweden)

Valentine Megan

2012-09-01

Full Text Available Abstract Background Changes in genes coding for ciliary proteins contribute to complex human syndromes called ciliopathies, such as Bardet-Biedl Syndrome (BBS. We used the model organism Paramecium to focus on ciliary ion channels that affect the beat form and sensory function of motile cilia and evaluate the effects of perturbing BBS proteins on these channels. Methods We used immunoprecipitations and mass spectrometry to explore whether Paramecium proteins interact as in mammalian cells. We used RNA interference (RNAi and swimming behavior assays to examine the effects of BBS depletion on ciliary ion channels that control ciliary beating. Combining RNA interference and epitope tagging, we examined the effects of BBS depletion of BBS 7, 8 and 9 on the location of three channels and a chemoreceptor in cilia. Results We found 10 orthologs of 8 BBS genes in P. tetraurelia. BBS1, 2, 4, 5, 7, 8 and 9 co-immunoprecipitate. While RNAi reduction of BBS 7 and 9 gene products caused loss and shortening of cilia, RNAi for all BBS genes except BBS2 affected patterns of ciliary motility that are governed by ciliary ion channels. Swimming behavior assays pointed to loss of ciliary K+ channel function. Combining RNAi and epitope tagged ciliary proteins we demonstrated that a calcium activated K+ channel was no longer located in the cilia upon depletion of BBS 7, 8 or 9, consistent with the cells’ swimming behavior. The TRPP channel PKD2 was also lost from the cilia. In contrast, the ciliary voltage gated calcium channel was unaffected by BBS depletion, consistent with behavioral assays. The ciliary location of a chemoreceptor for folate was similarly unperturbed by the depletion of BBS 7, 8 or 9. Conclusions The co-immunoprecipitation of BBS 1,2,4,5,7,8, and 9 suggests a complex of BBS proteins. RNAi for BBS 7, 8 or 9 gene products causes the selective loss of K+ and PKD2 channels from the cilia while the critical voltage gated calcium channel and a

Multi-scaled normal mode analysis method for dynamics simulation of protein-membrane complexes: A case study of potassium channel gating motion correlations

Energy Technology Data Exchange (ETDEWEB)

Wu, Xiaokun; Han, Min; Ming, Dengming, E-mail: dming@fudan.edu.cn [Department of Physiology and Biophysics, School of Life Sciences, Fudan University, Shanghai (China)

2015-10-07

Membrane proteins play critically important roles in many cellular activities such as ions and small molecule transportation, signal recognition, and transduction. In order to fulfill their functions, these proteins must be placed in different membrane environments and a variety of protein-lipid interactions may affect the behavior of these proteins. One of the key effects of protein-lipid interactions is their ability to change the dynamics status of membrane proteins, thus adjusting their functions. Here, we present a multi-scaled normal mode analysis (mNMA) method to study the dynamics perturbation to the membrane proteins imposed by lipid bi-layer membrane fluctuations. In mNMA, channel proteins are simulated at all-atom level while the membrane is described with a coarse-grained model. mNMA calculations clearly show that channel gating motion can tightly couple with a variety of membrane deformations, including bending and twisting. We then examined bi-channel systems where two channels were separated with different distances. From mNMA calculations, we observed both positive and negative gating correlations between two neighboring channels, and the correlation has a maximum as the channel center-to-center distance is close to 2.5 times of their diameter. This distance is larger than recently found maximum attraction distance between two proteins embedded in membrane which is 1.5 times of the protein size, indicating that membrane fluctuation might impose collective motions among proteins within a larger area. The hybrid resolution feature in mNMA provides atomic dynamics information for key components in the system without costing much computer resource. We expect it to be a conventional simulation tool for ordinary laboratories to study the dynamics of very complicated biological assemblies. The source code is available upon request to the authors.

Do TRPC channels support working memory? Comparing modulations of TRPC channels and working memory through G-protein coupled receptors and neuromodulators.

Science.gov (United States)

Reboreda, Antonio; Theissen, Frederik M; Valero-Aracama, Maria J; Arboit, Alberto; Corbu, Mihaela A; Yoshida, Motoharu

2018-03-01

Working memory is a crucial ability we use in daily life. However, the cellular mechanisms supporting working memory still remain largely unclear. A key component of working memory is persistent neural firing which is believed to serve short-term (hundreds of milliseconds up to tens of seconds) maintenance of necessary information. In this review, we will focus on the role of transient receptor potential canonical (TRPC) channels as a mechanism underlying persistent firing. Many years of in vitro work have been suggesting a crucial role of TRPC channels in working memory and temporal association tasks. If TRPC channels are indeed a central mechanism for working memory, manipulations which impair or facilitate working memory should have a similar effect on TRPC channel modulation. However, modulations of working memory and TRPC channels were never systematically compared, and it remains unanswered whether TRPC channels indeed contribute to working memory in vivo or not. In this article, we review the effects of G-protein coupled receptors (GPCR) and neuromodulators, including acetylcholine, noradrenalin, serotonin and dopamine, on working memory and TRPC channels. Based on comparisons, we argue that GPCR and downstream signaling pathways that activate TRPC, generally support working memory, while those that suppress TRPC channels impair it. However, depending on the channel types, areas, and systems tested, this is not the case in all studies. Further work to clarify involvement of specific TRPC channels in working memory tasks and how they are affected by neuromodulators is still necessary in the future. Copyright © 2018 Elsevier B.V. All rights reserved.

Inhibition of protein kinase A and GIRK channel reverses fentanyl-induced respiratory depression.

Science.gov (United States)

Liang, Xiaonan; Yong, Zheng; Su, Ruibin

2018-06-11

Opioid-induced respiratory depression is a major obstacle to improving the clinical management of moderate to severe chronic pain. Opioids inhibit neuronal activity via various pathways, including calcium channels, adenylyl cyclase, and potassium channels. Currently, the underlying molecular pathway of opioid-induced respiratory depression is only partially understood. This study aimed to investigate the mechanisms of opioid-induced respiratory depression in vivo by examining the effects of different pharmacological agents on fentanyl-induced respiratory depression. Respiratory parameters were detected using whole body plethysmography in conscious rats. We show that pre-treatment with the protein kinase A (PKA) inhibitor H89 reversed the fentanyl-related effects on respiratory rate, inspiratory time, and expiratory time. Pre-treatment with the G protein-gated inwardly rectifying potassium (GIRK) channel blocker Tertiapin-Q dose-dependently reversed the fentanyl-related effects on respiratory rate and inspiratory time. A phosphodiesterase 4 (PDE4) inhibitor and cyclic adenosine monophosphate (cAMP) analogs did not affect fentanyl-induced respiratory depression. These findings suggest that PKA and GIRK may be involved in fentanyl-induced respiratory depression and could represent useful therapeutic targets for the treatment of fentanyl-induced ventilatory depression. Copyright © 2018 Elsevier B.V. All rights reserved.

FPGA based, DSP integrated, 8-channel SIMCON, ver. 3.0. Initial results for 8-channel algorithm

Energy Technology Data Exchange (ETDEWEB)

Giergusiewicz, W.; Koprek, W.; Jalmuzna, W.; Pozniak, K.T.; Romaniuk, R.S. [Warsaw Univ. of Technology (Poland). Inst. of Electronic Systems

2005-07-01

The paper describes design, construction and initial measurements of an eight channel electronic LLRF device predicted for building of the control system for the VUV-FEL accelerator at DESY (Hamburg). The device, referred in the paper to as the SIMCON 3.0 (from the SC cavity simulator and controller) consists of a 16 layer, VME size, PCB, a large FPGA chip (VirtexII-4000 by Xilinx), eight fast ADCs and four DACs (by Analog Devices). To our knowledge, the proposed device is the first of this kind for the accelerator technology in which there was achieved (the FPGA based) DSP latency below 200 ns. With the optimized data transmission system, the overall LLRF system latency can be as low as 500 ns. The SIMCON 3.0 sub-system was applied for initial tests with the ACC1 module of the VUV FEL accelerator (eight channels) and with the CHECHIA test stand (single channel), both at the DESY. The promising results with the SIMCON 3.0. encouraged us to enter the design of SIMCON 3.1. possessing 10 measurement and control channels and some additional features to be reported in the next technical note. SIMCON 3.0. is a modular solution, while SIMCON 3.1. will be an integrated board of the all-in-one type. Two design approaches - modular and all-in-one, after branching off in this version of the Simcon, will be continued. (orig.)

Evidence for functional diversity between the voltage-gated proton channel Hv1 and its closest related protein HVRP1.

Directory of Open Access Journals (Sweden)

Iris H Kim

Full Text Available The Hv1 channel and voltage-sensitive phosphatases share with voltage-gated sodium, potassium, and calcium channels the ability to detect changes in membrane potential through voltage-sensing domains (VSDs. However, they lack the pore domain typical of these other channels. NaV, KV, and CaV proteins can be found in neurons and muscles, where they play important roles in electrical excitability. In contrast, VSD-containing proteins lacking a pore domain are found in non-excitable cells and are not involved in neuronal signaling. Here, we report the identification of HVRP1, a protein related to the Hv1 channel (from which the name Hv1 Related Protein 1 is derived, which we find to be expressed primarily in the central nervous system, and particularly in the cerebellum. Within the cerebellar tissue, HVRP1 is specifically expressed in granule neurons, as determined by in situ hybridization and immunohistochemistry. Analysis of subcellular distribution via electron microscopy and immunogold labeling reveals that the protein localizes on the post-synaptic side of contacts between glutamatergic mossy fibers and the granule cells. We also find that, despite the similarities in amino acid sequence and structural organization between Hv1 and HVRP1, the two proteins have distinct functional properties. The high conservation of HVRP1 in vertebrates and its cellular and subcellular localizations suggest an important function in the nervous system.

An evolutionarily conserved gene family encodes proton-selective ion channels.

Science.gov (United States)

Tu, Yu-Hsiang; Cooper, Alexander J; Teng, Bochuan; Chang, Rui B; Artiga, Daniel J; Turner, Heather N; Mulhall, Eric M; Ye, Wenlei; Smith, Andrew D; Liman, Emily R

2018-03-02

Ion channels form the basis for cellular electrical signaling. Despite the scores of genetically identified ion channels selective for other monatomic ions, only one type of proton-selective ion channel has been found in eukaryotic cells. By comparative transcriptome analysis of mouse taste receptor cells, we identified Otopetrin1 (OTOP1), a protein required for development of gravity-sensing otoconia in the vestibular system, as forming a proton-selective ion channel. We found that murine OTOP1 is enriched in acid-detecting taste receptor cells and is required for their zinc-sensitive proton conductance. Two related murine genes, Otop2 and Otop3 , and a Drosophila ortholog also encode proton channels. Evolutionary conservation of the gene family and its widespread tissue distribution suggest a broad role for proton channels in physiology and pathophysiology. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Estimation of Channel-Forming Discharge and Large-Event Geomorphic Response Using HEC-RAS

Science.gov (United States)

Hamilton, P.; Strom, K.; Hosseiny, S. M. H.

2015-12-01

The goal of the present work was to consider the functionality and applicability of HEC-RAS sediment transport simulations in two situations. The first was as a mode for obtaining quick estimates of the effective discharge, one measure of channel-forming discharge, and the second was as a mode to quickly estimate sediment transport and the commensurate potential erosion and deposition during large flood events. Though there are many other sediment transport and morphodynamic models available, e.g., CCHE1D, Nays2DH, we were interested in using HEC-RAS since this is the model of choice for many regulatory bodies, e.g., FEMA, cities, and counties. This makes using the sediment transport capability of HEC-RAS a natural extension of models that already otherwise exist and are well calibrated. In first looking at the utility of these models, we wanted to estimate the effective discharge of streams. Effective discharge is one way of defining the channel-forming discharge for a stream and is therefore an important parameter in natural channel design and restoration efforts. By running this range of floods, one can easily obtain an estimate for recurrence interval most responsible for moving the majority of sediment over a long time period. Results were compared to data collected within our research group on the Brazos River (TX). Effective discharge is an important estimate, particularly in understanding the equilibrium channel condition. Nevertheless, large floods are contemporaneously catastrophic and understanding their potential effects is desirable. Finally, we performed some sensitivity analysis to better understand the underlying assumptions of the various sediment transport model options and how they might affect the outcome of the aforementioned computations.

Investigating the role of viral integral membrane proteins in promoting the assembly of nepovirus and comovirus replication factories

Directory of Open Access Journals (Sweden)

Helene eSanfacon

2013-01-01

Full Text Available Formation of plant virus membrane-associated replication factories requires the association of viral replication proteins and viral RNA with intracellular membranes, the recruitment of host factors and the modification of membranes to form novel structures that house the replication complex. Many viruses encode integral membrane proteins that act as anchors for the replication complex. These hydrophobic proteins contain trans-membrane domains and/or amphipathic helices that associate with the membrane and modify its structure. The comovirus Co-Pro and NTP-binding (NTB, putative helicase proteins and the cognate nepovirus X2 and NTB proteins are among the best characterized plant virus integral membrane replication proteins and are functionally related to the picornavirus 2B, 2C and 3A membrane proteins. The identification of membrane-association domains and analysis of the membrane topology of these proteins is discussed. The evidence suggesting that these proteins have the ability to induce membrane proliferation, alter the structure and integrity of intracellular membranes and modulate the induction of symptoms in infected plants is also reviewed. Finally, areas of research that need further investigation are highlighted.

Molecular Evolution of Slow and Quick Anion Channels (SLACs and QUACs/ALMTs).

Science.gov (United States)

Dreyer, Ingo; Gomez-Porras, Judith Lucia; Riaño-Pachón, Diego Mauricio; Hedrich, Rainer; Geiger, Dietmar

2012-01-01

Electrophysiological analyses conducted about 25 years ago detected two types of anion channels in the plasma membrane of guard cells. One type of channel responds slowly to changes in membrane voltage while the other responds quickly. Consequently, they were named SLAC, for SLow Anion Channel, and QUAC, for QUick Anion Channel. Recently, genes SLAC1 and QUAC1/ALMT12, underlying the two different anion current components, could be identified in the model plant Arabidopsis thaliana. Expression of the gene products in Xenopus oocytes confirmed the quick and slow current kinetics. In this study we provide an overview on our current knowledge on slow and quick anion channels in plants and analyze the molecular evolution of ALMT/QUAC-like and SLAC-like channels. We discovered fingerprints that allow screening databases for these channel types and were able to identify 192 (177 non-redundant) SLAC-like and 422 (402 non-redundant) ALMT/QUAC-like proteins in the fully sequenced genomes of 32 plant species. Phylogenetic analyses provided new insights into the molecular evolution of these channel types. We also combined sequence alignment and clustering with predictions of protein features, leading to the identification of known conserved phosphorylation sites in SLAC1-like channels along with potential sites that have not been yet experimentally confirmed. Using a similar strategy to analyze the hydropathicity of ALMT/QUAC-like channels, we propose a modified topology with additional transmembrane regions that integrates structure and function of these membrane proteins. Our results suggest that cross-referencing phylogenetic analyses with position-specific protein properties and functional data could be a very powerful tool for genome research approaches in general.

Molecular evolution of slow and quick anion channels (SLACs and QUACs/ALMTs

Directory of Open Access Journals (Sweden)

Ingo eDreyer

2012-11-01

Full Text Available Electrophysiological analyses conducted about 25 years ago detected two types of anion channels in the plasma membrane of guard cells. One type of channel responds slowly to changes in membrane voltage while the other responds quickly. Consequently, they were named SLAC, for SLow Anion Channel, and QUAC, for QUick Anion Channel. Recently, genes SLAC1 and QUAC1/ALMT12, underlying the two different anion current components, could be identified in the model plant Arabidopsis thaliana. Expression of the gene products in Xenopus oocytes confirmed the quick and slow current kinetics. In this study we provide an overview on our current knowledge on slow and quick anion channels in plants and analyze the molecular evolution of ALMT/QUAC-like and SLAC-like channels. We discovered fingerprints that allow screening databases for these channel types and were able to identify 192 (177 non-redundant SLAC-like and 422 (402 non-redundant ALMT/QUAC-like proteins in the fully sequenced genomes of 32 plant species. Phylogenetic analyses provided new insights into the molecular evolution of these channel types. We also combined sequence alignment and clustering with predictions of protein features, leading to the identification of known conserved phosphorylation sites in SLAC1-like channels along with potential sites that have not been yet experimentally confirmed. Using a similar strategy to analyze the hydropathicity of ALMT/QUAC-like channels, we propose a modified topology with additional transmembrane regions that integrates structure and function of these membrane proteins. Our results suggest that cross-referencing phylogenetic analyses with position-specific protein properties and functional data could be a very powerful tool for genome research approaches in general.

Morphology of channels and channel-sand bodies in the Glauconitic sandstone member (Upper Mannville), Little Bow area, Alberta

Energy Technology Data Exchange (ETDEWEB)

Hopkins, J.C.; Hermanson, S.W.; Lawton, D.C.

1982-12-01

Large channels in the Glauconitic sandstone member of southern Alberta have proved to be difficult exploration targets because of an irregular distribution of reservoir sands within the channels. In the Little Bow area, two channels are present in the lower part of the Glauconitic member and have cut into the underlying Calcareous member. The channels can be recognized where they truncate regional markers of the Calcareous member -- the Bantry shale and Ostracod limestone, and where channel-fill deposits exhibit uniform geophysical log characters, indicating sandstone- or mudstone-filled reaches. Sediments of the Glauconitic member adjacent to the channels comprise a series of splay sandstones and mudstones that prograded into interdistributary bays of the lower delta plain. The two channels associated with these deposits are interpreted as distributary channels. Sand bodies within the channels formed as lateral bars and are isolated by mudstones which mark the abandoned stream course. Geological exploration for these discontinuous channel sandstones is difficult, and high-resolutio seismic data integrated with sound geologic modelling are critical for successful prospect delineation.

CLC-Nt1, a putative chloride channel protein of tobacco, co-localizes with mitochondrial membrane markers.

Science.gov (United States)

Lurin, C; Güclü, J; Cheniclet, C; Carde, J P; Barbier-Brygoo, H; Maurel, C

2000-06-01

The voltage-dependent chloride channel (CLC) family of membrane proteins has cognates in animals, yeast, bacteria and plants, and chloride-channel activity has been assigned to most of the animal homologues. Lack of evidence of CLC functions in plants prompted us to characterize the cellular localization of the tobacco CLC-Nt1 protein. Specific polyclonal antibodies were raised against an N-terminal polypeptide of CLC-Nt1. These antibodies were used to probe membrane proteins prepared by various cell-fractionation methods. These included aqueous two-phase partitioning (for plasma membranes), free-flow electrophoresis (for vacuolar and plasma membranes), intact vacuole isolation, Percoll-gradient centrifugation (for plastids and mitochondria) and stepped, linear, sucrose-density-gradient centrifugation (for mitochondria). Each purified membrane fraction was characterized with specific marker enzyme activities or antibodies. Our studies ruled out the possibility that the major cell localization of CLC-Nt1 was the vacuolar or plasma membranes, the endoplasmic reticulum, the Golgi apparatus or the plastids. In contrast, we showed that the tobacco CLC-Nt1 specifically co-localized with the markers of the mitochondrial inner membrane, cytochrome c oxidase and NAD9 protein. CLC-Nt1 may correspond to the inner membrane anion channel ('IMAC') described previously in animal and plant mitochondria.

Establishing homology between mitochondrial calcium uniporters, prokaryotic magnesium channels and chlamydial IncA proteins.

Science.gov (United States)

Lee, Andre; Vastermark, Ake; Saier, Milton H

2014-08-01

Mitochondrial calcium uniporters (MCUs) (TC no. 1.A.77) are oligomeric channel proteins found in the mitochondrial inner membrane. MCUs have two well-conserved transmembrane segments (TMSs), connected by a linker, similar to bacterial MCU homologues. These proteins and chlamydial IncA proteins (of unknown function; TC no. 9.B.159) are homologous to prokaryotic Mg(2+) transporters, AtpI and AtpZ, based on comparison scores of up to 14.5 sds. A phylogenetic tree containing all of these proteins showed that the AtpZ proteins cluster coherently as a subset within the large and diverse AtpI cluster, which branches separately from the MCUs and IncAs, both of which cluster coherently. The MCUs and AtpZs share the same two TMS topology, but the AtpIs have four TMSs, and IncAs can have either two (most frequent) or four (less frequent) TMSs. Binary alignments, comparison scores and motif analyses showed that TMSs 1 and 2 align with TMSs 3 and 4 of the AtpIs, suggesting that the four TMS AtpI proteins arose via an intragenic duplication event. These findings establish an evolutionary link interconnecting eukaryotic and prokaryotic Ca(2+) and Mg(2+) transporters with chlamydial IncAs, and lead us to suggest that all members of the MCU superfamily, including IncAs, function as divalent cation channels. © 2014 The Authors.

Neighborhood Walkable Urban Form and C-Reactive Protein

Science.gov (United States)

Background: Walkable urban form predicts physical activity and lower body mass index, which lower C-reactive protein (CRP). However, urban form is also related to pollution, noise, social and health behavior, crowding, and other stressors, which may complement or contravene walka...

Integration of relational and hierarchical network information for protein function prediction

Directory of Open Access Journals (Sweden)

Jiang Xiaoyu

2008-08-01

Full Text Available Abstract Background In the current climate of high-throughput computational biology, the inference of a protein's function from related measurements, such as protein-protein interaction relations, has become a canonical task. Most existing technologies pursue this task as a classification problem, on a term-by-term basis, for each term in a database, such as the Gene Ontology (GO database, a popular rigorous vocabulary for biological functions. However, ontology structures are essentially hierarchies, with certain top to bottom annotation rules which protein function predictions should in principle follow. Currently, the most common approach to imposing these hierarchical constraints on network-based classifiers is through the use of transitive closure to predictions. Results We propose a probabilistic framework to integrate information in relational data, in the form of a protein-protein interaction network, and a hierarchically structured database of terms, in the form of the GO database, for the purpose of protein function prediction. At the heart of our framework is a factorization of local neighborhood information in the protein-protein interaction network across successive ancestral terms in the GO hierarchy. We introduce a classifier within this framework, with computationally efficient implementation, that produces GO-term predictions that naturally obey a hierarchical 'true-path' consistency from root to leaves, without the need for further post-processing. Conclusion A cross-validation study, using data from the yeast Saccharomyces cerevisiae, shows our method offers substantial improvements over both standard 'guilt-by-association' (i.e., Nearest-Neighbor and more refined Markov random field methods, whether in their original form or when post-processed to artificially impose 'true-path' consistency. Further analysis of the results indicates that these improvements are associated with increased predictive capabilities (i.e., increased

Master integrals for the four-loop Sudakov form factor

Directory of Open Access Journals (Sweden)

Rutger H. Boels

2016-01-01

Full Text Available The light-like cusp anomalous dimension is a universal function in the analysis of infrared divergences. In maximally (N=4 supersymmetric Yang–Mills theory (SYM in the planar limit, it is known, in principle, to all loop orders. The non-planar corrections are not known in any theory, with the first appearing at the four-loop order. The simplest quantity which contains this correction is the four-loop two-point form factor of the stress tensor multiplet. This form factor was largely obtained in integrand form in a previous work for N=4 SYM, up to a free parameter. In this work, a reduction of the appearing integrals obtained by solving integration-by-parts (IBP identities using a modified version of Reduze is reported. The form factor is shown to be independent of the remaining parameter at integrand level due to an intricate pattern of cancellations after IBP reduction. Moreover, two of the integral topologies vanish after reduction. The appearing master integrals are cross-checked using independent algebraic-geometry techniques explored in the Mint package. The latter results provide the basis of master integrals applicable to generic form factors, including those in Quantum Chromodynamics. Discrepancies between explicitly solving the IBP relations and the MINT approach are highlighted. Remaining bottlenecks to completing the computation of the four-loop non-planar cusp anomalous dimension in N=4 SYM and beyond are identified.

Master integrals for the four-loop Sudakov form factor

International Nuclear Information System (INIS)

Boels, Rutger; Kniehl, Bernd A.; Yang, Gang; Chinese Academy of Sciences, Beijing

2015-08-01

The light-like cusp anomalous dimension is a universal function in the analysis of infrared divergences. In maximally (N=4) supersymmetric Yang-Mills theory (SYM) in the planar limit, it is known, in principle, to all loop orders. The non-planar corrections are not known in any theory, with the first appearing at the four-loop order. Probably the simplest quantity which contains this correction is the four-loop two-point form factor of the stress tensor multiplet. This form factor was obtained in integrand form in a previous work for N=4 SYM, up to a single parameter. In this work, a reduction of the appearing integrals obtained by solving integration-by-parts (IBP) identities using (a tweaked version of) Reduze is reported. The form factor is shown to be independent of the remaining parameter at integrand level due to an intricate pattern of cancellations after IBP reduction. The appearing master integrals are cross-checked using algebraic techniques explored in the Mint package. The latter results provide the basis of master integrals applicable to generic form factors, including those in Quantum Chromodynamics. Remaining bottlenecks to completing the computation of the four-loop non-planar cusp anomalous dimension in N=4 SYM and beyond are identified.

Chaperone-Mediated Sec61 Channel Gating during ER Import of Small Precursor Proteins Overcomes Sec61 Inhibitor-Reinforced Energy Barrier

Directory of Open Access Journals (Sweden)

Sarah Haßdenteufel

2018-05-01

Full Text Available Summary: Protein transport into the mammalian endoplasmic reticulum (ER is mediated by the heterotrimeric Sec61 channel. The signal recognition particle (SRP and TRC systems and Sec62 have all been characterized as membrane-targeting components for small presecretory proteins, whereas Sec63 and the lumenal chaperone BiP act as auxiliary translocation components. Here, we report the transport requirements of two natural, small presecretory proteins and engineered variants using semipermeabilized human cells after the depletion of specific ER components. Our results suggest that hSnd2, Sec62, and SRP and TRC receptor each provide alternative targeting pathways for short secretory proteins and define rules of engagement for the actions of Sec63 and BiP during their membrane translocation. We find that the Sec62/Sec63 complex plus BiP can facilitate Sec61 channel opening, thereby allowing precursors that have weak signal peptides or other inhibitory features to translocate. A Sec61 inhibitor can mimic the effect of BiP depletion on Sec61 gating, suggesting that they both act at the same essential membrane translocation step. : Protein transport into the human endoplasmic reticulum (ER is mediated by the heterotrimeric Sec61 channel. Haßdenteufel et al. map the determinants for requirement of different targeting pathways and different auxiliary components of the Sec61 channel in ER import of short presecretory proteins. Different characteristics of precursor polypeptides dictate the engagement of each component. Keywords: endoplasmic reticulum, protein targeting and translocation, Sec61 channel gating, Sec62, Sec63, BiP, CAM741, signal peptide, mature region, cluster of positive charges

Physiology and pathophysiology of ClC-K/barttin channels.

Science.gov (United States)

Fahlke, Christoph; Fischer, Martin

2010-01-01

ClC-K channels form a subgroup of anion channels within the ClC family of anion transport proteins. They are expressed predominantly in the kidney and in the inner ear, and are necessary for NaCl resorption in the loop of Henle and for K+ secretion by the stria vascularis. Subcellular distribution as well as the function of these channels are tightly regulated by an accessory subunit, barttin. Barttin improves the stability of ClC-K channel protein, stimulates the exit from the endoplasmic reticulum and insertion into the plasma membrane and changes its function by modifying voltage-dependent gating processes. The importance of ClC-K/barttin channels is highlighted by several genetic diseases. Dysfunctions of ClC-K channels result in Bartter syndrome, an inherited human condition characterized by impaired urinary concentration. Mutations in the gene encoding barttin, BSND, affect the urinary concentration as well as the sensory function of the inner ear. Surprisingly, there is one BSND mutation that causes deafness without affecting renal function, indicating that kidney function tolerates a reduction of anion channel activity that is not sufficient to support normal signal transduction in inner hair cells. This review summarizes recent work on molecular mechanisms, physiology, and pathophysiology of ClC-K/barttin channels.

Physiology and pathophysiology of ClC-K/barttin channels

Directory of Open Access Journals (Sweden)

Christoph eFahlke

2010-11-01

Full Text Available ClC-K channels form a subgroup of anion channels within the ClC family of anion transport proteins. They are expressed predominantly in the kidney and in the inner ear, and are necessary for NaCl resorption in the loop of Henle and for K+ secretion by the stria vascularis. Subcellular distribution as well as the function of these channels are tightly regulated by an accessory subunit, barttin. Barttin improves the stability of ClC-K channel protein, stimulates the exit from the endoplasmic reticulum and insertion into the plasma membrane and changes its function by modifying voltage-dependent gating processes. The importance of ClC-K/barttin channels is highlighted by several genetic diseases. Dysfunctions of ClC-K channels result in Bartter syndrome, an inherited human condition characterized by impaired urinary concentration. Mutations in the gene encoding barttin, BSND, affect the urinary concentration as well as the sensory function of the inner ear. Surprisingly, there is one BSND mutation that causes deafness without affecting renal function, indicating that kidney function tolerates a reduction of anion channel activity that is not sufficient to support normal signal transduction in inner hair cells. This review summarizes recent work on molecular mechanisms, physiology and pathophysiology of ClC-K/barttin channels.

A family of fluoride-specific ion channels with dual-topology architecture.

Science.gov (United States)

Stockbridge, Randy B; Robertson, Janice L; Kolmakova-Partensky, Ludmila; Miller, Christopher

2013-08-27

Fluoride ion, ubiquitous in soil, water, and marine environments, is a chronic threat to microorganisms. Many prokaryotes, archea, unicellular eukaryotes, and plants use a recently discovered family of F(-) exporter proteins to lower cytoplasmic F(-) levels to counteract the anion's toxicity. We show here that these 'Fluc' proteins, purified and reconstituted in liposomes and planar phospholipid bilayers, form constitutively open anion channels with extreme selectivity for F(-) over Cl(-). The active channel is a dimer of identical or homologous subunits arranged in antiparallel transmembrane orientation. This dual-topology assembly has not previously been seen in ion channels but is known in multidrug transporters of the SMR family, and is suggestive of an evolutionary antecedent of the inverted repeats found within the subunits of many membrane transport proteins. DOI:http://dx.doi.org/10.7554/eLife.01084.001.

Multi-channel normal speed gated integrator in the measurement of the laser scattering light energy

International Nuclear Information System (INIS)

Yang Dong; Yu Xiaoqi; Hu Yuanfeng

2005-01-01

With the method of integration in a limited time, a Multi-channel normal speed gated integrator based on VXI system has been developed for measuring the signals with changeable pulse width in laser scattering light experiment. It has been tested with signal sources in ICF experiment. In tests, the integral nonlinearity between the integral results of the gated integrator and that of an oscilloscope is less than 1%. In the ICF experiments the maximum error between the integral results of the gated integrator and that of oscilloscope is less than 3% of the full scale range of the gated integrator. (authors)

Transient bedrock channel evolution across a precipitation gradient: A case study from Kohala, Hawaii.

Science.gov (United States)

Gasparini, N. M.; Han, J.; Johnson, J. P.; Menking, J. A.

2011-12-01

integrative, relatively extreme precipitation gradients are required to produce anomalous channel profile forms.

Proton-beam writing channel based on an electrostatic accelerator

Science.gov (United States)

Lapin, A. S.; Rebrov, V. A.; Kolin'ko, S. V.; Salivon, V. F.; Ponomarev, A. G.

2016-09-01

We have described the structure of the proton-beam writing channel as a continuation of a nuclear scanning microprobe channel. The problem of the accuracy of positioning a probe by constructing a new high-frequency electrostatic scanning system has been solved. Special attention has been paid to designing the probe-forming system and its various configurations have been considered. The probe-forming system that best corresponds to the conditions of the lithographic process has been found based on solving the problem of optimizing proton beam formation. A system for controlling beam scanning using multifunctional module of integrated programmable logic systems has been developed.

Expression patterns of ion channels and structural proteins in a multimodal cell type of the avian optic tectum.

Science.gov (United States)

Lischka, Katharina; Ladel, Simone; Luksch, Harald; Weigel, Stefan

2018-02-15

The midbrain is an important subcortical area involved in distinct functions such as multimodal integration, movement initiation, bottom-up, and top-down attention. Our group is particularly interested in cellular computation of multisensory integration. We focus on the visual part of the avian midbrain, the optic tectum (TeO, counterpart to mammalian superior colliculus). This area has a layered structure with the great advantage of distinct input and output regions. In chicken, the TeO is organized in 15 layers where visual input targets the superficial layers while auditory input terminates in deeper layers. One specific cell type, the Shepherd's crook neuron (SCN), extends dendrites in both input regions. The characteristic feature of these neurons is the axon origin at the apical dendrite. The molecular identity of this characteristic region and thus, the site of action potential generation are of particular importance to understand signal flow and cellular computation in this neuron. We present immunohistochemical data of structural proteins (NF200, Ankyrin G, and Myelin) and ion channels (Pan-Na v , Na v 1.6, and K v 3.1b). NF200 is strongly expressed in the axon. Ankyrin G is mainly expressed at the axon initial segment (AIS). Myelination starts after the AIS as well as the distribution of Na v channels on the axon. The subtype Na v 1.6 has a high density in this region. K v 3.1b is restricted to the soma, the primary neurite and the axon branch. The distribution of functional molecules in SCNs provides insight into the information flow and the integration of sensory modalities in the TeO of the avian midbrain. © 2017 Wiley Periodicals, Inc.

The 128-channel fully differential digital integrated neural recording and stimulation interface.

Science.gov (United States)

Shahrokhi, Farzaneh; Abdelhalim, Karim; Serletis, Demitre; Carlen, Peter L; Genov, Roman

2010-06-01

We present a fully differential 128-channel integrated neural interface. It consists of an array of 8 X 16 low-power low-noise signal-recording and generation circuits for electrical neural activity monitoring and stimulation, respectively. The recording channel has two stages of signal amplification and conditioning with and a fully differential 8-b column-parallel successive approximation (SAR) analog-to-digital converter (ADC). The total measured power consumption of each recording channel, including the SAR ADC, is 15.5 ¿W. The measured input-referred noise is 6.08 ¿ Vrms over a 5-kHz bandwidth, resulting in a noise efficiency factor of 5.6. The stimulation channel performs monophasic or biphasic voltage-mode stimulation, with a maximum stimulation current of 5 mA and a quiescent power dissipation of 51.5 ¿W. The design is implemented in 0.35-¿m complementary metal-oxide semiconductor technology with the channel pitch of 200 ¿m for a total die size of 3.4 mm × 2.5 mm and a total power consumption of 9.33 mW. The neural interface was validated in in vitro recording of a low-Mg(2+)/high-K(+) epileptic seizure model in an intact hippocampus of a mouse.

Use of integrity control and automatic start of reserve in a multi-channel temperature and flow rate control device

International Nuclear Information System (INIS)

Strzalkowski, L.

1975-01-01

A way to increase reliability of process quantity control is control of the integrity of the control plants themselves. The possibilities of integrity control on control devices having simply duplicated control channels or working on the basis of the ''two-from-three'' principle are valued. A highly reliable integrity control is possible by use of test signals. For an appropriate control device, structure and function of the assemblies are described. The integrity control device may be used in the water coolant temperature and flow rate control system for all technological channels of the research reactor ''Maria''

Fluvial Channel Networks as Analogs for the Ridge-Forming Unit, Sinus Meridiani, Mars

Science.gov (United States)

Wilkinson, M. J.; du Bois, J. B.

2010-01-01

Fluvial models have been generally discounted as analogs for the younger layered rock units of Sinus Meridiani. A fluvial model based on the large fluvial fan provides a possibly close analog for various features of the sinuous ridges of the etched, ridge-forming unit (RFU) in particular. The close spacing of the RFU ridges, their apparently chaotic orientations, and their organization in dense networks all appear unlike classical stream channel patterns. However, drainage patterns on large fluvial fans low-angle, fluvial aggradational features, 100s of km long, documented worldwide by us provide parallels. Some large fan characteristics resemble those of classical floodplains, but many differences have been demonstrated. One major distinction relevant to the RFU is that channel landscapes of large fans can dominate large areas (1.2 million km2 in one S. American study area). We compare channel morphologies on large fans in the southern Sahara Desert with ridge patterns in Sinus Meridiani (fig 1). Stream channels are the dominant landform on large terrestrial fans: they may equate to the ubiquitous, sinuous, elongated ridges of the RFU that cover areas region wide. Networks of convergent/divergent and crossing channels may equate to similar features in the ridge networks. Downslope divergence is absent in channels of terrestrial upland erosional landscapes (fig. 1, left), whereas it is common to both large fans (fig. 1, center) and RFU ridge patterns (fig 1, right downslope defined as the regional NW slope of Sinus Meridiani). RFU ridge orientation, judged from those areas apparently devoid of impact crater control, is broadly parallel with the regional slope (arrow, fig. 1, right), as is mean orientation of major channels on large fans (arrow, fig. 1, center). High densities per unit area characterize fan channels and martian ridges reaching an order of magnitude higher than those in uplands just upstream of the terrestrial study areas fig. 1. In concert with

An improved integrally formed radio frequency quadrupole

Science.gov (United States)

Abbott, S.R.

1987-10-05

An improved radio frequency quadrupole is provided having an elongate housing with an elongate central axis and top, bottom and two side walls symmetrically disposed about the axis, and vanes formed integrally with the walls, the vanes each having a cross-section at right angles to the central axis which tapers inwardly toward the axis to form electrode tips spaced from each other by predetermined distances. Each of the four walls, and the vanes integral therewith, is a separate structural element having a central lengthwise plane passing through the tip of the vane, the walls having flat mounting surfaces at right angles to and parallel to the control plane, respectively, which are butted together to position the walls and vane tips relative to each other. 4 figs.

Channel belt architecture formed by a meandering river

NARCIS (Netherlands)

Lageweg, W.I. van de; Dijk, W.M. van; Kleinhans, M.G.

2013-01-01

Stratification in channel belts is the key to reconstructing formative channel dimensions and palaeoflow conditions; this requires an understanding of the relation between river morphodynamics and set thickness. So far, theories for reconstruction of the original morphology from preserved

Near-optimal integration of facial form and motion.

Science.gov (United States)

Dobs, Katharina; Ma, Wei Ji; Reddy, Leila

2017-09-08

Human perception consists of the continuous integration of sensory cues pertaining to the same object. While it has been fairly well shown that humans use an optimal strategy when integrating low-level cues proportional to their relative reliability, the integration processes underlying high-level perception are much less understood. Here we investigate cue integration in a complex high-level perceptual system, the human face processing system. We tested cue integration of facial form and motion in an identity categorization task and found that an optimal model could successfully predict subjects' identity choices. Our results suggest that optimal cue integration may be implemented across different levels of the visual processing hierarchy.

Sea Turtles Geolocalization in the Indian Ocean: An Over Sea Radio Channel framework integrating a trilateration technique

Science.gov (United States)

Guegan, Loic; Murad, Nour Mohammad; Bonhommeau, Sylvain

2018-03-01

This paper deals with the modeling of the over sea radio channel and aims to establish sea turtles localization off the coast of Reunion Island, and also on Europa Island in the Mozambique Channel. In order to model this radio channel, a framework measurement protocol is proposed. The over sea measured channel is integrated to the localization algorithm to estimate the turtle trajectory based on Power of Arrival (PoA) technique compared to GPS localization. Moreover, cross correlation tool is used to characterize the over sea propagation channel. First measurement of the radio channel on the Reunion Island coast combine to the POA algorithm show an error of 18 m for 45% of the approximated points.

Experiments on laser driven beatwave acceleration in a ponderomotively formed plasma channel

International Nuclear Information System (INIS)

Tochitsky, S.Ya.; Narang, R.; Filip, C.V.; Clayton, C.E.; Marsh, K.A.; Joshi, C.; Musumeci, P.; Yoder, R.B.; Rosenzweig, J.B.; Pellegrini, C.

2004-01-01

A 10 ps long beam of 12 MeV electrons is externally injected into a ∼3-cm long plasma beatwave excited in a laser ionized hydrogen gas. The electrons have been accelerated to 50 MeV with a gradient of ∼1.3 GeV/m. It is shown that when the effective plasma wave amplitude-length product is limited by ionization-induced defocusing (IID), acceleration of electrons is significantly enhanced by using a laser pulse with a duration longer than the time required for ions to move across the laser spot size. Both experiments and two-dimensional simulations reveal that, in this case, self-guiding of the laser pulse in a ponderomotively formed plasma channel occurs. This compensates for IID and drives the beatwave over the longer length compared to when such a channel is not present

Effect of ceramic membrane channel geometry and uniform transmembrane pressure on limiting flux and serum protein removal during skim milk microfiltration.

Science.gov (United States)

Adams, Michael C; Hurt, Emily E; Barbano, David M

2015-11-01

Our objectives were to determine the effects of a ceramic microfiltration (MF) membrane's retentate flow channel geometry (round or diamond-shaped) and uniform transmembrane pressure (UTP) on limiting flux (LF) and serum protein (SP) removal during skim milk MF at a temperature of 50°C, a retentate protein concentration of 8.5%, and an average cross-flow velocity of 7 m·s(-1). Performance of membranes with round and diamond flow channels was compared in UTP mode. Performance of the membrane with round flow channels was compared with and without UTP. Using UTP with round flow channel MF membranes increased the LF by 5% when compared with not using UTP, but SP removal was not affected by the use of UTP. Using membranes with round channels instead of diamond-shaped channels in UTP mode increased the LF by 24%. This increase was associated with a 25% increase in Reynolds number and can be explained by lower shear at the vertices of the diamond-shaped channel's surface. The SP removal factor of the diamond channel system was higher than the SP removal factor of the round channel system below the LF. However, the diamond channel system passed more casein into the MF permeate than the round channel system. Because only one batch of each membrane was tested in our study, it was not possible to determine if the differences in protein rejection between channel geometries were due to the membrane design or random manufacturing variation. Despite the lower LF of the diamond channel system, the 47% increase in membrane module surface area of the diamond channel system produced a modular permeate removal rate that was at least 19% higher than the round channel system. Consequently, using diamond channel membranes instead of round channel membranes could reduce some of the costs associated with ceramic MF of skim milk if fewer membrane modules could be used to attain the required membrane area. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All

Underestimated Halogen Bonds Forming with Protein Backbone in Protein Data Bank.

Science.gov (United States)

Zhang, Qian; Xu, Zhijian; Shi, Jiye; Zhu, Weiliang

2017-07-24

Halogen bonds (XBs) are attracting increasing attention in biological systems. Protein Data Bank (PDB) archives experimentally determined XBs in biological macromolecules. However, no software for structure refinement in X-ray crystallography takes into account XBs, which might result in the weakening or even vanishing of experimentally determined XBs in PDB. In our previous study, we showed that side-chain XBs forming with protein side chains are underestimated in PDB on the basis of the phenomenon that the proportion of side-chain XBs to overall XBs decreases as structural resolution becomes lower and lower. However, whether the dominant backbone XBs forming with protein backbone are overlooked is still a mystery. Here, with the help of the ratio (R F ) of the observed XBs' frequency of occurrence to their frequency expected at random, we demonstrated that backbone XBs are largely overlooked in PDB, too. Furthermore, three cases were discovered possessing backbone XBs in high resolution structures while losing the XBs in low resolution structures. In the last two cases, even at 1.80 Å resolution, the backbone XBs were lost, manifesting the urgent need to consider XBs in the refinement process during X-ray crystallography study.

A Closed Form Solution for an Unorthodox Trigonometric Integral

Science.gov (United States)

Wu, Yan

2009-01-01

A closed form solution for the trigonometric integral [integral]sec[superscript 2k+1]xdx, k=0,1,2,..., is presented in this article. The result will fill the gap in another trigonometric integral [integral]sec[superscript 2m+1] x tan[superscript 2n]xdx, which is neglected by most of the calculus textbooks due to its foreseeable unorthodox solution…

RBMK fuel channel integrity. A publication of the extrabudgetary programme on the safety of WWER and RBMK nuclear power plants

International Nuclear Information System (INIS)

1999-01-01

The fuel channel integrity in the RBMK NPPs is an issue of high safety concern. To date, three single fuel channel ruptures have occurred. Fuel channel rupture results in release of radioactivity to the reactor cavity and may lead to a release of radioactivity to the environment if the confinement safety system does not function properly. A multiple fuel channel rupture exceeding the venting capacity of the reactor cavity overpressure protection system poses a major impact on the plant safety. Further, due to incorrect prediction at the design stage the gas gap between the fuel channel pressure tube and the graphite blocks closes after approximately 17 years of plant operation. There is no safety justification available for the continued plant operation in this condition and the reactors are being retubed to avoid operation in this out of design condition, which may have negative impact on the fuel channel integrity. The loss of the mechanical integrity of fuel channel pressure tubes is a major safety concern for RBMK reactors since it may lead to overpressurization of the reactor cavity and consequently develop into a severe accident. In this report, information on the main design features of the RBMK reactor related to the fuel channel integrity is given. Further, detailed information on the fuel channel pressure tube and the graphite blocks with respect to their design, manufacture, in-service inspection, operating experience, ageing behaviour including degradation mechanisms is discussed in detail. The behaviour of the system fuel channel-graphite core including the corrective actions developed and implemented is discussed. Both normal operating conditions and accident conditions are addressed, considering also the gas gap closure process and its impact. The report also covers the fuel channel ducts. It is concluded in the report that for RBMK-1000 reactors and the adopted retubing strategy, limited local gas gap closure occurs at the time of pressure tube

Some closed form expressions for the generalized secant integrals

International Nuclear Information System (INIS)

Nadarajah, Saralees

2007-01-01

The generalized secant integrals of the form I a (ψ,b)=b a ∫ 0 ψ exp(-bsecθ)(secθ) a dθ for b>0 and 0 a (ψ,b) have been known except when both a and ψ are fixed. In this note, we provide several closed form expressions for I a (ψ,b) applicable for a wide range of values of a and b. We establish their numerical accuracy over the methods presented in Michieli [1998. Point kernel calculations of dose fields from line sources using expanded polynomial form of buildup factor data: generalized secant integral series representations. Radiat. Phys. Chem. 51, 121-128; 2001. Some properties of generalized secant integrals: extended definition and recurrence relations. Radiat. Phys. Chem. 60, 551-554]. Finally, a simple computer program is provided for I a (ψ,b) that could be used widely

Structure of synaptophysin: a hexameric MARVEL-domain channel protein.

Science.gov (United States)

Arthur, Christopher P; Stowell, Michael H B

2007-06-01

Synaptophysin I (SypI) is an archetypal member of the MARVEL-domain family of integral membrane proteins and one of the first synaptic vesicle proteins to be identified and cloned. Most all MARVEL-domain proteins are involved in membrane apposition and vesicle-trafficking events, but their precise role in these processes is unclear. We have purified mammalian SypI and determined its three-dimensional (3D) structure by using electron microscopy and single-particle 3D reconstruction. The hexameric structure resembles an open basket with a large pore and tenuous interactions within the cytosolic domain. The structure suggests a model for Synaptophysin's role in fusion and recycling that is regulated by known interactions with the SNARE machinery. This 3D structure of a MARVEL-domain protein provides a structural foundation for understanding the role of these important proteins in a variety of biological processes.

Integrable couplings of relativistic Toda lattice systems in polynomial form and rational form, their hierarchies and bi-Hamiltonian structures

Energy Technology Data Exchange (ETDEWEB)

Xu Xixiang [College of Science, Shandong University of Science and Technology, Qingdao 266510 (China)], E-mail: xixiang_xu@yahoo.com.cn

2009-10-02

Integrable couplings of relativistic Toda lattice systems in polynomial form and rational form, and their hierarchies, are derived from a four-by-four discrete matrix eigenvalue problem. The bi-Hamiltonian structure for every integrable coupling in the two hierarchies obtained is established by means of the discrete variational identity. Ultimately, Liouvolle integrability of the obtained integrable couplings is demonstrated.

A channel-by-channel method of reducing the errors associated with peak area integration

International Nuclear Information System (INIS)

Luedeke, T.P.; Tripard, G.E.

1996-01-01

A new method of reducing the errors associated with peak area integration has been developed. This method utilizes the signal content of each channel as an estimate of the overall peak area. These individual estimates can then be weighted according to the precision with which each estimate is known, producing an overall area estimate. Experimental measurements were performed on a small peak sitting on a large background, and the results compared to those obtained from a commercial software program. Results showed a marked decrease in the spread of results around the true value (obtained by counting for a long period of time), and a reduction in the statistical uncertainty associated with the peak area. (orig.)

Modeling Oligo-Miocence channel sands (Dezful Embayment, SW Iran): an integrated facies classification workflow

International Nuclear Information System (INIS)

Heydari, Mostafa; Maddahi, Iradj; Moradpour, Mehran; Esmaeilpour, Sajjad

2014-01-01

This study has been conducted on Mansuri onshore oilfield located in Dezful Embayment, south-west Iran. One of the hydrocarbon-bearing formations is a Oligo-Miocene Asmari formation—the most prolific Iranian reservoir rock. Like many other oilfields in the area, the trap in this field is deemed structural (anticline), formed during the collision of the Arabian plate with the Iranian plate and the folding of Neotethys deposits with a NW–SE trend. This study integrates three different quantitative studies from geology, geophysics and petrophysics disciplines to quantitate ‘the qualitative study of seismic facies analysis based on trace shapes and 3D multi-attribute clustering’. First, stratigraphic sequences and seismic detectable facies were derived at one well location using the available high resolution core facies analysis and depositional environment assessment reports. Paleo and petrophysical logs from other wells were subsequently used for the extrapolation of stratigraphic sequences interpreted at the first well. Utilizing lithology discrimination obtained by wire-line log interpretation, facies were extrapolated to all wells in the area. Seismic 3D attribute analysis and seismic facies classification established a 3D facies volume accordingly, which was finally calibrated to geological facies at well locations. The ultimate extracted facies-guided geobody shows that good reservoir-quality channel sands have accumulated with NW/SE elongation at the ridge of the structure. As a result, this type of geometry has created a stratigraphic/structural hydrocarbon trap in this oilfield. Moreover, seismic facies analysis shows that buried channels do not parallel the predominant Arabian plate-originated channels (with SW–NE trends) in SW Zagros and are locally swerved in this area. (paper)

Biological Membrane Ion Channels Dynamics, Structure, and Applications

CERN Document Server

Chung, Shin-Ho; Krishnamurthy, Vikram

2007-01-01

Ion channels are biological nanotubes that are formed by membrane proteins. Because ion channels regulate all electrical activities in living cells, understanding their mechanisms at a molecular level is a fundamental problem in biology. This book deals with recent breakthroughs in ion-channel research that have been brought about by the combined effort of experimental biophysicists and computational physicists, who together are beginning to unravel the story of these exquisitely designed biomolecules. With chapters by leading experts, the book is aimed at researchers in nanodevices and biosensors, as well as advanced undergraduate and graduate students in biology and the physical sciences. Key Features Presents the latest information on the molecular mechanisms of ion permeation through membrane ion channels Uses schematic diagrams to illustrate important concepts in biophysics Written by leading researchers in the area of ion channel investigations

Single channel recording of a mitochondrial calcium uniporter.

Science.gov (United States)

Wu, Guangyan; Li, Shunjin; Zong, Guangning; Liu, Xiaofen; Fei, Shuang; Shen, Linda; Guan, Xiangchen; Yang, Xue; Shen, Yuequan

2018-01-29

Mitochondrial calcium uniporter (MCU) is the pore-forming subunit of the entire uniporter complex and plays an important role in mitochondrial calcium uptake. However, the single channel recording of MCU remains controversial. Here, we expressed and purified different MCU proteins and then reconstituted them into planar lipid bilayers for single channel recording. We showed that MCU alone from Pyronema omphalodes (pMCU) is active with prominent single channel Ca 2+ currents. In sharp contrast, MCU alone from Homo sapiens (hMCU) is inactive. The essential MCU regulator (EMRE) activates hMCU, and therefore, the complex (hMCU-hEMRE) shows prominent single channel Ca 2+ currents. These single channel currents are sensitive to the specific MCU inhibitor Ruthenium Red. Our results clearly demonstrate that active MCU can conduct large amounts of calcium into the mitochondria. Copyright © 2018 Elsevier Inc. All rights reserved.

A Suboptimal Scheme for Multi-User Scheduling in Gaussian Broadcast Channels

KAUST Repository

Zafar, Ammar; Alouini, Mohamed-Slim; Shaqfeh, Mohammad

2014-01-01

This work proposes a suboptimal multi-user scheduling scheme for Gaussian broadcast channels which improves upon the classical single user selection, while considerably reducing complexity as compared to the optimal superposition coding with successful interference cancellation. The proposed scheme combines the two users with the maximum weighted instantaneous rate using superposition coding. The instantaneous rate and power allocation are derived in closed-form, while the long term rate of each user is derived in integral form for all channel distributions. Numerical results are then provided to characterize the prospected gains of the proposed scheme.

A Suboptimal Scheme for Multi-User Scheduling in Gaussian Broadcast Channels

KAUST Repository

Zafar, Ammar

2014-05-28

This work proposes a suboptimal multi-user scheduling scheme for Gaussian broadcast channels which improves upon the classical single user selection, while considerably reducing complexity as compared to the optimal superposition coding with successful interference cancellation. The proposed scheme combines the two users with the maximum weighted instantaneous rate using superposition coding. The instantaneous rate and power allocation are derived in closed-form, while the long term rate of each user is derived in integral form for all channel distributions. Numerical results are then provided to characterize the prospected gains of the proposed scheme.

Surface dynamics of voltage-gated ion channels

Science.gov (United States)

Heine, Martin; Ciuraszkiewicz, Anna; Voigt, Andreas; Heck, Jennifer; Bikbaev, Arthur

2016-01-01

ABSTRACT Neurons encode information in fast changes of the membrane potential, and thus electrical membrane properties are critically important for the integration and processing of synaptic inputs by a neuron. These electrical properties are largely determined by ion channels embedded in the membrane. The distribution of most ion channels in the membrane is not spatially uniform: they undergo activity-driven changes in the range of minutes to days. Even in the range of milliseconds, the composition and topology of ion channels are not static but engage in highly dynamic processes including stochastic or activity-dependent transient association of the pore-forming and auxiliary subunits, lateral diffusion, as well as clustering of different channels. In this review we briefly discuss the potential impact of mobile sodium, calcium and potassium ion channels and the functional significance of this for individual neurons and neuronal networks. PMID:26891382

Manufacturing method of fuel assembly and channel box for the fuel assembly

International Nuclear Information System (INIS)

Fujieda, Tadashi; Inagaki, Masatoshi; Takase, Iwao; Nishino, Yoshitaka; Yamashita, Jun-ichi; Yamanaka, Akihiro; Ito, Ken-ichi; Nakajima, Junjiro; Seto, Takehiro.

1998-01-01

An MOX fuel assembly to be used for a BWR type reactor comprises a channel box, a great number of fuel rod bundles and a water rod. BP members incorporated with a burnable neutron absorbing poison (BP) are buried in the vicinity of corners of four sides of the channel box in the longitudinal direction. The channel box is formed by fitting the BP members in concaves formed in the longitudinal direction of zircaloy plates, laminating other zircaloy plates and welding the seams. Then, hot rolling, cold rolling and annealing are conducted to form them into a single plate. Integrated two single plates after bending treatment are abutted and welded, and heat-treatment is applied to complete the channel box. With such a constitution, since the BP member is not brought into contact with reactor water directly, crevice corrosion or galvanic corrosion can be prevented. (I.N.)

Gating of Connexin Channels by transjunctional-voltage: Conformations and models of open and closed states.

Science.gov (United States)

Bargiello, Thaddeus A; Oh, Seunghoon; Tang, Qingxiu; Bargiello, Nicholas K; Dowd, Terry L; Kwon, Taekyung

2018-01-01

Voltage is an important physiologic regulator of channels formed by the connexin gene family. Connexins are unique among ion channels in that both plasma membrane inserted hemichannels (undocked hemichannels) and intercellular channels (aggregates of which form gap junctions) have important physiological roles. The hemichannel is the fundamental unit of gap junction voltage-gating. Each hemichannel displays two distinct voltage-gating mechanisms that are primarily sensitive to a voltage gradient formed along the length of the channel pore (the transjunctional voltage) rather than sensitivity to the absolute membrane potential (V m or V i-o ). These transjunctional voltage dependent processes have been termed V j - or fast-gating and loop- or slow-gating. Understanding the mechanism of voltage-gating, defined as the sequence of voltage-driven transitions that connect open and closed states, first and foremost requires atomic resolution models of the end states. Although ion channels formed by connexins were among the first to be characterized structurally by electron microscopy and x-ray diffraction in the early 1980's, subsequent progress has been slow. Much of the current understanding of the structure-function relations of connexin channels is based on two crystal structures of Cx26 gap junction channels. Refinement of crystal structure by all-atom molecular dynamics and incorporation of charge changing protein modifications has resulted in an atomic model of the open state that arguably corresponds to the physiologic open state. Obtaining validated atomic models of voltage-dependent closed states is more challenging, as there are currently no methods to solve protein structure while a stable voltage gradient is applied across the length of an oriented channel. It is widely believed that the best approach to solve the atomic structure of a voltage-gated closed ion channel is to apply different but complementary experimental and computational methods and to use

Zinc oxide integrated area efficient high output low power wavy channel thin film transistor

International Nuclear Information System (INIS)

Hanna, A. N.; Ghoneim, M. T.; Bahabry, R. R.; Hussain, A. M.; Hussain, M. M.

2013-01-01

We report an atomic layer deposition based zinc oxide channel material integrated thin film transistor using wavy channel architecture allowing expansion of the transistor width in the vertical direction using the fin type features. The experimental devices show area efficiency, higher normalized output current, and relatively lower power consumption compared to the planar architecture. This performance gain is attributed to the increased device width and an enhanced applied electric field due to the architecture when compared to a back gated planar device with the same process conditions

Zinc oxide integrated area efficient high output low power wavy channel thin film transistor

KAUST Repository

Hanna, Amir; Ghoneim, Mohamed T.; Bahabry, Rabab R.; Hussain, Aftab M.; Hussain, Muhammad Mustafa

2013-01-01

We report an atomic layer deposition based zinc oxide channel material integrated thin film transistor using wavy channel architecture allowing expansion of the transistor width in the vertical direction using the fin type features. The experimental devices show area efficiency, higher normalized output current, and relatively lower power consumption compared to the planar architecture. This performance gain is attributed to the increased device width and an enhanced applied electric field due to the architecture when compared to a back gated planar device with the same process conditions.

Zinc oxide integrated area efficient high output low power wavy channel thin film transistor

KAUST Repository

Hanna, Amir

2013-11-26

We report an atomic layer deposition based zinc oxide channel material integrated thin film transistor using wavy channel architecture allowing expansion of the transistor width in the vertical direction using the fin type features. The experimental devices show area efficiency, higher normalized output current, and relatively lower power consumption compared to the planar architecture. This performance gain is attributed to the increased device width and an enhanced applied electric field due to the architecture when compared to a back gated planar device with the same process conditions.

Application of gamma irradiation on forming protein-based edible films

International Nuclear Information System (INIS)

Sabato, Susy Frey

2000-01-01

In the last decade considerable interest has been addressed to the development of protein-based edible films due to their application in the food industry, as a substitute to traditional plastic films. The use of soy and whey proteins to form those films has been investigated, using heat, chemical and enzymatic processes. Gamma irradiation was recently reported to form caseinate-based edible films, due to the increase of the cohesive strength of the proteins by the formation of cross-links. This work aimed to verify the role of the gamma irradiation in the process of forming edible films from soy protein isolate (SPI) alone and in complex mixtures, that is, mixed with whey protein isolate (WPI), with carbethoxymethyl cellulose (CMC) and with poly(vinyl)alcohol (PVA). Gamma irradiation treatment improved significantly the mechanical properties for all films. The mechanical behavior is strongly related to the formulation, showing synergy between the gamma irradiation and the CMC, mainly for SPI-based films. SPI-based films presented a trend to decrease the water vapor permeability values when irradiated. The CMC addition showed significant improvements on the permeability for films from SPI and from the mixture of SPI with WPI. (author)

Properties of the intracellular transient receptor potential (TRP) channel in yeast, Yvc1.

Science.gov (United States)

Chang, Yiming; Schlenstedt, Gabriel; Flockerzi, Veit; Beck, Andreas

2010-05-17

Transient receptor potential (TRP) channels are found among mammals, flies, worms, ciliates, Chlamydomonas, and yeast but are absent in plants. These channels are believed to be tetramers of proteins containing six transmembrane domains (TMs). Their primary structures are diverse with sequence similarities only in some short amino acid sequence motifs mainly within sequences covering TM5, TM6, and adjacent domains. In the yeast genome, there is one gene encoding a TRP-like sequence. This protein forms an ion channel in the vacuolar membrane and is therefore called Yvc1 for yeast vacuolar conductance 1. In the following we summarize its prominent features. Copyright 2009 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Expression of G-protein inwardly rectifying potassium channels (GIRKs in lung cancer cell lines

Directory of Open Access Journals (Sweden)

Schuller Hildegard M

2005-08-01

Full Text Available Abstract Background Previous data from our laboratory has indicated that there is a functional link between the β-adrenergic receptor signaling pathway and the G-protein inwardly rectifying potassium channel (GIRK1 in human breast cancer cell lines. We wanted to determine if GIRK channels were expressed in lung cancers and if a similar link exists in lung cancer. Methods GIRK1-4 expression and levels were determined by reverse transcription polymerase chain reaction (RT-PCR and real-time PCR. GIRK protein levels were determined by western blots and cell proliferation was determined by a 5-bromo-2'-deoxyuridine (BrdU assay. Results GIRK1 mRNA was expressed in three of six small cell lung cancer (SCLC cell lines, and either GIRK2, 3 or 4 mRNA expression was detected in all six SCLC cell lines. Treatment of NCI-H69 with β2-adrenergic antagonist ICI 118,551 (100 μM daily for seven days led to slight decreases of GIRK1 mRNA expression levels. Treatment of NCI-H69 with the β-adrenergic agonist isoproterenol (10 μM decreased growth rates in these cells. The GIRK inhibitor U50488H (2 μM also inhibited proliferation, and this decrease was potentiated by isoproterenol. In the SCLC cell lines that demonstrated GIRK1 mRNA expression, we also saw GIRK1 protein expression. We feel these may be important regulatory pathways since no expression of mRNA of the GIRK channels (1 & 2 was found in hamster pulmonary neuroendocrine cells, a suggested cell of origin for SCLC, nor was GIRK1 or 2 expression found in human small airway epithelial cells. GIRK (1,2,3,4 mRNA expression was also seen in A549 adenocarcinoma and NCI-H727 carcinoid cell lines. GIRK1 mRNA expression was not found in tissue samples from adenocarcinoma or squamous cancer patients, nor was it found in NCI-H322 or NCI-H441 adenocarcinoma cell lines. GIRK (1,3,4 mRNA expression was seen in three squamous cell lines, GIRK2 was only expressed in one squamous cell line. However, GIRK1 protein

Sub-cellular distribution and translocation of TRP channels.

Science.gov (United States)

Toro, Carlos A; Arias, Luis A; Brauchi, Sebastian

2011-01-01

Cellular electrical activity is the result of a highly complex processes that involve the activation of ion channel proteins. Ion channels make pores on cell membranes that rapidly transit between conductive and non-conductive states, allowing different ions to flow down their electrochemical gradients across cell membranes. In the case of neuronal cells, ion channel activity orchestrates action potentials traveling through axons, enabling electrical communication between cells in distant parts of the body. Somatic sensation -our ability to feel touch, temperature and noxious stimuli- require ion channels able to sense and respond to our peripheral environment. Sensory integration involves the summing of various environmental cues and their conversion into electrical signals. Members of the Transient Receptor Potential (TRP) family of ion channels have emerged as important mediators of both cellular sensing and sensory integration. The regulation of the spatial and temporal distribution of membrane receptors is recognized as an important mechanism for controlling the magnitude of the cellular response and the time scale on which cellular signaling occurs. Several studies have shown that this mechanism is also used by TRP channels to modulate cellular response and ultimately fulfill their physiological function as sensors. However, the inner-working of this mode of control for TRP channels remains poorly understood. The question of whether TRPs intrinsically regulate their own vesicular trafficking or weather the dynamic regulation of TRP channel residence on the cell surface is caused by extrinsic changes in the rates of vesicle insertion or retrieval remain open. This review will examine the evidence that sub-cellular redistribution of TRP channels plays an important role in regulating their activity and explore the mechanisms that control the trafficking of vesicles containing TRP channels.

Computer Simulation Studies of Ion Channels at High Temperatures

Science.gov (United States)

Song, Hyun Deok

The gramicidin channel is the smallest known biological ion channel, and it exhibits cation selectivity. Recently, Dr. John Cuppoletti's group at the University of Cincinnati showed that the gramicidin channel can function at high temperatures (360 ˜ 380K) with significant currents. This finding may have significant implications for fuel cell technology. In this thesis, we have examined the gramicidin channel at 300K, 330K, and 360K by computer simulation. We have investigated how the temperature affects the current and differences in magnitude of free energy between the two gramicidin forms, the helical dimer (HD) and the double helix (DH). A slight decrease of the free energy barrier inside the gramicidin channel and increased diffusion at high temperatures result in an increase of current. An applied external field of 0.2V/nm along the membrane normal results in directly observable ion transport across the channels at high temperatures for both HD and DH forms. We found that higher temperatures also affect the probability distribution of hydrogen bonds, the bending angle, the distance between dimers, and the size of the pore radius for the helical dimer structure. These findings may be related to the gating of the gramicidin channel. Methanococcus jannaschii (MJ) is a methane-producing thermophile, which was discovered at a depth of 2600m in a Pacific Ocean vent in 1983. It has the ability to thrive at high temperatures and high pressures, which are unfavorable for most life forms. There have been some experiments to study its stability under extreme conditions, but still the origin of the stability of MJ is not exactly known. MJ0305 is the chloride channel protein from the thermophile MJ. After generating a structure of MJ0305 by homology modeling based on the Ecoli ClC templates, we examined the thermal stability, and the network stability from the change of network entropy calculated from the adjacency matrices of the protein. High temperatures increase the

Integrated Manufacturing of Aerospace Components by Superplastic Forming Technology

Directory of Open Access Journals (Sweden)

Ju Min Kyung

2015-01-01

Full Text Available Aerospace vehicle requires lightweight structures to obtain weight saving and fuel efficiency. It is known that superplastic characteristics of some materials provide significant opportunity for forming complicated, lightweight components of aerospace structure. One of the most important advantages of using superplastic forming process is its simplicity to form integral parts and economy in tooling[1]. For instance, it can be applied to blow-forming, in which a metal sheet is deformed due to the pressure difference of hydrostatic gas on both sides of the sheet. Since the loading medium is gas pressure difference, this forming is different from conventional sheet metal forming technique in that this is stress-controlled rather than strain and strain rate controlled. This method is especially advantageous when several sheet metals are formed into complex shapes. In this study, it is demonstrated that superplastic forming process with titanium and steel alloy can be applied to manufacturing lightweight integral structures of aerospace structural parts and rocket propulsion components. The result shows that the technology to design and develop the forming process of superplastic forming can be applied for near net shape forming of a complex contour of a thrust chamber and a toroidal fuel tank.

Expression of Water Channel Proteins in Mesembryanthemum crystallinum1

Science.gov (United States)

Kirch, Hans-Hubert; Vera-Estrella, Rosario; Golldack, Dortje; Quigley, Francoise; Michalowski, Christine B.; Barkla, Bronwyn J.; Bohnert, Hans J.

2000-01-01

We have characterized transcripts for nine major intrinsic proteins (MIPs), some of which function as water channels (aquaporins), from the ice plant Mesembryanthemum crystallinum. To determine the cellular distribution and expression of these MIPs, oligopeptide-based antibodies were generated against MIP-A, MIP-B, MIP-C, or MIP-F, which, according to sequence and functional characteristics, are located in the plasma membrane (PM) and tonoplast, respectively. MIPs were most abundant in cells involved in bulk water flow and solute flux. The tonoplast MIP-F was found in all cells, while signature cell types identified different PM-MIPs: MIP-A predominantly in phloem-associated cells, MIP-B in xylem parenchyma, and MIP-C in the epidermis and endodermis of immature roots. Membrane protein analysis confirmed MIP-F as tonoplast located. MIP-A and MIP-B were found in tonoplast fractions and also in fractions distinct from either the tonoplast or PM. MIP-C was most abundant but not exclusive to PM fractions, where it is expected based on its sequence signature. We suggest that within the cell, MIPs are mobile, which is similar to aquaporins cycling through animal endosomes. MIP cycling and the differential regulation of these proteins observed under conditions of salt stress may be fundamental for the control of tissue water flux. PMID:10806230

Conditions for integrability of a 3-form

Czech Academy of Sciences Publication Activity Database

Vanžura, Jiří

2017-01-01

Roč. 53, č. 5 (2017), s. 371-380 ISSN 0044-8753 Institutional support: RVO:67985840 Keywords : multisymplectic 3-form * 6-dimensional manifold * integrability Subject RIV: BA - General Mathematics OBOR OECD: Applied mathematics http://hdl.handle.net/10338.dmlcz/147026

Claudin-21 Has a Paracellular Channel Role at Tight Junctions

KAUST Repository

Tanaka, Hiroo; Yamamoto, Yasuko; Kashihara, Hiroka; Yamazaki, Yuji; Tani, Kazutoshi; Fujiyoshi, Yoshinori; Mineta, Katsuhiko; Takeuchi, Kosei; Tamura, Atsushi; Tsukita, Sachiko

2016-01-01

Claudin protein family members, of which there are at least 27 in humans and mice, polymerize to form tight junctions (TJs) between epithelial cells, in a tissue- and developmental stage-specific manner. Claudins have a paracellular barrier function. In addition, certain claudins function as paracellular channels for small ions and/or solutes by forming selective pores at the TJs, although the specific claudins involved and their functional mechanisms are still in question. Here we show for the first time that claudin-21, which is more highly expressed in the embryonic than the postnatal stages, acts as a paracellular channel for small cations, such as Na+, similar to the typical channel-type claudins claudin-2 and -15. Claudin-21 also allows the paracellular passage of larger solutes. Our findings suggest that claudin-21-based TJs allow the passage of small and larger solutes by both paracellular channel-based and some additional mechanisms. © 2016, American Society for Microbiology. All Rights Reserved.

Claudin-21 Has a Paracellular Channel Role at Tight Junctions

KAUST Repository

Tanaka, Hiroo

2016-01-05

Claudin protein family members, of which there are at least 27 in humans and mice, polymerize to form tight junctions (TJs) between epithelial cells, in a tissue- and developmental stage-specific manner. Claudins have a paracellular barrier function. In addition, certain claudins function as paracellular channels for small ions and/or solutes by forming selective pores at the TJs, although the specific claudins involved and their functional mechanisms are still in question. Here we show for the first time that claudin-21, which is more highly expressed in the embryonic than the postnatal stages, acts as a paracellular channel for small cations, such as Na+, similar to the typical channel-type claudins claudin-2 and -15. Claudin-21 also allows the paracellular passage of larger solutes. Our findings suggest that claudin-21-based TJs allow the passage of small and larger solutes by both paracellular channel-based and some additional mechanisms. © 2016, American Society for Microbiology. All Rights Reserved.

Crystallographic and single-particle analyses of native- and nucleotide-bound forms of the cystic fibrosis transmembrane conductance regulator (CFTR) protein.

Science.gov (United States)

Awayn, N H; Rosenberg, M F; Kamis, A B; Aleksandrov, L A; Riordan, J R; Ford, R C

2005-11-01

Cystic fibrosis, one of the major human inherited diseases, is caused by defects in the CFTR (cystic fibrosis transmembrane conductance regulator), a cell-membrane protein. CFTR acts as a chloride channel which can be opened by ATP. Low-resolution structural studies of purified recombinant human CFTR are described in the present paper. Localization of the C-terminal decahistidine tag in CFTR was achieved by Ni2+-nitriloacetate nanogold labelling, followed by electron microscopy and single-particle analysis. The presence of the gold label appears to improve the single-particle-alignment procedure. Projection structures of CFTR from two-dimensional crystals analysed by electron crystallography displayed two alternative conformational states in the presence of nucleotide and nanogold, but only one form of the protein was observed in the quiescent (nucleotide-free) state.

A novel form of the membrane protein CD147 that contains an extra Ig-like domain and interacts homophilically

Directory of Open Access Journals (Sweden)

Brown Marion H

2003-11-01

Full Text Available Abstract Background CD147 is a broadly distributed integral membrane glycoprotein with two Ig-like domains implicated in a wide range of functions. It is associated at the cell surface with the monocarboxylate transporters MCT1 and 4 but interactions of the extracellular region have not been characterised. Results We report the characterisation of a form of CD147 with an additional membrane-distal Ig-like domain. In contrast to the two domain form, this three domain form of CD147 interacts homophilically. Surface plasmon resonance analysis using recombinant proteins showed that the interaction was of low affinity (KD ~ 40 μM and this is typical of many interactions between membrane proteins. cDNA for the 3 domain form are rare but have been identified in human and mouse retina. Conclusion The finding that the three domain form of CD147 has an extracellular ligand, that is it interacts homophilically, suggests this interaction may be important in aligning lactate transporters in the retina where lactate is an important metabolite.

DipA, a pore-forming protein in the outer membrane of Lyme disease spirochetes exhibits specificity for the permeation of dicarboxylates.

Directory of Open Access Journals (Sweden)

Marcus Thein

Full Text Available Lyme disease Borreliae are highly dependent on the uptake of nutrients provided by their hosts. Our study describes the identification of a 36 kDa protein that functions as putative dicarboxylate-specific porin in the outer membrane of Lyme disease Borrelia. The protein was purified by hydroxyapatite chromatography from Borrelia burgdorferi B31 and designated as DipA, for dicarboxylate-specific porin A. DipA was partially sequenced, and corresponding genes were identified in the genomes of B. burgdorferi B31, Borrelia garinii PBi and Borrelia afzelii PKo. DipA exhibits high homology to the Oms38 porins of relapsing fever Borreliae. B. burgdorferi DipA was characterized using the black lipid bilayer assay. The protein has a single-channel conductance of 50 pS in 1 M KCl, is slightly selective for anions with a permeability ratio for cations over anions of 0.57 in KCl and is not voltage-dependent. The channel could be partly blocked by different di- and tricarboxylic anions. Particular high stability constants up to about 28,000 l/mol (in 0.1 M KCl were obtained among the 11 tested anions for oxaloacetate, 2-oxoglutarate and citrate. The results imply that DipA forms a porin specific for dicarboxylates which may play an important role for the uptake of specific nutrients in different Borrelia species.

Legionella pneumophila secretes a mitochondrial carrier protein during infection.

Directory of Open Access Journals (Sweden)

Pavel Dolezal

2012-01-01

Full Text Available The Mitochondrial Carrier Family (MCF is a signature group of integral membrane proteins that transport metabolites across the mitochondrial inner membrane in eukaryotes. MCF proteins are characterized by six transmembrane segments that assemble to form a highly-selective channel for metabolite transport. We discovered a novel MCF member, termed Legionellanucleotide carrier Protein (LncP, encoded in the genome of Legionella pneumophila, the causative agent of Legionnaire's disease. LncP was secreted via the bacterial Dot/Icm type IV secretion system into macrophages and assembled in the mitochondrial inner membrane. In a yeast cellular system, LncP induced a dominant-negative phenotype that was rescued by deleting an endogenous ATP carrier. Substrate transport studies on purified LncP reconstituted in liposomes revealed that it catalyzes unidirectional transport and exchange of ATP transport across membranes, thereby supporting a role for LncP as an ATP transporter. A hidden Markov model revealed further MCF proteins in the intracellular pathogens, Legionella longbeachae and Neorickettsia sennetsu, thereby challenging the notion that MCF proteins exist exclusively in eukaryotic organisms.

[Mechanism of action of neurotoxins acting on the inactivation of voltage-gated sodium channels].

Science.gov (United States)

Benoit, E

1998-01-01

This review focuses on the mechanism(s) of action of neurotoxins acting on the inactivation of voltage-gated Na channels. Na channels are transmembrane proteins which are fundamental for cellular communication. These proteins form pores in the plasma membrane allowing passive ionic movements to occur. Their opening and closing are controlled by gating systems which depend on both membrane potential and time. Na channels have three functional properties, mainly studied using electrophysiological and biochemical techniques, to ensure their role in the generation and propagation of action potentials: 1) a highly selectivity for Na ions, 2) a rapid opening ("activation"), responsible for the depolarizing phase of the action potential, and 3) a late closing ("inactivation") involved in the repolarizing phase of the action potential. As an essential protein for membrane excitability, the Na channel is the specific target of a number of vegetal and animal toxins which, by binding to the channel, alter its activity by affecting one or more of its properties. At least six toxin receptor sites have been identified on the neuronal Na channel on the basis of binding studies. However, only toxins interacting with four of these sites (sites 2, 3, 5 et 6) produce alterations of channel inactivation. The maximal percentage of Na channels modified by the binding of neurotoxins to sites 2 (batrachotoxin and some alkaloids), 3 (alpha-scorpion and sea anemone toxins), 5 (brevetoxins and ciguatoxins) et 6 (delta-conotoxins) is different according to the site considered. However, in all cases, these channels do not inactivate. Moreover, Na channels modified by toxins which bind to sites 2, 5 and 6 activate at membrane potentials more negative than do unmodified channels. The physiological consequences of Na channel modifications, induced by the binding of neurotoxins to sites 2, 3, 5 and 6, are (i) an inhibition of cellular excitability due to an important membrane depolarization (site

Cholesterol up-regulates neuronal G protein-gated inwardly rectifying potassium (GIRK) channel activity in the hippocampus.

Science.gov (United States)

Bukiya, Anna N; Durdagi, Serdar; Noskov, Sergei; Rosenhouse-Dantsker, Avia

2017-04-14

Hypercholesterolemia is a well known risk factor for the development of neurodegenerative disease. However, the underlying mechanisms are mostly unknown. In recent years, it has become increasingly evident that cholesterol-driven effects on physiology and pathophysiology derive from its ability to alter the function of a variety of membrane proteins including ion channels. Yet, the effect of cholesterol on G protein-gated inwardly rectifying potassium (GIRK) channels expressed in the brain is unknown. GIRK channels mediate the actions of inhibitory brain neurotransmitters. As a result, loss of GIRK function can enhance neuron excitability, whereas gain of GIRK function can reduce neuronal activity. Here we show that in rats on a high-cholesterol diet, cholesterol levels in hippocampal neurons are increased. We also demonstrate that cholesterol plays a critical role in modulating neuronal GIRK currents. Specifically, cholesterol enrichment of rat hippocampal neurons resulted in enhanced channel activity. In accordance, elevated currents upon cholesterol enrichment were also observed in Xenopus oocytes expressing GIRK2 channels, the primary GIRK subunit expressed in the brain. Furthermore, using planar lipid bilayers, we show that although cholesterol did not affect the unitary conductance of GIRK2, it significantly enhanced the frequency of channel openings. Last, combining computational and functional approaches, we identified two putative cholesterol-binding sites in the transmembrane domain of GIRK2. These findings establish that cholesterol plays a critical role in modulating GIRK activity in the brain. Because up-regulation of GIRK function can reduce neuronal activity, our findings may lead to novel approaches for prevention and therapy of cholesterol-driven neurodegenerative disease. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Ion channel activity of membrane vesicles released from sea urchin sperm during the acrosome reaction

International Nuclear Information System (INIS)

Schulz, Joseph R.; Vega-Beltran, Jose L. de la; Beltran, Carmen; Vacquier, Victor D.; Darszon, Alberto

2004-01-01

The sperm acrosome reaction (AR) involves ion channel activation. In sea urchin sperm, the AR requires Ca 2+ and Na + influx and K + and H + efflux. During the AR, the plasma membrane fuses with the acrosomal vesicle membrane forming hybrid membrane vesicles that are released from sperm into the medium. This paper reports the isolation and preliminary characterization of these acrosome reaction vesicles (ARVs), using synaptosome-associated protein of 25 kDa (SNAP-25) as a marker. Isolated ARVs have a unique protein composition. The exocytosis regulatory proteins vesicle-associated membrane protein and SNAP-25 are inside ARVs, as judged by protease protection experiments, and membrane associated based on Triton X-114 partitioning. ARVs fused with planar bilayers display three main types of single channel activity. The most frequently recorded channel is cationic, weakly voltage dependent and has a low open probability that increases with negative potentials. This channel is activated by cAMP, blocked by Ba 2+ , and has a PK + /PNa + selectivity of 4.5. ARVs represent a novel membrane preparation suitable to deepen our understanding of ion channel activity in the AR and during fertilization

Slack sodium-activated potassium channel membrane expression requires p38 mitogen-activated protein kinase phosphorylation.

Science.gov (United States)

Gururaj, Sushmitha; Fleites, John; Bhattacharjee, Arin

2016-04-01

p38 MAPK has long been understood as an inducible kinase under conditions of cellular stress, but there is now increasing evidence to support its role in the regulation of neuronal function. Several phosphorylation targets have been identified, an appreciable number of which are ion channels, implicating the possible involvement of p38 MAPK in neuronal excitability. The KNa channel Slack is an important protein to be studied as it is highly and ubiquitously expressed in DRG neurons and is important in the maintenance of their firing accommodation. We sought to examine if the Slack channel could be a substrate of p38 MAPK activity. First, we found that the Slack C-terminus contains two putative p38 MAPK phosphorylation sites that are highly conserved across species. Second, we show via electrophysiology experiments that KNa currents and further, Slack currents, are subject to tonic modulation by p38 MAPK. Third, biochemical approaches revealed that Slack channel regulation by p38 MAPK occurs through direct phosphorylation at the two putative sites of interaction, and mutating both sites prevented surface expression of Slack channels. Based on these results, we conclude that p38 MAPK is an obligate regulator of Slack channel function via the trafficking of channels into the membrane. The present study identifies Slack KNa channels as p38 MAPK substrates. Copyright © 2015 Elsevier Ltd. All rights reserved.

The Proteins API: accessing key integrated protein and genome information.

Science.gov (United States)

Nightingale, Andrew; Antunes, Ricardo; Alpi, Emanuele; Bursteinas, Borisas; Gonzales, Leonardo; Liu, Wudong; Luo, Jie; Qi, Guoying; Turner, Edd; Martin, Maria

2017-07-03

The Proteins API provides searching and programmatic access to protein and associated genomics data such as curated protein sequence positional annotations from UniProtKB, as well as mapped variation and proteomics data from large scale data sources (LSS). Using the coordinates service, researchers are able to retrieve the genomic sequence coordinates for proteins in UniProtKB. This, the LSS genomics and proteomics data for UniProt proteins is programmatically only available through this service. A Swagger UI has been implemented to provide documentation, an interface for users, with little or no programming experience, to 'talk' to the services to quickly and easily formulate queries with the services and obtain dynamically generated source code for popular programming languages, such as Java, Perl, Python and Ruby. Search results are returned as standard JSON, XML or GFF data objects. The Proteins API is a scalable, reliable, fast, easy to use RESTful services that provides a broad protein information resource for users to ask questions based upon their field of expertise and allowing them to gain an integrated overview of protein annotations available to aid their knowledge gain on proteins in biological processes. The Proteins API is available at (http://www.ebi.ac.uk/proteins/api/doc). © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

ProNormz--an integrated approach for human proteins and protein kinases normalization.

Science.gov (United States)

Subramani, Suresh; Raja, Kalpana; Natarajan, Jeyakumar

2014-02-01

The task of recognizing and normalizing protein name mentions in biomedical literature is a challenging task and important for text mining applications such as protein-protein interactions, pathway reconstruction and many more. In this paper, we present ProNormz, an integrated approach for human proteins (HPs) tagging and normalization. In Homo sapiens, a greater number of biological processes are regulated by a large human gene family called protein kinases by post translational phosphorylation. Recognition and normalization of human protein kinases (HPKs) is considered to be important for the extraction of the underlying information on its regulatory mechanism from biomedical literature. ProNormz distinguishes HPKs from other HPs besides tagging and normalization. To our knowledge, ProNormz is the first normalization system available to distinguish HPKs from other HPs in addition to gene normalization task. ProNormz incorporates a specialized synonyms dictionary for human proteins and protein kinases, a set of 15 string matching rules and a disambiguation module to achieve the normalization. Experimental results on benchmark BioCreative II training and test datasets show that our integrated approach achieve a fairly good performance and outperforms more sophisticated semantic similarity and disambiguation systems presented in BioCreative II GN task. As a freely available web tool, ProNormz is useful to developers as extensible gene normalization implementation, to researchers as a standard for comparing their innovative techniques, and to biologists for normalization and categorization of HPs and HPKs mentions in biomedical literature. URL: http://www.biominingbu.org/pronormz. Copyright © 2013 Elsevier Inc. All rights reserved.

An integrated analog O/E/O link for multi-channel laser neurons

Energy Technology Data Exchange (ETDEWEB)

Nahmias, Mitchell A., E-mail: mnahmias@princeton.edu; Tait, Alexander N.; Tolias, Leonidas; Chang, Matthew P.; Ferreira de Lima, Thomas; Shastri, Bhavin J.; Prucnal, Paul R. [Electrical Engineering Department, Princeton University, 41 Olden St, Princeton, New Jersey 08540 (United States)

2016-04-11

We demonstrate an analog O/E/O electronic link to allow integrated laser neurons to accept many distinguishable, high bandwidth input signals simultaneously. This device utilizes wavelength division multiplexing to achieve multi-channel fan-in, a photodetector to sum signals together, and a laser cavity to perform a nonlinear operation. Its speed outpaces accelerated-time neuromorphic electronics, and it represents a viable direction towards scalable networking approaches.

Subcellular localization and logistics of integral membrane protein biogenesis in Escherichia coli.

Science.gov (United States)

Bogdanov, Mikhail; Aboulwafa, Mohammad; Saier, Milton H

2013-01-01

Transporters catalyze entry and exit of molecules into and out of cells and organelles, and protein-lipid interactions influence their activities. The bacterial phosphoenolpyruvate: sugar phosphotransferase system (PTS) catalyzes transport-coupled sugar phosphorylation as well as nonvectorial sugar phosphorylation in the cytoplasm. The vectorial process is much more sensitive to the lipid environment than the nonvectorial process. Moreover, cytoplasmic micellar forms of these enzyme-porters have been identified, and non-PTS permeases have similarly been shown to exist in 'soluble' forms. The latter porters exhibit lipid-dependent activities and can adopt altered topologies by simply changing the lipid composition. Finally, intracellular membranes and vesicles exist in Escherichia coli leading to the following unanswered questions: (1) what determines whether a PTS permease catalyzes vectorial or nonvectorial sugar phosphorylation? (2) How do phospholipids influence relative amounts of the plasma membrane, intracellular membrane, inner membrane-derived vesicles and cytoplasmic micelles? (3) What regulates the route(s) of permease insertion and transfer into and between the different subcellular sites? (4) Do these various membranous forms have distinct physiological functions? (5) What methods should be utilized to study the biogenesis and interconversion of these membranous structures? While research concerning these questions is still in its infancy, answers will greatly enhance our understanding of protein-lipid interactions and how they control the activities, conformations, cellular locations and biogenesis of integral membrane proteins. Copyright © 2013 S. Karger AG, Basel.

The gastric H,K-ATPase blocker lansoprazole is an inhibitor of chloride channels

Science.gov (United States)

Schmarda, Andreas; Dinkhauser, Patrick; Gschwentner, Martin; Ritter, Markus; Fürst, Johannes; Scandella, Elke; Wöll, Ewald; Laich, Andreas; Rossmann, Heidi; Seidler, Ursula; Lang, Florian; Paulmichl, Markus

2000-01-01

It was postulated that swelling dependent chloride channels are involved in the proton secretion of parietal cells. Since omeprazole, lansoprazole and its acid activated sulphenamide form AG2000 are structurally related to phenol derivatives known to block swelling dependent chloride channels, we set out to test, whether these substances – which are known to block the H,K-ATPase – could also lead to an inhibition of swelling-dependent chloride channels. Swelling-dependent chloride channels – characterized in many different cell types – show highly conserved biophysical and pharmacological features, therefore we investigated the effect of omeprazole, lansoprazole and its acid activated sulphenamide form AG2000 on swelling-dependent chloride channels elicited in fibroblasts, after the reduction of the extracellular osmolarity. Omeprazole, lansoprazole and its acid activated sulphenamide form AG2000 are able to block swelling-dependent chloride channels (IClswell). Lansoprazole and its protonated metabolite AG2000 act on at least two different sites of the IClswell protein: on an extracellular site which seems to be in a functional proximity to the nucleotide binding site, and on an intracellular site which allows the formation of disulfide-bridges. The inhibition of the proton pump and the simultaneous blocking of chloride channels by omeprazole, lansoprazole and its acid activated sulphenamide form AG2000, as described here could be an effective mode to restrict proton secretion in parietal cells. PMID:10711360

Bromodomain-containing Protein 4 Activates Voltage-gated Sodium Channel 1.7 Transcription in Dorsal Root Ganglia Neurons to Mediate Thermal Hyperalgesia in Rats.

Science.gov (United States)

Hsieh, Ming-Chun; Ho, Yu-Cheng; Lai, Cheng-Yuan; Wang, Hsueh-Hsiao; Lee, An-Sheng; Cheng, Jen-Kun; Chau, Yat-Pang; Peng, Hsien-Yu

2017-11-01

Bromodomain-containing protein 4 binds acetylated promoter histones and promotes transcription; however, the role of bromodomain-containing protein 4 in inflammatory hyperalgesia remains unclear. Male Sprague-Dawley rats received hind paw injections of complete Freund's adjuvant to induce hyperalgesia. The dorsal root ganglia were examined to detect changes in bromodomain-containing protein 4 expression and the activation of genes involved in the expression of voltage-gated sodium channel 1.7, which is a key pain-related ion channel. The intraplantar complete Freund's adjuvant injections resulted in thermal hyperalgesia (4.0 ± 1.5 s; n = 7). The immunohistochemistry and immunoblotting results demonstrated an increase in the bromodomain-containing protein 4-expressing dorsal root ganglia neurons (3.78 ± 0.38 fold; n = 7) and bromodomain-containing protein 4 protein levels (2.62 ± 0.39 fold; n = 6). After the complete Freund's adjuvant injection, histone H3 protein acetylation was enhanced in the voltage-gated sodium channel 1.7 promoter, and cyclin-dependent kinase 9 and phosphorylation of RNA polymerase II were recruited to this area. Furthermore, the voltage-gated sodium channel 1.7-mediated currents were enhanced in neurons of the complete Freund's adjuvant rats (55 ± 11 vs. 19 ± 9 pA/pF; n = 4 to 6 neurons). Using bromodomain-containing protein 4-targeted antisense small interfering RNA to the complete Freund's adjuvant-treated rats, the authors demonstrated a reduction in the expression of bromodomain-containing protein 4 (0.68 ± 0.16 fold; n = 7), a reduction in thermal hyperalgesia (7.5 ± 1.5 s; n = 7), and a reduction in the increased voltage-gated sodium channel 1.7 currents (21 ± 4 pA/pF; n = 4 to 6 neurons). Complete Freund's adjuvant triggers enhanced bromodomain-containing protein 4 expression, ultimately leading to the enhanced excitability of nociceptive neurons and thermal hyperalgesia. This effect is

Structure, Function, Self-Assembly and Origin of Simple Membrane Proteins

Science.gov (United States)

Pohorille, Andrew

2003-01-01

Integral membrane proteins perform such essential cellular functions as transport of ions, nutrients and waste products across cell walls, transduction of environmental signals, regulation of cell fusion, recognition of other cells, energy capture and its conversion into high-energy compounds. In fact, 30-40% of genes in modem organisms codes for membrane proteins. Although contemporary membrane proteins or their functional assemblies can be quite complex, their transmembrane fragments are usually remarkably simple. The most common structural motif for these fragments is a bundle of alpha-helices, but occasionally it could be a beta-barrel. In a series of molecular dynamics computer simulations we investigated self-organizing properties of simple membrane proteins based on these structural motifs. Specifically, we studied folding and insertion into membranes of short, nonpolar or amphiphatic peptides. We also investigated glycophorin A, a peptide that forms sequence-specific dimers, and a transmembrane aggregate of four identical alpha-helices that forms an efficient and selective voltage-gated proton channel was investigated. Many peptides are attracted to water-membrane interfaces. Once at the interface, nonpolar peptides spontaneously fold to a-helices. Whenever the sequence permits, peptides that contain both polar and nonpolar amino also adopt helical structures, in which polar and nonpolar amino acid side chains are immersed in water and membrane, respectively. Specific identity of side chains is less important. Helical peptides at the interface could insert into the membrane and adopt a transmembrane conformation. However, insertion of a single helix is unfavorable because polar groups in the peptide become completely dehydrated upon insertion. The unfavorable free energy of insertion can be regained by spontaneous association of peptides in the membrane. The first step in this process is the formation of dimers, although the most common are aggregates of 4

Four crystal forms of a Bence-Jones protein

International Nuclear Information System (INIS)

Makino, Debora L.; Henschen-Edman, Agnes H.; McPherson, Alexander

2004-01-01

Four crystal forms have been grown and characterized by X-ray diffraction of a Bence-Jones protein collected from the urine of a multiple myeloma patient more than 40 y ago. The trigonal crystal form may shed some light on the formation of fibrils common to certain storage diseases. Four crystal forms have been grown and characterized by X-ray diffraction of a Bence-Jones protein collected from the urine of a multiple myeloma patient more than 40 years ago. Closely related tetragonal and orthorhombic forms belonging to space groups P4 3 2 1 2 and P2 1 2 1 2 1 , with unit-cell parameters a = b = 68.7, c = 182.1 and a = 67.7, b = 69.4, c = 87.3 Å, diffract to 1.5 and 1.9 Å, respectively. Two closely related trigonal forms, both belonging to space group P3 1 21 with unit-cell parameters a = b = 154.3 Å but differing by a doubling of the c axis, one 46.9 Å and the other 94.0 Å, diffract to 2.9 and 2.6 Å resolution, respectively. The trigonal crystal of short c-axis length shows a positive indication of twinning. The trigonal crystal of longer c axis, which appeared only after eight months of incubation at room temperature, is likely to be composed of proteolytically degraded molecules and unlike the other crystal forms contains two entire Bence-Jones dimers in the asymmetric unit. This latter crystal form may shed some light on the formation of fibrils common to certain storage diseases

The stress protein heat shock cognate 70 (Hsc70) inhibits the Transient Receptor Potential Vanilloid type 1 (TRPV1) channel.

Science.gov (United States)

Iftinca, Mircea; Flynn, Robyn; Basso, Lilian; Melo, Helvira; Aboushousha, Reem; Taylor, Lauren; Altier, Christophe

2016-01-01

Specialized cellular defense mechanisms prevent damage from chemical, biological, and physical hazards. The heat shock proteins have been recognized as key chaperones that maintain cell survival against a variety of exogenous and endogenous stress signals including noxious temperature. However, the role of heat shock proteins in nociception remains poorly understood. We carried out an expression analysis of the constitutively expressed 70 kDa heat-shock cognate protein, a member of the stress-induced HSP70 family in lumbar dorsal root ganglia from a mouse model of Complete Freund's Adjuvant-induced chronic inflammatory pain. We used immunolabeling of dorsal root ganglion neurons, behavioral analysis and patch clamp electrophysiology in both dorsal root ganglion neurons and HEK cells transfected with Hsc70 and Transient Receptor Potential Channels to examine their functional interaction in heat shock stress condition. We report an increase in protein levels of Hsc70 in mouse dorsal root ganglia, 3 days post Complete Freund's Adjuvant injection in the hind paw. Immunostaining of Hsc70 was observed in most of the dorsal root ganglion neurons, including the small size nociceptors immunoreactive to the TRPV1 channel. Standard whole-cell patch-clamp technique was used to record Transient Receptor Potential Vanilloid type 1 current after exposure to heat shock. We found that capsaicin-evoked currents are inhibited by heat shock in dorsal root ganglion neurons and transfected HEK cells expressing Hsc70 and TRPV1. Blocking Hsc70 with matrine or spergualin compounds prevented heat shock-induced inhibition of the channel. We also found that, in contrast to TRPV1, both the cold sensor channels TRPA1 and TRPM8 were unresponsive to heat shock stress. Finally, we show that inhibition of TRPV1 depends on the ATPase activity of Hsc70 and involves the rho-associated protein kinase. Our work identified Hsc70 and its ATPase activity as a central cofactor of TRPV1 channel function

Optical integrated circuit of a 40-channel electrooptical LiNbO/sub 3/ modulator for data-processing devices

Energy Technology Data Exchange (ETDEWEB)

Bukreev, I.N.; Venediktov, V.V.; Gorbatovskii, M.V.; Demina, T.P.; Kashintsev, M.A.

1988-06-01

An optical integrated circuit for a 40-channel electrooptical phase modulator has been developed. The channel waveguides are prepared through Ti thermal diffusion into a Y-cut LiNbO/sub 3/ substrate. The half-wave voltage for each channel is 1.6 V at a modulating frequency bandwidth of 0-290 MHz. Results are presented from an experiment concerning the use of the modulator as an input device for the optical processing of radio signals.

Predicting Protein Function via Semantic Integration of Multiple Networks.

Science.gov (United States)

Yu, Guoxian; Fu, Guangyuan; Wang, Jun; Zhu, Hailong

2016-01-01

Determining the biological functions of proteins is one of the key challenges in the post-genomic era. The rapidly accumulated large volumes of proteomic and genomic data drives to develop computational models for automatically predicting protein function in large scale. Recent approaches focus on integrating multiple heterogeneous data sources and they often get better results than methods that use single data source alone. In this paper, we investigate how to integrate multiple biological data sources with the biological knowledge, i.e., Gene Ontology (GO), for protein function prediction. We propose a method, called SimNet, to Semantically integrate multiple functional association Networks derived from heterogenous data sources. SimNet firstly utilizes GO annotations of proteins to capture the semantic similarity between proteins and introduces a semantic kernel based on the similarity. Next, SimNet constructs a composite network, obtained as a weighted summation of individual networks, and aligns the network with the kernel to get the weights assigned to individual networks. Then, it applies a network-based classifier on the composite network to predict protein function. Experiment results on heterogenous proteomic data sources of Yeast, Human, Mouse, and Fly show that, SimNet not only achieves better (or comparable) results than other related competitive approaches, but also takes much less time. The Matlab codes of SimNet are available at https://sites.google.com/site/guoxian85/simnet.

Discharge on boiling in a channel: effect of channel geometry on the performance characteristics of determining metals in a liquid flow by atomic emission spectrometry

International Nuclear Information System (INIS)

Zuev, B.K.; Yagov, V.V.; Grachev, A.S.

2006-01-01

Discharge on boiling in a channel was studied as a new atomization and excitation source for spectrochemical analysis in a flow of electrolyte solutions. The discharge arises between the liquid walls of a vapor lock formed in the channel of a dielectric membrane because of the rapid Joule heating of the liquid in the channel. The effect of channel geometry on the reproducibility of the integrated light intensity was studied. The background radiation spectrum was measured over the range 220-900 nm, and the possibility of determining alkali and alkaline earth metals in a flow was studied. The parameters of linear calibration equations and the detection limits for these metals are given [ru

Sedimentary processes of the lower Monterey Fan channel and channel-mouth lobe

Science.gov (United States)

Klaucke, I.; Masson, D.G.; Kenyon, Neil H.; Gardner, J.V.

2004-01-01

The distribution of deposits, sediment transport pathways and processes on the lower Monterey Fan channel and channel-mouth lobe (CML) are studied through the integration of GLORIA and TOBI sidescan sonar data with 7-kHz subbottom profiler records and sediment cores for ground-truthing. The lower Monterey channel is characterised by an up to 30-m-deep channel with poorly developed levees and alternating muddy and silty muddy overbank deposits. The channel is discontinuous, disappearing where gradients are less than about 1:350. Ground-truthing of the large CML shows that the entire CML is characterised by widespread deposits of generally fine sand, with coarser sand at the base of turbidites. Sand is particularly concentrated in finger-like areas of low-backscatter intensity and is interpreted as the result of non-turbulent sediment-gravity flows depositing metres thick massive, fine sand. TOBI sidescan sonar data reveal recent erosional features in the form of scours, secondary channels, large flow slides, and trains of blocks at the distal end of the CML. Erosion is probably related to increasing gradient as the CML approaches Murray Fracture zone and to differential loading of sandy submarine fan deposits onto pelagic clays. Reworking of older flow slides by sediment transport processes on the lobe produces trains of blocks that are several metres in diameter and aligned parallel to the flow direction. ?? 2004 Elsevier B.V. All rights reserved.

Fabrication and characterization of gels with integrated channels using 3D printing with microfluidic nozzle for tissue engineering applications.

Science.gov (United States)

Attalla, R; Ling, C; Selvaganapathy, P

2016-02-01

The lack of a simple and effective method to integrate vascular network with engineered scaffolds and tissue constructs remains one of the biggest challenges in true 3D tissue engineering. Here, we detail the use of a commercially available, low-cost, open-source 3D printer modified with a microfluidic print-head in order to develop a method for the generation of instantly perfusable vascular network integrated with gel scaffolds seeded with cells. The print-head features an integrated coaxial nozzle that allows the fabrication of hollow, calcium-polymerized alginate tubes that can be easily patterned using 3D printing techniques. The diameter of the hollow channel can be precisely controlled and varied between 500 μm - 2 mm by changing applied flow rates or print-head speed. These channels are integrated into gel layers with a thickness of 800 μm - 2.5 mm. The structural rigidity of these constructs allows the fabrication of multi-layered structures without causing the collapse of hollow channels in lower layers. The 3D printing method was fully characterized at a range of operating speeds (0-40 m/min) and corresponding flow rates (1-30 mL/min) were identified to produce precise definition. This microfluidic design also allows the incorporation of a wide range of scaffold materials as well as biological constituents such as cells, growth factors, and ECM material. Media perfusion of the channels causes a significant viability increase in the bulk of cell-laden structures over the long-term. With this setup, gel constructs with embedded arrays of hollow channels can be created and used as a potential substitute for blood vessel networks.

Role of protein sulfation in vasodilation induced by minoxidil sulfate, a K+ channel opener

International Nuclear Information System (INIS)

Meisheri, K.D.; Oleynek, J.J.; Puddington, L.

1991-01-01

Evidence from contractile, radioisotope ion flux and electrophysiological studies suggest that minoxidil sulfate (MNXS) acts as a K+ channel opener in vascular smooth muscle. This study was designed to examine possible biochemical mechanisms by which MNXS exerts such an effect. Experiments performed in the isolated rabbit mesenteric artery (RMA) showed that MNXS, 5 microM, but not the parent compound minoxidil, was a potent vasodilator. Whereas the relaxant effects of an another K+ channel opener vasodilator, BRL-34915 (cromakalim), were removed by washing with physiological saline solution, the effects of MNXS persisted after repeated washout attempts. Furthermore, after an initial exposure of segments of intact RMA to [35S] MNXS, greater than 30% of the radiolabel was retained 2 hr after removal of the drug. In contrast, retention of radiolabel was not detected with either [3H]MNXS (label on the piperidine ring of MNXS) or [3H]minoxidil (each less than 3% after a 2-hr washout). These data suggested that the sulfate moiety from MNXS was closely associated with the vascular tissue. To determine if proteins were the acceptors of sulfate from MNXS, intact RMAs were incubated with [35S]MNXS, and then 35S-labeled proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and analyzed by fluorography. Preferential labeling of a 116 kD protein was detected by 2 and 5 min of treatment. A 43 kD protein (resembling actin) also showed significant labeling. A similar profile of 35S-labeled proteins was observed in [35S] MNXS-treated A7r5 rat aortic smooth muscle cells, suggesting that the majority of proteins labeled by [35S]MNXS in intact RMA were components of smooth muscle cells

Integrating protein structures and precomputed genealogies in the Magnum database: Examples with cellular retinoid binding proteins

Directory of Open Access Journals (Sweden)

Bradley Michael E

2006-02-01

Full Text Available Abstract Background When accurate models for the divergent evolution of protein sequences are integrated with complementary biological information, such as folded protein structures, analyses of the combined data often lead to new hypotheses about molecular physiology. This represents an excellent example of how bioinformatics can be used to guide experimental research. However, progress in this direction has been slowed by the lack of a publicly available resource suitable for general use. Results The precomputed Magnum database offers a solution to this problem for ca. 1,800 full-length protein families with at least one crystal structure. The Magnum deliverables include 1 multiple sequence alignments, 2 mapping of alignment sites to crystal structure sites, 3 phylogenetic trees, 4 inferred ancestral sequences at internal tree nodes, and 5 amino acid replacements along tree branches. Comprehensive evaluations revealed that the automated procedures used to construct Magnum produced accurate models of how proteins divergently evolve, or genealogies, and correctly integrated these with the structural data. To demonstrate Magnum's capabilities, we asked for amino acid replacements requiring three nucleotide substitutions, located at internal protein structure sites, and occurring on short phylogenetic tree branches. In the cellular retinoid binding protein family a site that potentially modulates ligand binding affinity was discovered. Recruitment of cellular retinol binding protein to function as a lens crystallin in the diurnal gecko afforded another opportunity to showcase the predictive value of a browsable database containing branch replacement patterns integrated with protein structures. Conclusion We integrated two areas of protein science, evolution and structure, on a large scale and created a precomputed database, known as Magnum, which is the first freely available resource of its kind. Magnum provides evolutionary and structural

Angiotensin-2-mediated Ca2+ signaling in the retinal pigment epithelium: role of angiotensin-receptor-associated-protein and TRPV2 channel.

Directory of Open Access Journals (Sweden)

Rene Barro-Soria

Full Text Available Angiotensin II (AngII receptor (ATR is involved in pathologic local events such as neovascularisation and inflammation including in the brain and retina. The retinal pigment epithelium (RPE expresses ATR in its AT1R form, angiotensin-receptor-associated protein (Atrap, and transient-receptor-potential channel-V2 (TRPV2. AT1R and Atrap co-localize to the basolateral membrane of the RPE, as shown by immunostaining. Stimulation of porcine RPE (pRPE cells by AngII results in biphasic increases in intracellular free Ca(2+inhibited by losartan. Xestospongin C (xest C and U-73122, blockers of IP3R and PLC respectively, reduced AngII-evoked Ca(2+response. RPE cells from Atrap(-/- mice showed smaller AngII-evoked Ca(2+peak (by 22% and loss of sustained Ca(2+elevation compared to wild-type. The TRPV channel activator cannabidiol (CBD at 15 µM stimulates intracellular Ca(2+-rise suggesting that porcine RPE cells express TRPV2 channels. Further evidence supporting the functional expression of TRPV2 channels comes from experiments in which 100 µM SKF96365 (a TRPV channel inhibitor reduced the cannabidiol-induced Ca(2+-rise. Application of SKF96365 or reduction of TRPV2 expression by siRNA reduced the sustained phase of AngII-mediated Ca(2+transients by 53%. Thus systemic AngII, an effector of the local renin-angiotensin system stimulates biphasic Ca(2+transients in the RPE by releasing Ca(2+from cytosolic IP3-dependent stores and activating ATR/Atrap and TRPV2 channels to generate a sustained Ca(2+elevation.

Protein self-assembly and lipid binding in the folding of the potassium channel KcsA

NARCIS (Netherlands)

Barrera, F.N.; Renard, M.L.; Poveda, J.A.; de Kruijff, B.; Killian, J.A.; González-Ros, J.M.

2008-01-01

Moderate concentrations of the alcohol 2,2,2-trifluoroethanol (TFE) cause the coupled unfolding and dissociation into subunits of the homotetrameric potassium channel KcsA, in a process that is partially irreversible when the protein is solubilized in plain dodecyl â-D-maltoside (DDM) micelles

Assembling filamentous phage occlude pIV channels.

Science.gov (United States)

Marciano, D K; Russel, M; Simon, S M

2001-07-31

Filamentous phage f1 is exported from its Escherichia coli host without killing the bacterial cell. Phage-encoded protein pIV, which is required for phage assembly and secretion, forms large highly conductive channels in the outer membrane of E. coli. It has been proposed that the phage are extruded across the bacterial outer membrane through pIV channels. To test this prediction, we developed an in vivo assay by using a mutant pIV that functions in phage export but whose channel opens in the absence of phage extrusion. In E. coli lacking its native maltooligosacharride transporter LamB, this pIV variant allowed oligosaccharide transport across the outer membrane. This entry of oligosaccharide was decreased by phage production and still further decreased by production of phage that cannot be released from the cell surface. Thus, exiting phage block the pIV-dependent entry of oligosaccharide, suggesting that phage occupy the lumen of pIV channels. This study provides the first evidence, to our knowledge, for viral exit through a large aqueous channel.

Four crystal forms of a Bence-Jones protein

Energy Technology Data Exchange (ETDEWEB)

Makino, Debora L.; Henschen-Edman, Agnes H.; McPherson, Alexander, E-mail: amcphers@uci.edu [Molecular Biology and Biochemistry, University of California, Irvine, 560 Steinhaus Hall, Irvine, CA 92697-3900 (United States)

2005-01-01

Four crystal forms have been grown and characterized by X-ray diffraction of a Bence-Jones protein collected from the urine of a multiple myeloma patient more than 40 y ago. The trigonal crystal form may shed some light on the formation of fibrils common to certain storage diseases. Four crystal forms have been grown and characterized by X-ray diffraction of a Bence-Jones protein collected from the urine of a multiple myeloma patient more than 40 years ago. Closely related tetragonal and orthorhombic forms belonging to space groups P4{sub 3}2{sub 1}2 and P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = b = 68.7, c = 182.1 and a = 67.7, b = 69.4, c = 87.3 Å, diffract to 1.5 and 1.9 Å, respectively. Two closely related trigonal forms, both belonging to space group P3{sub 1}21 with unit-cell parameters a = b = 154.3 Å but differing by a doubling of the c axis, one 46.9 Å and the other 94.0 Å, diffract to 2.9 and 2.6 Å resolution, respectively. The trigonal crystal of short c-axis length shows a positive indication of twinning. The trigonal crystal of longer c axis, which appeared only after eight months of incubation at room temperature, is likely to be composed of proteolytically degraded molecules and unlike the other crystal forms contains two entire Bence-Jones dimers in the asymmetric unit. This latter crystal form may shed some light on the formation of fibrils common to certain storage diseases.

Comparative architecture of octahedral protein cages. I. Indexed enclosing forms

Science.gov (United States)

Janner, A.

2008-07-01

The architecture of four protein cages (bacterio ferritin, human mitochondrial ferritin, sulfur oxygenase reductase and small heat-shock protein) are compared top-to-bottom, starting from polyhedra with vertices at cubic lattice points enclosing the cage down to indexed polyhedral forms of single monomers.

Lipid bilayer regulation of membrane protein function: gramicidin channels as molecular force probes

DEFF Research Database (Denmark)

Lundbæk, Jens August; Collingwood, S.A.; Ingolfsson, H.I.

2010-01-01

with collective physical properties (e.g. thickness, intrinsic monolayer curvature or elastic moduli). Studies in physico-chemical model systems have demonstrated that changes in bilayer physical properties can regulate membrane protein function by altering the energetic cost of the bilayer deformation associated...... with a protein conformational change. This type of regulation is well characterized, and its mechanistic elucidation is an interdisciplinary field bordering on physics, chemistry and biology. Changes in lipid composition that alter bilayer physical properties (including cholesterol, polyunsaturated fatty acids...... channels as molecular force probes for studying this mechanism, with a unique ability to discriminate between consequences of changes in monolayer curvature and bilayer elastic moduli....

The ε-form of the differential equations for Feynman integrals in the elliptic case

Science.gov (United States)

Adams, Luise; Weinzierl, Stefan

2018-06-01

Feynman integrals are easily solved if their system of differential equations is in ε-form. In this letter we show by the explicit example of the kite integral family that an ε-form can even be achieved, if the Feynman integrals do not evaluate to multiple polylogarithms. The ε-form is obtained by a (non-algebraic) change of basis for the master integrals.

Diversity and evolution of ABC proteins in mycorrhiza-forming fungi.

Science.gov (United States)

Kovalchuk, Andriy; Kohler, Annegret; Martin, Francis; Asiegbu, Fred O

2015-12-28

Transporter proteins are predicted to have an important role in the mycorrhizal symbiosis, due to the fact that this type of an interaction between plants and fungi requires a continuous nutrient and signalling exchange. ABC transporters are one of the large groups of transporter proteins found both in plants and in fungi. The crucial role of plant ABC transporters in the formation of the mycorrhizal symbiosis has been demonstrated recently. Some of the fungal ABC transporter-encoding genes are also induced during the mycorrhiza formation. However, no experimental evidences of the direct involvement of fungal ABC transporters in this process are available so far. To facilitate the identification of fungal ABC proteins with a potential role in the establishment of the mycorrhizal symbiosis, we have performed an inventory of the ABC protein-encoding genes in the genomes of 25 species of mycorrhiza-forming fungi. We have identified, manually annotated and curated more than 1300 gene models of putative ABC protein-encoding genes. Out of those, more than 1000 models are predicted to encode functional proteins, whereas about 300 models represent gene fragments or putative pseudogenes. We have also performed the phylogenetic analysis of the identified sequences. The sets of ABC proteins in the mycorrhiza-forming species were compared to the related saprotrophic or plant-pathogenic fungal species. Our results demonstrate the high diversity of ABC genes in the genomes of mycorrhiza-forming fungi. Via comparison of transcriptomics data from different species, we have identified candidate groups of ABC transporters that might have a role in the process of the mycorrhiza formation. Results of our inventory will facilitate the identification of fungal transporters with a role in the mycorrhiza formation. We also provide the first data on ABC protein-coding genes for the phylum Glomeromycota and for orders Pezizales, Atheliales, Cantharellales and Sebacinales, contributing to

Discovery of functional monoclonal antibodies targeting G-protein-coupled receptors and ion channels.

Science.gov (United States)

Wilkinson, Trevor C I

2016-06-15

The development of recombinant antibody therapeutics is a significant area of growth in the pharmaceutical industry with almost 50 approved monoclonal antibodies on the market in the US and Europe. Despite this growth, however, certain classes of important molecular targets have remained intractable to therapeutic antibodies due to complexity of the target molecules. These complex target molecules include G-protein-coupled receptors and ion channels which represent a large potential target class for therapeutic intervention with monoclonal antibodies. Although these targets have typically been addressed by small molecule approaches, the exquisite specificity of antibodies provides a significant opportunity to provide selective modulation of these target proteins. Given this opportunity, substantial effort has been applied to address the technical challenges of targeting these complex membrane proteins with monoclonal antibodies. In this review recent progress made in the strategies for discovery of functional monoclonal antibodies for these challenging membrane protein targets is addressed. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

A mother cell-to-forespore channel: current understanding and future challenges.

Science.gov (United States)

Crawshaw, Adam D; Serrano, Mónica; Stanley, Will A; Henriques, Adriano O; Salgado, Paula S

2014-09-01

Formation of endospores allows some bacteria to survive extreme nutrient limitation. The resulting dormant cell, the spore, persists in the environment and is highly resistant to physical and chemical stresses. During spore formation, cells divide asymmetrically and the mother cell engulfs the developing spore, encasing it within a double membrane and isolating it from the medium. Communication between mother cell and isolated forespore involves a specialised connection system that allows nurturing of the forespore and continued macromolecular synthesis, required to finalise spore maturation. Here, we review current understanding of this feeding channel formed by a forespore protein, SpoIIQ, and a mother cell protein, SpoIIIAH, in the model organism Bacillus subtilis and the important human pathogen Clostridium difficile. We also analyse the presence of this channel across endospore-forming bacteria and highlight the main questions still remaining. © 2014 The Authors FEMS Microbiology Letters published by John Wiley & Sons Ltd on behalf of Federation of European Microbiological Societies.

Cytoplasmic Domain of MscS Interacts with Cell Division Protein FtsZ: A Possible Non-Channel Function of the Mechanosensitive Channel in Escherichia Coli.

Directory of Open Access Journals (Sweden)

Piotr Koprowski

Full Text Available Bacterial mechano-sensitive (MS channels reside in the inner membrane and are considered to act as emergency valves whose role is to lower cell turgor when bacteria enter hypo-osmotic environments. However, there is emerging evidence that members of the Mechano-sensitive channel Small (MscS family play additional roles in bacterial and plant cell physiology. MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel. Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein. We identify point mutations in the MscS C-terminal domain that reduce binding to FtsZ and show that bacteria expressing these mutants are compromised in growth on sublethal concentrations of β-lactam antibiotics. Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

Cytoplasmic Domain of MscS Interacts with Cell Division Protein FtsZ: A Possible Non-Channel Function of the Mechanosensitive Channel in Escherichia Coli.

Science.gov (United States)

Koprowski, Piotr; Grajkowski, Wojciech; Balcerzak, Marcin; Filipiuk, Iwona; Fabczak, Hanna; Kubalski, Andrzej

2015-01-01

Bacterial mechano-sensitive (MS) channels reside in the inner membrane and are considered to act as emergency valves whose role is to lower cell turgor when bacteria enter hypo-osmotic environments. However, there is emerging evidence that members of the Mechano-sensitive channel Small (MscS) family play additional roles in bacterial and plant cell physiology. MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel. Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein. We identify point mutations in the MscS C-terminal domain that reduce binding to FtsZ and show that bacteria expressing these mutants are compromised in growth on sublethal concentrations of β-lactam antibiotics. Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

Use of the Le Chatelier principle in calculating the uniform flux in channels formed by packed rods

International Nuclear Information System (INIS)

Skrebkov, G.P.; Lozhkin, S.N.

1986-01-01

A method is proposed for calculating the hydrodynamics of a uniform flow in channels with a cross section of complex form. The method takes into account the anisotropy of the momentum transfer. The anisotropy coefficient of the momentum transfer is determined by using the Le Chatelier principle in a virtual process of transition to the kinematic structure of a uniform flux in equilibrium with a specified set of external conditions which include the channel geometry, wall roughness, and the value of the piezometric gradient

Pado, a fluorescent protein with proton channel activity can optically monitor membrane potential, intracellular pH, and map gap junctions.

Science.gov (United States)

Kang, Bok Eum; Baker, Bradley J

2016-04-04

An in silico search strategy was developed to identify potential voltage-sensing domains (VSD) for the development of genetically encoded voltage indicators (GEVIs). Using a conserved charge distribution in the S2 α-helix, a single in silico search yielded most voltage-sensing proteins including voltage-gated potassium channels, voltage-gated calcium channels, voltage-gated sodium channels, voltage-gated proton channels, and voltage-sensing phosphatases from organisms ranging from mammals to bacteria and plants. A GEVI utilizing the VSD from a voltage-gated proton channel identified from that search was able to optically report changes in membrane potential. In addition this sensor was capable of manipulating the internal pH while simultaneously reporting that change optically since it maintains the voltage-gated proton channel activity of the VSD. Biophysical characterization of this GEVI, Pado, demonstrated that the voltage-dependent signal was distinct from the pH-dependent signal and was dependent on the movement of the S4 α-helix. Further investigation into the mechanism of the voltage-dependent optical signal revealed that inhibiting the dimerization of the fluorescent protein greatly reduced the optical signal. Dimerization of the FP thereby enabled the movement of the S4 α-helix to mediate a fluorescent response.

A fully integrated, monolithic, cryogenic charge sensitive preamplifier using N-channel JFETs and polysilicon resistors

International Nuclear Information System (INIS)

Jung, T.S.; Guckel, H.; Seefeldt, J.; Ott, G.; Ahn, Y.C.

1994-01-01

In this paper, an integrated charge preamplifier to be used with small (10--30 mm 2 ) Si(Li) and Ge(Li) X-ray detectors is described. The preamplifier is designed to operate at cryogenic temperatures (∼100 K to 160 K) for the best performance. An N-channel JFET process technology for integrated charge sensitive preamplifiers has been developed. The process integrates multiple pinch-off voltage JFETs fabricated in an n-type epitaxial layer on a low resistivity p-type substrate. The process also incorporates polysilicon resistors integrated on the same die as the JFETs. The optimized polysilicon resistors exhibit 1/f noise nearly as good as metal film resistors at the same current. Results for integrated amplifier are discussed

Cryogenic microwave channelized receiver

International Nuclear Information System (INIS)

Rauscher, C.; Pond, J.M.; Tait, G.B.

1996-01-01

The channelized receiver being presented demonstrates the use of high temperature superconductor technology in a microwave system setting where superconductor, microwave-monolithic-integrated-circuit, and hybrid-integrated-circuit components are united in one package and cooled to liquid-nitrogen temperatures. The receiver consists of a superconducting X-band four-channel demultiplexer with 100-MHz-wide channels, four commercial monolithically integrated mixers, and four custom-designed hybrid-circuit detectors containing heterostructure ramp diodes. The composite receiver unit has been integrated into the payload of the second-phase NRL high temperature superconductor space experiment (HTSSE-II). Prior to payload assembly, the response characteristics of the receiver were measured as functions of frequency, temperature, and drive levels. The article describes the circuitry, discusses the key issues related to design and implementation, and summarizes the experimental results

Cardiac sodium channel Na(v)1.5 interacts with and is regulated by the protein tyrosine phosphatase PTPH1

DEFF Research Database (Denmark)

Jespersen, Thomas; Gavillet, Bruno; van Bemmelen, Miguel X

2006-01-01

In order to identify proteins interacting with the cardiac voltage-gated sodium channel Na(v)1.5, we used the last 66 amino acids of the C-terminus of the channel as bait to screen a human cardiac cDNA library. We identified the protein tyrosine phosphatase PTPH1 as an interacting protein. Pull......-down experiments confirmed the interaction, and indicated that it depends on the PDZ-domain binding motif of Na(v)1.5. Co-expression experiments in HEK293 cells showed that PTPH1 shifts the Na(v)1.5 availability relationship toward hyperpolarized potentials, whereas an inactive PTPH1 or the tyrosine kinase Fyn...... does the opposite. The results of this study suggest that tyrosine phosphorylation destabilizes the inactivated state of Na(v)1.5....

A linearization of quantum channels

Science.gov (United States)

Crowder, Tanner

2015-06-01

Because the quantum channels form a compact, convex set, we can express any quantum channel as a convex combination of extremal channels. We give a Euclidean representation for the channels whose inverses are also valid channels; these are a subset of the extreme points. They form a compact, connected Lie group, and we calculate its Lie algebra. Lastly, we calculate a maximal torus for the group and provide a constructive approach to decomposing any invertible channel into a product of elementary channels.

Nucleic acid and protein extraction from electropermeabilized E. coli cells on a microfluidic chip

DEFF Research Database (Denmark)

Matos, T.; Senkbeil, Silja; Mendonça, A.

2013-01-01

technique has been developed which is based on exposing E. coli cells to low voltages to allow extraction of nucleic acids and proteins. The flow-through electropermeability chip used consists of a microfluidic channel with integrated gold electrodes that promote cell envelope channel formation at low...

A new essential protein discovery method based on the integration of protein-protein interaction and gene expression data

Directory of Open Access Journals (Sweden)

Li Min

2012-03-01

Full Text Available Abstract Background Identification of essential proteins is always a challenging task since it requires experimental approaches that are time-consuming and laborious. With the advances in high throughput technologies, a large number of protein-protein interactions are available, which have produced unprecedented opportunities for detecting proteins' essentialities from the network level. There have been a series of computational approaches proposed for predicting essential proteins based on network topologies. However, the network topology-based centrality measures are very sensitive to the robustness of network. Therefore, a new robust essential protein discovery method would be of great value. Results In this paper, we propose a new centrality measure, named PeC, based on the integration of protein-protein interaction and gene expression data. The performance of PeC is validated based on the protein-protein interaction network of Saccharomyces cerevisiae. The experimental results show that the predicted precision of PeC clearly exceeds that of the other fifteen previously proposed centrality measures: Degree Centrality (DC, Betweenness Centrality (BC, Closeness Centrality (CC, Subgraph Centrality (SC, Eigenvector Centrality (EC, Information Centrality (IC, Bottle Neck (BN, Density of Maximum Neighborhood Component (DMNC, Local Average Connectivity-based method (LAC, Sum of ECC (SoECC, Range-Limited Centrality (RL, L-index (LI, Leader Rank (LR, Normalized α-Centrality (NC, and Moduland-Centrality (MC. Especially, the improvement of PeC over the classic centrality measures (BC, CC, SC, EC, and BN is more than 50% when predicting no more than 500 proteins. Conclusions We demonstrate that the integration of protein-protein interaction network and gene expression data can help improve the precision of predicting essential proteins. The new centrality measure, PeC, is an effective essential protein discovery method.

Oxidized Low-density Lipoprotein (ox-LDL) Cholesterol Induces the Expression of miRNA-223 and L-type Calcium Channel Protein in Atrial Fibrillation

Science.gov (United States)

He, Fengping; Xu, Xin; Yuan, Shuguo; Tan, Liangqiu; Gao, Lingjun; Ma, Shaochun; Zhang, Shebin; Ma, Zhanzhong; Jiang, Wei; Liu, Fenglian; Chen, Baofeng; Zhang, Beibei; Pang, Jungang; Huang, Xiuyan; Weng, Jiaqiang

2016-08-01

Atrial fibrillation (AF) is the most common sustained arrhythmia causing high morbidity and mortality. While changing of the cellular calcium homeostasis plays a critical role in AF, the L-type calcium channel α1c protein has suggested as an important regulator of reentrant spiral dynamics and is a major component of AF-related electrical remodeling. Our computational modeling predicted that miRNA-223 may regulate the CACNA1C gene which encodes the cardiac L-type calcium channel α1c subunit. We found that oxidized low-density lipoprotein (ox-LDL) cholesterol significantly up-regulates both the expression of miRNA-223 and L-type calcium channel protein. In contrast, knockdown of miRNA-223 reduced L-type calcium channel protein expression, while genetic knockdown of endogenous miRNA-223 dampened AF vulnerability. Transfection of miRNA-223 by adenovirus-mediated expression enhanced L-type calcium currents and promoted AF in mice while co-injection of a CACNA1C-specific miR-mimic counteracted the effect. Taken together, ox-LDL, as a known factor in AF-associated remodeling, positively regulates miRNA-223 transcription and L-type calcium channel protein expression. Our results implicate a new molecular mechanism for AF in which miRNA-223 can be used as an biomarker of AF rheumatic heart disease.

Emerging role of the KCNT1 Slack channel in intellectual disability.

Science.gov (United States)

Kim, Grace E; Kaczmarek, Leonard K

2014-01-01

The sodium-activated potassium KNa channels Slack and Slick are encoded by KCNT1 and KCNT2, respectively. These channels are found in neurons throughout the brain, and are responsible for a delayed outward current termed I KNa. These currents integrate into shaping neuronal excitability, as well as adaptation in response to maintained stimulation. Abnormal Slack channel activity may play a role in Fragile X syndrome, the most common cause for intellectual disability and inherited autism. Slack channels interact directly with the fragile X mental retardation protein (FMRP) and I KNa is reduced in animal models of Fragile X syndrome that lack FMRP. Human Slack mutations that alter channel activity can also lead to intellectual disability, as has been found for several childhood epileptic disorders. Ongoing research is elucidating the relationship between mutant Slack channel activity, development of early onset epilepsies and intellectual impairment. This review describes the emerging role of Slack channels in intellectual disability, coupled with an overview of the physiological role of neuronal I KNa currents.

Franchising as an Integrated Form of Business Organization

Directory of Open Access Journals (Sweden)

Goncharenko Margaryta L.

2018-03-01

Full Text Available The article is aimed at substantiating the possibilities of using franchising for effective integration of enterprises in the market economy. A study on the essence of franchising relations with consideration of their historical formation and the current legal representation was carried out. Features of such relationships for franchisor and franchisees in the form of two business models of franchising together with different types of payments for franchise have been provided. It has been identified that franchising allows for integrative relations, which are established between the representatives of big and small business over a certain period of time, forming for each party some positive and negative moments of interaction. It is proposed to carry out an analysis of economic problems leading to delays in the formation of franchising schemes at both the State and regional levels.

Plan form changes of Gumara River channel over 50 years (Upper Blue Nile basin, Ethiopia)

Science.gov (United States)

Abate, Mengiste; Nyssen, Jan; Mehari, Michael

2014-05-01

Channel plan form changes were investigated along the 65 km long Gumara River in Lake Tana basin (Ethiopia) by overlaying information from aerial photographs and SPOT imagery. Two sets of aerial photographs (1957 and 1980) were scanned, and then orthorectified in ENVI 4.2 environment. Recent channel plan form information was extracted from SPOT images of 2006. ERDAS 2010 and ArcGIS 10.1 tools were used for the data preparation and analysis. The information on river plan form changes spans from 1957 to 2006 (49 years), during which time the Gumara catchment has been subjected to changes in land use/cover and increasing water abstraction, which may have affected its hydrogeomorphology. The results indicated that the lower reach of Gumara at its mouth has undergone major plan form changes. A delta of 1.12 km² was created between 1957 and 1980 and additional 1.00 km² land has been created between 1980 and 2006. The sinuosity of the plan form changed only slightly through the study period: 1.78 in 1957, 1.76 in 1980, and 1.81 in 2006. Comparison of cross sections at the hydrological gauging station showed that the river bed aggraded in the order of 1.5 m to 2.5 m for the period 1963-2009. The trend analysis of stream flow of Gumara River versus rainfall in the catchment also indicated that the bed level of the Gumara river at its gauging station has risen. From field observations, the impact of direct human interventions was identified. The building of artificial levees along the river banks has contributed to huge deposition in the river bed. At locations where intensive irrigation takes place in the floodplain, seepage water through the banks created river bank failure and modifications in plan form. The unstable segments of the river reach were identified and will be further analysed.

Role of TRP channels in the cardiovascular system.

Science.gov (United States)

Yue, Zhichao; Xie, Jia; Yu, Albert S; Stock, Jonathan; Du, Jianyang; Yue, Lixia

2015-02-01

The transient receptor potential (TRP) superfamily consists of a large number of nonselective cation channels with variable degree of Ca(2+)-permeability. The 28 mammalian TRP channel proteins can be grouped into six subfamilies: canonical, vanilloid, melastatin, ankyrin, polycystic, and mucolipin TRPs. The majority of these TRP channels are expressed in different cell types including both excitable and nonexcitable cells of the cardiovascular system. Unlike voltage-gated ion channels, TRP channels do not have a typical voltage sensor, but instead can sense a variety of other stimuli including pressure, shear stress, mechanical stretch, oxidative stress, lipid environment alterations, hypertrophic signals, and inflammation products. By integrating multiple stimuli and transducing their activity to downstream cellular signal pathways via Ca(2+) entry and/or membrane depolarization, TRP channels play an essential role in regulating fundamental cell functions such as contraction, relaxation, proliferation, differentiation, and cell death. With the use of targeted deletion and transgenic mouse models, recent studies have revealed that TRP channels are involved in numerous cellular functions and play an important role in the pathophysiology of many diseases in the cardiovascular system. Moreover, several TRP channels are involved in inherited diseases of the cardiovascular system. This review presents an overview of current knowledge concerning the physiological functions of TRP channels in the cardiovascular system and their contributions to cardiovascular diseases. Ultimately, TRP channels may become potential therapeutic targets for cardiovascular diseases. Copyright © 2015 the American Physiological Society.

Alternative function for the mitochondrial SAM complex in biogenesis of alpha-helical TOM proteins.

Science.gov (United States)

Stojanovski, Diana; Guiard, Bernard; Kozjak-Pavlovic, Vera; Pfanner, Nikolaus; Meisinger, Chris

2007-12-03

The mitochondrial outer membrane contains two preprotein translocases: the general translocase of outer membrane (TOM) and the beta-barrel-specific sorting and assembly machinery (SAM). TOM functions as the central entry gate for nuclear-encoded proteins. The channel-forming Tom40 is a beta-barrel protein, whereas all Tom receptors and small Tom proteins are membrane anchored by a transmembrane alpha-helical segment in their N- or C-terminal portion. Synthesis of Tom precursors takes place in the cytosol, and their import occurs via preexisting TOM complexes. The precursor of Tom40 is then transferred to SAM for membrane insertion and assembly. Unexpectedly, we find that the biogenesis of alpha-helical Tom proteins with a membrane anchor in the C-terminal portion is SAM dependent. Each SAM protein is necessary for efficient membrane integration of the receptor Tom22, whereas assembly of the small Tom proteins depends on Sam37. Thus, the substrate specificity of SAM is not restricted to beta-barrel proteins but also includes the majority of alpha-helical Tom proteins.

Wiring assembly and method of forming a channel in a wiring assembly for receiving conductor and providing separate regions of conductor contact with the channel

Energy Technology Data Exchange (ETDEWEB)

Stelzer, Gerald; Meinke, Rainer; Senti, Mark

2018-03-06

A conductor assembly and method for constructing an assembly of the type which, when conducting current, generates a magnetic field or which, in the presence of a changing magnetic field, induces a voltage. In one embodiment the method provides a first insulative layer tubular in shape and including a surface along which a conductor segment may be positioned. A channel formed in the surface of the insulative layer defines a first conductor path and includes a surface of first contour in cross section along a first plane transverse to the conductor path. A segment of conductor having a surface of second contour in cross section is positioned at least partly in the channel and extends along the conductor path. Along the first plane, contact between the conductor surface of second contour and the channel surface of first contour includes at least two separate regions of contact.

Ion Permeation and Mechanotransduction Mechanisms of Mechanosensitive Piezo Channels.

Science.gov (United States)

Zhao, Qiancheng; Wu, Kun; Geng, Jie; Chi, Shaopeng; Wang, Yanfeng; Zhi, Peng; Zhang, Mingmin; Xiao, Bailong

2016-03-16

Piezo proteins have been proposed as the long-sought-after mechanosensitive cation channels in mammals that play critical roles in various mechanotransduction processes. However, the molecular bases that underlie their ion permeation and mechanotransduction have remained functionally undefined. Here we report our finding of the miniature pore-forming module of Piezo1 that resembles the pore architecture of other trimeric channels and encodes the essential pore properties. We further identified specific residues within the pore module that determine unitary conductance, pore blockage and ion selectivity for divalent and monovalent cations and anions. The non-pore-containing region of Piezo1 confers mechanosensitivity to mechano-insensitive trimeric acid-sensing ion channels, demonstrating that Piezo1 channels possess intrinsic mechanotransduction modules separate from their pore modules. In conclusion, this is the first report on the bona fide pore module and mechanotransduction components of Piezo channels, which define their ion-conducting properties and gating by mechanical stimuli, respectively. Copyright © 2016 Elsevier Inc. All rights reserved.

Association of cardiac myosin binding protein-C with the ryanodine receptor channel: putative retrograde regulation?

Science.gov (United States)

Stanczyk, Paulina J; Seidel, Monika; White, Judith; Viero, Cedric; George, Christopher H; Zissimopoulos, Spyros; Lai, F Anthony

2018-06-21

The cardiac muscle ryanodine receptor-Ca 2+ release channel (RyR2) constitutes the sarcoplasmic reticulum (SR) Ca 2+ efflux mechanism that initiates myocyte contraction, while cardiac myosin binding protein-C (cMyBP-C) mediates regulation of acto-myosin cross-bridge cycling. In this report, we provide the first evidence for the presence of direct interaction between these two proteins, forming a RyR2:cMyBP-C complex. The C-terminus of cMyBP-C binds with the RyR2 N-terminus in mammalian cells and is not mediated by a fibronectin-like domain. Notably, we detected complex formation between both recombinant cMyBP-C and RyR2, as well as with the native proteins in cardiac tissue. Cellular Ca 2+ dynamics in HEK293 cells is altered upon co-expression of cMyBP-C and RyR2, with lowered frequency of RyR2-mediated spontaneous Ca 2+ oscillations, suggesting cMyBP-C exerts a potential inhibitory effect on RyR2-dependent Ca 2+ release. Discovery of a functional RyR2 association with cMyBP-C provides direct evidence for a putative mechanistic link between cytosolic soluble cMyBP-C and SR-mediated Ca 2+ release, via RyR2. Importantly, this interaction may have clinical relevance to the observed cMyBP-C and RyR2 dysfunction in cardiac pathologies, such as hypertrophic cardiomyopathy. © 2018. Published by The Company of Biologists Ltd.

High throughput platforms for structural genomics of integral membrane proteins.

Science.gov (United States)

Mancia, Filippo; Love, James

2011-08-01

Structural genomics approaches on integral membrane proteins have been postulated for over a decade, yet specific efforts are lagging years behind their soluble counterparts. Indeed, high throughput methodologies for production and characterization of prokaryotic integral membrane proteins are only now emerging, while large-scale efforts for eukaryotic ones are still in their infancy. Presented here is a review of recent literature on actively ongoing structural genomics of membrane protein initiatives, with a focus on those aimed at implementing interesting techniques aimed at increasing our rate of success for this class of macromolecules. Copyright © 2011 Elsevier Ltd. All rights reserved.

Structure of the cleavage-activated prefusion form of the parainfluenza virus 5 fusion protein.

Science.gov (United States)

Welch, Brett D; Liu, Yuanyuan; Kors, Christopher A; Leser, George P; Jardetzky, Theodore S; Lamb, Robert A

2012-10-09

The paramyxovirus parainfluenza virus 5 (PIV5) enters cells by fusion of the viral envelope with the plasma membrane through the concerted action of the fusion (F) protein and the receptor binding protein hemagglutinin-neuraminidase. The F protein folds initially to form a trimeric metastable prefusion form that is triggered to undergo large-scale irreversible conformational changes to form the trimeric postfusion conformation. It is thought that F refolding couples the energy released with membrane fusion. The F protein is synthesized as a precursor (F0) that must be cleaved by a host protease to form a biologically active molecule, F1,F2. Cleavage of F protein is a prerequisite for fusion and virus infectivity. Cleavage creates a new N terminus on F1 that contains a hydrophobic region, known as the FP, which intercalates target membranes during F protein refolding. The crystal structure of the soluble ectodomain of the uncleaved form of PIV5 F is known; here we report the crystal structure of the cleavage-activated prefusion form of PIV5 F. The structure shows minimal movement of the residues adjacent to the protease cleavage site. Most of the hydrophobic FP residues are buried in the uncleaved F protein, and only F103 at the newly created N terminus becomes more solvent-accessible after cleavage. The conformational freedom of the charged arginine residues that compose the protease recognition site increases on cleavage of F protein.

Molecular determinants of voltage-gated sodium channel regulation by the Nedd4/Nedd4-like proteins

DEFF Research Database (Denmark)

Rougier, Jean-Sébastien; van Bemmelen, Miguel X; Bruce, M Christine

2004-01-01

-ubiquitin ligases of the Nedd4 family. We recently reported that cardiac Na(v)1.5 is regulated by Nedd4-2. In this study, we further investigated the molecular determinants of regulation of Na(v) proteins. When expressed in HEK-293 cells and studied using whole cell voltage clamping, the neuronal Na(v)1.2 and Na...... that Nedd4-dependent ubiquitination of Na(v) channels may represent a general mechanism regulating the excitability of neurons and myocytes via modulation of channel density at the plasma membrane....

Structure of haze forming proteins in white wines: Vitis vinifera thaumatin-like proteins.

Science.gov (United States)

Marangon, Matteo; Van Sluyter, Steven C; Waters, Elizabeth J; Menz, Robert I

2014-01-01

Grape thaumatin-like proteins (TLPs) play roles in plant-pathogen interactions and can cause protein haze in white wine unless removed prior to bottling. Different isoforms of TLPs have different hazing potential and aggregation behavior. Here we present the elucidation of the molecular structures of three grape TLPs that display different hazing potential. The three TLPs have very similar structures despite belonging to two different classes (F2/4JRU is a thaumatin-like protein while I/4L5H and H2/4MBT are VVTL1), and having different unfolding temperatures (56 vs. 62°C), with protein F2/4JRU being heat unstable and forming haze, while I/4L5H does not. These differences in properties are attributable to the conformation of a single loop and the amino acid composition of its flanking regions.

Ion Channel Conformation and Oligomerization Assessment by Site-Directed Spin Labeling and Pulsed-EPR.

Science.gov (United States)

Pliotas, Christos

2017-01-01

Mechanosensitive (MS) ion channels are multimeric integral membrane proteins that respond to increased lipid bilayer tension by opening their nonselective pores to release solutes and relieve increased cytoplasmic pressure. These systems undergo major conformational changes during gating and the elucidation of their mechanism requires a deep understanding of the interplay between lipids and proteins. Lipids are responsible for transmitting lateral tension to MS channels and therefore play a key role in obtaining a molecular-detail model for mechanosensation. Site-directed spin labeling combined with electron paramagnetic resonance (EPR) spectroscopy is a powerful spectroscopic tool in the study of proteins. The main bottleneck for its use relates to challenges associated with successful isolation of the protein of interest, introduction of paramagnetic labels on desired sites, and access to specialized instrumentation and expertise. The design of sophisticated experiments, which combine a variety of existing EPR methodologies to address a diversity of specific questions, require knowledge of the limitations and strengths, characteristic of each particular EPR method. This chapter is using the MS ion channels as paradigms and focuses on the application of different EPR techniques to ion channels, in order to investigate oligomerization, conformation, and the effect of lipids on their regulation. The methodology we followed, from the initial strategic selection of mutants and sample preparation, including protein purification, spin labeling, reconstitution into lipid mimics to the complete set-up of the pulsed-EPR experiments, is described in detail. © 2017 Elsevier Inc. All rights reserved.

Channel Choice: A Literature Review

DEFF Research Database (Denmark)

Østergaard Madsen, Christian; Kræmmergaard, Pernille

2015-01-01

The channel choice branch of e-government studies citizens’ and businesses’ choice of channels for interacting with government, and how government organizations can integrate channels and migrate users towards the most cost-efficient channels. In spite of the valuable contributions offered...

FORMING OF THE SUSTAINABLE SUPPLY CHAINS BASED ON INCREASING OF THEIR INTEGRATED ENERGY EFFICIENCY

Directory of Open Access Journals (Sweden)

SHVETS V. J.

2016-03-01

Full Text Available Purpose. Key challenges and barriers of mining machinery enterprises in the context of forming sustainable supply chains in the mining industry are specified. Methodology. The algorithm of forming the industrial relations, which are carried out by the enterprises of mining machinery in order to increase the sustainable development of a mining industry, is offered. Conclusions.. Modern supply chains are confronted with dynamic trends and developments that are hard to predict. In order to respond to these changes and remain competitive advantage, companies should be able to identify and understand new sustainability issues in their branches. It is established that in the conditions of globalization of the market and the general orientation of Ukraine’s economy to the European principles of development, for mining machinery enterprises a proof of the commitment and observance of requirements of energy efficiency, i.e. energy saving, energy safety and social energy responsibility are very important. Supply channels should increase not only energy efficiency, economic and production stability of the company, but also strengthen the stability of all participants in the production chain, as well as other stakeholders (business partners, customers, investors, local communities, etc.. Originality. The interrelation of an indicator of energy efficiency in forming sustainable supply chain in a mining industry is established. Practical value. Using the principles of sustainable development in the forming of value chains will improve energy efficiency of industry in general; provide a competitive advantage to entry into the global production chains in terms of European integration of Ukraine.

Predicting protein-protein interactions in Arabidopsis thaliana through integration of orthology, gene ontology and co-expression

Directory of Open Access Journals (Sweden)

Vandepoele Klaas

2009-06-01

Full Text Available Abstract Background Large-scale identification of the interrelationships between different components of the cell, such as the interactions between proteins, has recently gained great interest. However, unraveling large-scale protein-protein interaction maps is laborious and expensive. Moreover, assessing the reliability of the interactions can be cumbersome. Results In this study, we have developed a computational method that exploits the existing knowledge on protein-protein interactions in diverse species through orthologous relations on the one hand, and functional association data on the other hand to predict and filter protein-protein interactions in Arabidopsis thaliana. A highly reliable set of protein-protein interactions is predicted through this integrative approach making use of existing protein-protein interaction data from yeast, human, C. elegans and D. melanogaster. Localization, biological process, and co-expression data are used as powerful indicators for protein-protein interactions. The functional repertoire of the identified interactome reveals interactions between proteins functioning in well-conserved as well as plant-specific biological processes. We observe that although common mechanisms (e.g. actin polymerization and components (e.g. ARPs, actin-related proteins exist between different lineages, they are active in specific processes such as growth, cancer metastasis and trichome development in yeast, human and Arabidopsis, respectively. Conclusion We conclude that the integration of orthology with functional association data is adequate to predict protein-protein interactions. Through this approach, a high number of novel protein-protein interactions with diverse biological roles is discovered. Overall, we have predicted a reliable set of protein-protein interactions suitable for further computational as well as experimental analyses.

Fundamental Characteristics of AAA+ Protein Family Structure and Function.

Science.gov (United States)

Miller, Justin M; Enemark, Eric J

2016-01-01

Many complex cellular events depend on multiprotein complexes known as molecular machines to efficiently couple the energy derived from adenosine triphosphate hydrolysis to the generation of mechanical force. Members of the AAA+ ATPase superfamily (ATPases Associated with various cellular Activities) are critical components of many molecular machines. AAA+ proteins are defined by conserved modules that precisely position the active site elements of two adjacent subunits to catalyze ATP hydrolysis. In many cases, AAA+ proteins form a ring structure that translocates a polymeric substrate through the central channel using specialized loops that project into the central channel. We discuss the major features of AAA+ protein structure and function with an emphasis on pivotal aspects elucidated with archaeal proteins.

Integrated continuous processing of proteins expressed as inclusion bodies: GCSF as a case study.

Science.gov (United States)

Kateja, Nikhil; Agarwal, Harshit; Hebbi, Vishwanath; Rathore, Anurag S

2017-07-01

Affordability of biopharmaceuticals continues to be a challenge, particularly in developing economies. This has fuelled advancements in manufacturing that can offer higher productivity and better economics without sacrificing product quality in the form of an integrated continuous manufacturing platform. While platform processes for monoclonal antibodies have existed for more than a decade, development of an integrated continuous manufacturing process for bacterial proteins has received relatively scant attention. In this study, we propose an end-to-end integrated continuous downstream process (from inclusion bodies to unformulated drug substance) for a therapeutic protein expressed in Escherichia coli as inclusion body. The final process consisted of a continuous refolding in a coiled flow inverter reactor directly coupled to a three-column periodic counter-current chromatography for capture of the product followed by a three-column con-current chromatography for polishing. The continuous bioprocessing train was run uninterrupted for 26 h to demonstrate its capability and the resulting output was analyzed for the various critical quality attributes, namely product purity (>99%), high molecular weight impurities (<0.5%), host cell proteins (<100 ppm), and host cell DNA (<10 ppb). All attributes were found to be consistent over the period of operation. The developed assembly offers smaller facility footprint, higher productivity, fewer hold steps, and significantly higher equipment and resin utilization. The complexities of process integration in the context of continuous processing have been highlighted. We hope that the study presented here will promote development of highly efficient, universal, end-to-end, fully continuous platforms for manufacturing of biotherapeutics. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:998-1009, 2017. © 2016 American Institute of Chemical Engineers.

Ca2+-dependent K+ Channels in Exocrine Salivary Glands

Science.gov (United States)

Catalán, Marcelo A.; Peña-Munzenmayer, Gaspar; Melvin, James E.

2014-01-01

In the last 15 years, remarkable progress has been realized in identifying the genes that encode the ion-transporting proteins involved in exocrine gland function, including salivary glands. Among these proteins, Ca2+-dependent K+ channels take part in key functions including membrane potential regulation, fluid movement and K+ secretion in exocrine glands. Two K+ channels have been identified in exocrine salivary glands: 1) a Ca2+-activated K+ channel of intermediate single channel conductance encoded by the KCNN4 gene; and, 2) a voltage- and Ca2+-dependent K+ channel of large single channel conductance encoded by the KCNMA1 gene. This review focuses on the physiological roles of Ca2+-dependent K+ channels in exocrine salivary glands. We also discuss interesting recent findings on the regulation of Ca2+-dependent K+ channels by protein-protein interactions that may significantly impact exocrine gland physiology. PMID:24559652

Sub-Nanometer Channels Embedded in Two-Dimensional Materials

KAUST Repository

Han, Yimo

2017-07-31

Two-dimensional (2D) materials are among the most promising candidates for next-generation electronics due to their atomic thinness, allowing for flexible transparent electronics and ultimate length scaling1. Thus far, atomically-thin p-n junctions2-7, metal-semiconductor contacts8-10, and metal-insulator barriers11-13 have been demonstrated. While 2D materials achieve the thinnest possible devices, precise nanoscale control over the lateral dimensions are also necessary. Although external one-dimensional (1D) carbon nanotubes14 can be used to locally gate 2D materials, this adds a non-trivial third dimension, complicating device integration and flexibility. Here, we report the direct synthesis of sub-nanometer 1D MoS2 channels embedded within WSe2 monolayers, using a dislocation-catalyzed approach. The 1D channels have edges free of misfit dislocations and dangling bonds, forming a coherent interface with the embedding 2D matrix. Periodic dislocation arrays produce 2D superlattices of coherent MoS2 1D channels in WSe2. Molecular dynamics (MD) simulations have identified other combinations of 2D materials that could form 1D channels. Density function theory (DFT) calculation predicts these 1D channels display type II band alignment needed for carrier confinement and charge separation to access the ultimate length scales necessary for future electronic applications.

Association of lipids with integral membrane surface proteins of Mycoplasma hyorhinis

International Nuclear Information System (INIS)

Bricker, T.M.; Boyer, M.J.; Keith, J.; Watson-McKown, R.; Wise, K.S.

1988-01-01

Triton X-114 (TX-114)-phase fractionation was used to identify and characterize integral membrane surface proteins of the wall-less procaryote Mycoplasma hyorhinis GDL. Phase fractionation of mycoplasmas followed by analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed selective partitioning of approximately 30 [ 35 S]methionine-labeled intrinsic membrane proteins into the TX-114 phase. Similar analysis of [ 3 H]palmitate-labeled cells showed that approximately 20 proteins of this organism were associated with lipid, all of which also efficiently partitioned as integral membrane components into the detergent phase. Immunoblotting and immunoprecipitation of TX-114-phase proteins from 125 I-surface-labeled cells with four monoclonal antibodies to distinct surface epitopes of M. hyorhinis identified surface proteins p120, p70, p42, and p23 as intrinsic membrane components. Immunoprecipitation of [ 3 H]palmitate-labeled TX-114-phase proteins further established that surface proteins p120, p70, and p23 (a molecule that mediates complement-dependent mycoplasmacidal monoclonal antibody activity) were among the lipid-associated proteins of this organism. Two of these proteins, p120 and p123, were acidic (pI less than or equal to 4.5), as shown by two-dimensional isoelectric focusing. This study established that M. hyorhinis contains an abundance of integral membrane proteins tightly associated with lipids and that many of these proteins are exposed at the external surface of the single limiting plasma membrane. Monoclonal antibodies are reported that will allow detailed analysis of the structure and processing of lipid-associated mycoplasma proteins

Influence of the Communication Channel on the Forms of Impoliteness in Company-Customer Interactions

OpenAIRE

Schwab, Pierre-Nicolas; Rosier, Laurence

2015-01-01

The present study aims to examine verbal violence in companies’ answers sent in response to customers’ complaints through two different channels: online (on a public forum) vs. offline (by postal mail). We draw on a recent body of marketing literature pertaining to employees’ dysfunctional behaviors, as well as on conceptualizations of impoliteness, to analyze which role the communication method plays on the forms of impoliteness taken in those interactions.The online dataset of impolite answ...

Identifying protein complex by integrating characteristic of core-attachment into dynamic PPI network.

Directory of Open Access Journals (Sweden)

Xianjun Shen

Full Text Available How to identify protein complex is an important and challenging task in proteomics. It would make great contribution to our knowledge of molecular mechanism in cell life activities. However, the inherent organization and dynamic characteristic of cell system have rarely been incorporated into the existing algorithms for detecting protein complexes because of the limitation of protein-protein interaction (PPI data produced by high throughput techniques. The availability of time course gene expression profile enables us to uncover the dynamics of molecular networks and improve the detection of protein complexes. In order to achieve this goal, this paper proposes a novel algorithm DCA (Dynamic Core-Attachment. It detects protein-complex core comprising of continually expressed and highly connected proteins in dynamic PPI network, and then the protein complex is formed by including the attachments with high adhesion into the core. The integration of core-attachment feature into the dynamic PPI network is responsible for the superiority of our algorithm. DCA has been applied on two different yeast dynamic PPI networks and the experimental results show that it performs significantly better than the state-of-the-art techniques in terms of prediction accuracy, hF-measure and statistical significance in biology. In addition, the identified complexes with strong biological significance provide potential candidate complexes for biologists to validate.

Hydrophobic interaction between contiguous residues in the S6 transmembrane segment acts as a stimuli integration node in the BK channel

Science.gov (United States)

Carrasquel-Ursulaez, Willy; Contreras, Gustavo F.; Sepúlveda, Romina V.; Aguayo, Daniel; González-Nilo, Fernando

2015-01-01

Large-conductance Ca2+- and voltage-activated K+ channel (BK) open probability is enhanced by depolarization, increasing Ca2+ concentration, or both. These stimuli activate modular voltage and Ca2+ sensors that are allosterically coupled to channel gating. Here, we report a point mutation of a phenylalanine (F380A) in the S6 transmembrane helix that, in the absence of internal Ca2+, profoundly hinders channel opening while showing only minor effects on the voltage sensor active–resting equilibrium. Interpretation of these results using an allosteric model suggests that the F380A mutation greatly increases the free energy difference between open and closed states and uncouples Ca2+ binding from voltage sensor activation and voltage sensor activation from channel opening. However, the presence of a bulky and more hydrophobic amino acid in the F380 position (F380W) increases the intrinsic open–closed equilibrium, weakening the coupling between both sensors with the pore domain. Based on these functional experiments and molecular dynamics simulations, we propose that F380 interacts with another S6 hydrophobic residue (L377) in contiguous subunits. This pair forms a hydrophobic ring important in determining the open–closed equilibrium and, like an integration node, participates in the communication between sensors and between the sensors and pore. Moreover, because of its effects on open probabilities, the F380A mutant can be used for detailed voltage sensor experiments in the presence of permeant cations. PMID:25548136

Voltage-gated calcium channels of Paramecium cilia.

Science.gov (United States)

Lodh, Sukanya; Yano, Junji; Valentine, Megan S; Van Houten, Judith L

2016-10-01

Paramecium cells swim by beating their cilia, and make turns by transiently reversing their power stroke. Reversal is caused by Ca 2+ entering the cilium through voltage-gated Ca 2+ (Ca V ) channels that are found exclusively in the cilia. As ciliary Ca 2+ levels return to normal, the cell pivots and swims forward in a new direction. Thus, the activation of the Ca V channels causes cells to make a turn in their swimming paths. For 45 years, the physiological characteristics of the Paramecium ciliary Ca V channels have been known, but the proteins were not identified until recently, when the P. tetraurelia ciliary membrane proteome was determined. Three Ca V α1 subunits that were identified among the proteins were cloned and confirmed to be expressed in the cilia. We demonstrate using RNA interference that these channels function as the ciliary Ca V channels that are responsible for the reversal of ciliary beating. Furthermore, we show that Pawn (pw) mutants of Paramecium that cannot swim backward for lack of Ca V channel activity do not express any of the three Ca V 1 channels in their ciliary membrane, until they are rescued from the mutant phenotype by expression of the wild-type PW gene. These results reinforce the correlation of the three Ca V channels with backward swimming through ciliary reversal. The PwB protein, found in endoplasmic reticulum fractions, co-immunoprecipitates with the Ca V 1c channel and perhaps functions in trafficking. The PwA protein does not appear to have an interaction with the channel proteins but affects their appearance in the cilia. © 2016. Published by The Company of Biologists Ltd.

Host Proteins Determine MRSA Biofilm Structure and Integrity

DEFF Research Database (Denmark)

Dreier, Cindy; Nielsen, Astrid; Jørgensen, Nis Pedersen

Human extracellular matrix (hECM) proteins aids the initial attachment and initiation of an infection, by specific binding to bacterial cell surface proteins. However, the importance of hECM proteins in structure, integrity and antibiotic resilience of a biofilm is unknown. This study aims...... to determine how specific hECM proteins affect S. aureus USA300 JE2 biofilms. Biofilms were grown in the presence of synovial fluid from rheumatoid arteritis patients to mimic in vivo conditions, where bacteria incorporate hECM proteins into the biofilm matrix. Difference in biofilm structure, with and without...... addition of hECM to growth media, was visualized by confocal laser scanning microscopy. Two enzymatic degradation experiments were used to study biofilm matrix composition and importance of hECM proteins: enzymatic removal of specific hECM proteins from growth media, before biofilm formation, and enzymatic...

Structure of haze forming proteins in white wines: Vitis vinifera thaumatin-like proteins.

Directory of Open Access Journals (Sweden)

Matteo Marangon

Full Text Available Grape thaumatin-like proteins (TLPs play roles in plant-pathogen interactions and can cause protein haze in white wine unless removed prior to bottling. Different isoforms of TLPs have different hazing potential and aggregation behavior. Here we present the elucidation of the molecular structures of three grape TLPs that display different hazing potential. The three TLPs have very similar structures despite belonging to two different classes (F2/4JRU is a thaumatin-like protein while I/4L5H and H2/4MBT are VVTL1, and having different unfolding temperatures (56 vs. 62°C, with protein F2/4JRU being heat unstable and forming haze, while I/4L5H does not. These differences in properties are attributable to the conformation of a single loop and the amino acid composition of its flanking regions.

The Fanconi anemia protein FANCF forms a nuclear complex with FANCA, FANCC and FANCG.

Science.gov (United States)

de Winter, J P; van der Weel, L; de Groot, J; Stone, S; Waisfisz, Q; Arwert, F; Scheper, R J; Kruyt, F A; Hoatlin, M E; Joenje, H

2000-11-01

Fanconi anemia (FA) is a chromosomal instability syndrome associated with a strong predisposition to cancer, particularly acute myeloid leukemia and squamous cell carcinoma. At the cellular level, FA is characterized by spontaneous chromosomal breakage and a unique hypersensitivity to DNA cross-linking agents. Complementation analysis has indicated that at least seven distinct genes are involved in the pathogenesis of FA. Despite the identification of four of these genes (FANCA, FANCC, FANCF and FANCG), the nature of the 'FA pathway' has remained enigmatic, as the FA proteins lack sequence homologies or motifs that could point to a molecular function. To further define this pathway, we studied the subcellular localizations and mutual interactions of the FA proteins, including the recently identified FANCF protein, in human lymphoblasts. FANCF was found predominantly in the nucleus, where it complexes with FANCA, FANCC and FANCG. These interactions were detected in wild-type and FA-D lymphoblasts, but not in lymphoblasts of other FA complementation groups. This implies that each of the FA proteins, except FANCD, is required for these complexes to form. Similarly, we show that the interaction between FANCA and FANCC is restricted to wild-type and FA-D cells. Furthermore, we document the subcellular localization of FANCA and the FANCA/FANCG complex in all FA complementation groups. Our results, along with published data, culminate in a model in which a multi-protein FA complex serves a nuclear function to maintain genomic integrity.

SLITHER: a web server for generating contiguous conformations of substrate molecules entering into deep active sites of proteins or migrating through channels in membrane transporters.

Science.gov (United States)

Lee, Po-Hsien; Kuo, Kuei-Ling; Chu, Pei-Ying; Liu, Eric M; Lin, Jung-Hsin

2009-07-01

Many proteins use a long channel to guide the substrate or ligand molecules into the well-defined active sites for catalytic reactions or for switching molecular states. In addition, substrates of membrane transporters can migrate to another side of cellular compartment by means of certain selective mechanisms. SLITHER (http://bioinfo.mc.ntu.edu.tw/slither/or http://slither.rcas.sinica.edu.tw/) is a web server that can generate contiguous conformations of a molecule along a curved tunnel inside a protein, and the binding free energy profile along the predicted channel pathway. SLITHER adopts an iterative docking scheme, which combines with a puddle-skimming procedure, i.e. repeatedly elevating the potential energies of the identified global minima, thereby determines the contiguous binding modes of substrates inside the protein. In contrast to some programs that are widely used to determine the geometric dimensions in the ion channels, SLITHER can be applied to predict whether a substrate molecule can crawl through an inner channel or a half-channel of proteins across surmountable energy barriers. Besides, SLITHER also provides the list of the pore-facing residues, which can be directly compared with many genetic diseases. Finally, the adjacent binding poses determined by SLITHER can also be used for fragment-based drug design.

The role of oligomerization and cooperative regulation in protein function: the case of tryptophan synthase.

Directory of Open Access Journals (Sweden)

M Qaiser Fatmi

Full Text Available The oligomerization/co-localization of protein complexes and their cooperative regulation in protein function is a key feature in many biological systems. The synergistic regulation in different subunits often enhances the functional properties of the multi-enzyme complex. The present study used molecular dynamics and Brownian dynamics simulations to study the effects of allostery, oligomerization and intermediate channeling on enhancing the protein function of tryptophan synthase (TRPS. TRPS uses a set of α/β-dimeric units to catalyze the last two steps of L-tryptophan biosynthesis, and the rate is remarkably slower in the isolated monomers. Our work shows that without their binding partner, the isolated monomers are stable and more rigid. The substrates can form fairly stable interactions with the protein in both forms when the protein reaches the final ligand-bound conformations. Our simulations also revealed that the α/β-dimeric unit stabilizes the substrate-protein conformation in the ligand binding process, which lowers the conformation transition barrier and helps the protein conformations shift from an open/inactive form to a closed/active form. Brownian dynamics simulations with a coarse-grained model illustrate how protein conformations affect substrate channeling. The results highlight the complex roles of protein oligomerization and the fine balance between rigidity and dynamics in protein function.

Membrane shape modulates transmembrane protein distribution.

Science.gov (United States)

Aimon, Sophie; Callan-Jones, Andrew; Berthaud, Alice; Pinot, Mathieu; Toombes, Gilman E S; Bassereau, Patricia

2014-01-27

Although membrane shape varies greatly throughout the cell, the contribution of membrane curvature to transmembrane protein targeting is unknown because of the numerous sorting mechanisms that take place concurrently in cells. To isolate the effect of membrane shape, we used cell-sized giant unilamellar vesicles (GUVs) containing either the potassium channel KvAP or the water channel AQP0 to form membrane nanotubes with controlled radii. Whereas the AQP0 concentrations in flat and curved membranes were indistinguishable, KvAP was enriched in the tubes, with greater enrichment in more highly curved membranes. Fluorescence recovery after photobleaching measurements showed that both proteins could freely diffuse through the neck between the tube and GUV, and the effect of each protein on membrane shape and stiffness was characterized using a thermodynamic sorting model. This study establishes the importance of membrane shape for targeting transmembrane proteins and provides a method for determining the effective shape and flexibility of membrane proteins. Copyright © 2014 Elsevier Inc. All rights reserved.

Determining Membrane Protein-Lipid Binding Thermodynamics Using Native Mass Spectrometry.

Science.gov (United States)

Cong, Xiao; Liu, Yang; Liu, Wen; Liang, Xiaowen; Russell, David H; Laganowsky, Arthur

2016-04-06

Membrane proteins are embedded in the biological membrane where the chemically diverse lipid environment can modulate their structure and function. However, the thermodynamics governing the molecular recognition and interaction of lipids with membrane proteins is poorly understood. Here, we report a method using native mass spectrometry (MS), to determine thermodynamics of individual ligand binding events to proteins. Unlike conventional methods, native MS can resolve individual ligand binding events and, coupled with an apparatus to control the temperature, determine binding thermodynamic parameters, such as for protein-lipid interactions. We validated our approach using three soluble protein-ligand systems (maltose binding protein, lysozyme, and nitrogen regulatory protein) and obtained similar results to those using isothermal titration calorimetry and surface plasmon resonance. We also determined for the first time the thermodynamics of individual lipid binding to the ammonia channel (AmtB), an integral membrane protein from Escherichia coli. Remarkably, we observed distinct thermodynamic signatures for the binding of different lipids and entropy-enthalpy compensation for binding lipids of variable chain length. Additionally, using a mutant form of AmtB that abolishes a specific phosphatidylglycerol (PG) binding site, we observed distinct changes in the thermodynamic signatures for binding PG, implying these signatures can identify key residues involved in specific lipid binding and potentially differentiate between specific lipid binding sites.

Centrosome proteins form an insoluble perinuclear matrix during muscle cell differentiation

Directory of Open Access Journals (Sweden)

Srsen Vlastimil

2009-04-01

Full Text Available Abstract Background Muscle fibres are formed by elongation and fusion of myoblasts into myotubes. During this differentiation process, the cytoskeleton is reorganized, and proteins of the centrosome re-localize to the surface of the nucleus. The exact timing of this event, and the underlying molecular mechanisms are still poorly understood. Results We performed studies on mouse myoblast cell lines that were induced to differentiate in culture, to characterize the early events of centrosome protein re-localization. We demonstrate that this re-localization occurs already at the single cell stage, prior to fusion into myotubes. Centrosome proteins that accumulate at the nuclear surface form an insoluble matrix that can be reversibly disassembled if isolated nuclei are exposed to mitotic cytoplasm from Xenopus egg extract. Our microscopy data suggest that this perinuclear matrix of centrosome proteins consists of a system of interconnected fibrils. Conclusion Our data provide new insights into the reorganization of centrosome proteins during muscular differentiation, at the structural and biochemical level. Because we observe that centrosome protein re-localization occurs early during differentiation, we believe that it is of functional importance for the reorganization of the cytoskeleton in the differentiation process.

Brownian dynamic study of an enzyme metabolon in the TCA cycle: Substrate kinetics and channeling.

Science.gov (United States)

Huang, Yu-Ming M; Huber, Gary A; Wang, Nuo; Minteer, Shelley D; McCammon, J Andrew

2018-02-01

Malate dehydrogenase (MDH) and citrate synthase (CS) are two pacemaking enzymes involved in the tricarboxylic acid (TCA) cycle. Oxaloacetate (OAA) molecules are the intermediate substrates that are transferred from the MDH to CS to carry out sequential catalysis. It is known that, to achieve a high flux of intermediate transport and reduce the probability of substrate leaking, a MDH-CS metabolon forms to enhance the OAA substrate channeling. In this study, we aim to understand the OAA channeling within possible MDH-CS metabolons that have different structural orientations in their complexes. Three MDH-CS metabolons from native bovine, wild-type porcine, and recombinant sources, published in recent work, were selected to calculate OAA transfer efficiency by Brownian dynamics (BD) simulations and to study, through electrostatic potential calculations, a possible role of charges that drive the substrate channeling. Our results show that an electrostatic channel is formed in the metabolons of native bovine and recombinant porcine enzymes, which guides the oppositely charged OAA molecules passing through the channel and enhances the transfer efficiency. However, the channeling probability in a suggested wild-type porcine metabolon conformation is reduced due to an extended diffusion length between the MDH and CS active sites, implying that the corresponding arrangements of MDH and CS result in the decrease of electrostatic steering between substrates and protein surface and then reduce the substrate transfer efficiency from one active site to another. © 2017 The Protein Society.

Strategic positioning of retail entrepreneurship in marketing channels

Directory of Open Access Journals (Sweden)

Ristić Miloš

2005-01-01

Full Text Available Both the actual market flows and dynamiting of power ratios bring about the revolutionary changes within the marketing strategies and basic management approaches, while completely deranging the traditional value system (i.e. system established through marketing channel members' interaction. What is being altered are the market performance patterns, channel communication standards, and forms of institutional pressure manifestation toward the remainder of members which is being performed by the institution with highest economic and negotiating power. Retail company becomes the leader in development of break-the-rule strategies as well as the mutual revolutionary and innovational processes in marketing channel functioning. The final objectives of initiation of co-revolutionary retail strategy with the remainder of marketing channel members are identification and removal of all non-efficient points in the process of development and the process of adding value at all levels of distribution. In addition, these goals are the advanced management of supply chain and integration of consumers with appropriate marketing channel on a long-term basis.

Inhibition of G protein-activated inwardly rectifying K+ channels by different classes of antidepressants.

Directory of Open Access Journals (Sweden)

Toru Kobayashi

Full Text Available Various antidepressants are commonly used for the treatment of depression and several other neuropsychiatric disorders. In addition to their primary effects on serotonergic or noradrenergic neurotransmitter systems, antidepressants have been shown to interact with several receptors and ion channels. However, the molecular mechanisms that underlie the effects of antidepressants have not yet been sufficiently clarified. G protein-activated inwardly rectifying K(+ (GIRK, Kir3 channels play an important role in regulating neuronal excitability and heart rate, and GIRK channel modulation has been suggested to have therapeutic potential for several neuropsychiatric disorders and cardiac arrhythmias. In the present study, we investigated the effects of various classes of antidepressants on GIRK channels using the Xenopus oocyte expression assay. In oocytes injected with mRNA for GIRK1/GIRK2 or GIRK1/GIRK4 subunits, extracellular application of sertraline, duloxetine, and amoxapine effectively reduced GIRK currents, whereas nefazodone, venlafaxine, mianserin, and mirtazapine weakly inhibited GIRK currents even at toxic levels. The inhibitory effects were concentration-dependent, with various degrees of potency and effectiveness. Furthermore, the effects of sertraline were voltage-independent and time-independent during each voltage pulse, whereas the effects of duloxetine were voltage-dependent with weaker inhibition with negative membrane potentials and time-dependent with a gradual decrease in each voltage pulse. However, Kir2.1 channels were insensitive to all of the drugs. Moreover, the GIRK currents induced by ethanol were inhibited by sertraline but not by intracellularly applied sertraline. The present results suggest that GIRK channel inhibition may reveal a novel characteristic of the commonly used antidepressants, particularly sertraline, and contributes to some of the therapeutic effects and adverse effects.

The Integration of Bacteriorhodopsin Proteins with Semiconductor Heterostructure Devices

Science.gov (United States)

Xu, Jian

2008-03-01

Bioelectronics has emerged as one of the most rapidly developing fields among the active frontiers of interdisciplinary research. A major thrust in this field is aimed at the coupling of the technologically-unmatched performance of biological systems, such as neural and sensing functions, with the well developed technology of microelectronics and optoelectronics. To this end we have studied the integration of a suitably engineered protein, bacteriorhodopsin (BR), with semiconductor optoelectronic devices and circuits. Successful integration will potentially lead to ultrasensitive sensors with polarization selectivity and built-in preprocessing capabilities that will be useful for high speed tracking, motion and edge detection, biological detection, and artificial vision systems. In this presentation we will summarize our progresses in this area, which include fundamental studies on the transient dynamics of photo-induced charge shift in BR and the coupling mechanism at protein-semiconductor interface for effective immobilizing and selectively integrating light sensitive proteins with microelectronic devices and circuits, and the device engineering of BR-transistor-integrated optical sensors as well as their applications in phototransceiver circuits. Work done in collaboration with Pallab Bhattacharya, Jonghyun Shin, Department of Electrical Engineering and Computer Science, University of Michigan, Ann Arbor, MI; Robert R. Birge, Department of Chemistry, University of Connecticut, Storrs, CT 06269; and György V'ar'o, Institute of Biophysics, Biological Research Center of the Hungarian Academy of Science, H-6701 Szeged, Hungary.

A resolvase-like protein is requered for the site-specific integration of the temperate lactococcal bacteriophage TP901-1

DEFF Research Database (Denmark)

Christiansen, Bettina; Brøndsted, Lone; Vogensen, Finn K.

1996-01-01

upstream of attP. The N-terminal 150 to 1180 amino acids of Orf1 showed 38 to 44% similarity to the resolvase group of site-specific integrases, while no similarity to know proteins was found in the C-terminal end. Bacteriophage 'TP901-1 therefore contains a unique integration system that does not resemble...... the Int class of site-specific integrases usually found in temperate bacteriophages. The constructed integration vector, pBC170, integrates into the chromosomal attachment site very efficiently and forms stable transformants with a frequency corresponding to 20% of the transformation efficiency....

Multidimensional models for contaminants dispersion in rivers and channels: hybrid solutions via integral transforms

International Nuclear Information System (INIS)

Barros, Felipe Pereira Jorge de

2004-05-01

The aims of the present work were to use the Generalized Integral Transform Technique (GITT) to solve steady state multidimensional models for contaminants dispersion in rivers and channels, as well as to analyze the reduction of computational costs associated with convection-diffusion models that contains more than one space variable. The main focus of this work is the development of models that include variable coefficients such as variable velocity fields along and across the channel. The mathematical formulations also allow the use of different inlet conditions such as point sources, linear sources and plane sources. Several test cases were simulated and the models were validated numerically and with experimental data taken from the literature. The models were implemented in the symbolic computation platform, Mathematica 4.2. (author)

Alpha-helical hydrophobic polypeptides form proton-selective channels in lipid bilayers

Science.gov (United States)

Oliver, A. E.; Deamer, D. W.

1994-01-01

Proton translocation is important in membrane-mediated processes such as ATP-dependent proton pumps, ATP synthesis, bacteriorhodopsin, and cytochrome oxidase function. The fundamental mechanism, however, is poorly understood. To test the theoretical possibility that bundles of hydrophobic alpha-helices could provide a low energy pathway for ion translocation through the lipid bilayer, polyamino acids were incorporated into extruded liposomes and planar lipid membranes, and proton translocation was measured. Liposomes with incorporated long-chain poly-L-alanine or poly-L-leucine were found to have proton permeability coefficients 5 to 7 times greater than control liposomes, whereas short-chain polyamino acids had relatively little effect. Potassium permeability was not increased markedly by any of the polyamino acids tested. Analytical thin layer chromatography measurements of lipid content and a fluorescamine assay for amino acids showed that there were approximately 135 polyleucine or 65 polyalanine molecules associated with each liposome. Fourier transform infrared spectroscopy indicated that a major fraction of the long-chain hydrophobic peptides existed in an alpha-helical conformation. Single-channel recording in both 0.1 N HCl and 0.1 M KCl was also used to determine whether proton-conducting channels formed in planar lipid membranes (phosphatidylcholine/phosphatidylethanolamine, 1:1). Poly-L-leucine and poly-L-alanine in HCl caused a 10- to 30-fold increase in frequency of conductive events compared to that seen in KCl or by the other polyamino acids in either solution. This finding correlates well with the liposome observations in which these two polyamino acids caused the largest increase in membrane proton permeability but had little effect on potassium permeability. Poly-L-leucine was considerably more conductive than poly-L-alanine due primarily to larger event amplitudes and, to a lesser extent, a higher event frequency. Poly-L-leucine caused two

Regulation of ATP-sensitive K+ channels in insulinoma cells: Activation by somatostatin and protein kinase C and the role of cAMP

International Nuclear Information System (INIS)

De Weille, J.R.; Schmid-Antomarchi, H.; Fosset, M.; Lazdunski, M.

1989-01-01

The actions of somatostatin and of the phorbol ester 4β-phorbol 12-myristate 13-acetate (PMA) were studied in rat insulinoma (RINm5F) cells by electrophysiological and 86 Rb + flux techniques. Both PMA and somatostatin hyperpolarize insulinoma cells by activating ATP-sensitive K + channels. The presence of intracellular GTP is required for the somatostatin effects. PMA- and somatostatin-induced hyperpolarization and channel activity are inhibited by the sulfonylurea glibenclamide. Glibenclamide-sensitive 86 Rb + efflux from insulinoma cells is stimulated by somatostatin in a dose-dependent manner (half maximal effect at 0.7 nM) and abolished by pertussis toxin pretreatment. Mutual roles of a GTP-binding protein, of protein kinase C, and of cAMP in the regulation of ATP-sensitive K + channels are discussed

From membrane tension to channel gating: A principal energy transfer mechanism for mechanosensitive channels.

Science.gov (United States)

Zhang, Xuejun C; Liu, Zhenfeng; Li, Jie

2016-11-01

Mechanosensitive (MS) channels are evolutionarily conserved membrane proteins that play essential roles in multiple cellular processes, including sensing mechanical forces and regulating osmotic pressure. Bacterial MscL and MscS are two prototypes of MS channels. Numerous structural studies, in combination with biochemical and cellular data, provide valuable insights into the mechanism of energy transfer from membrane tension to gating of the channel. We discuss these data in a unified two-state model of thermodynamics. In addition, we propose a lipid diffusion-mediated mechanism to explain the adaptation phenomenon of MscS. © 2016 The Protein Society.

Fabrication of high performance microlenses for an integrated capillary channel electrochromatograph with fluorescence detection

International Nuclear Information System (INIS)

Wendt, J. R.; Warren, M. E.; Sweatt, W. C.; Bailey, C. G.; Matzke, C. M.; Arnold, D. W.; Allerman, A. A.; Carter, T. R.; Asbill, R. E.; Samora, S.

1999-01-01

We describe the microfabrication of an extremely compact optical system as a key element in an integrated capillary channel electrochromatograph with fluorescence detection. The optical system consists of a vertical cavity surface-emitting laser (VCSEL), two high performance microlenses, and a commercial photodetector. The microlenses are multilevel diffractive optics patterned by electron beam lithography and etched by reactive ion etching in fused silica. The design uses substrate-mode propagation within the fused silica substrate. Two generations of optical subsystems are described. The first generation design has a 6 mm optical length and is integrated directly onto the capillary channel-containing substrate. The second generation design separates the optical system onto its own substrate module and the optical path length is further compressed to 3.5 mm. The first generation design has been tested using direct fluorescence detection with a 750 nm VCSEL pumping a 10 -4 M solution of CY-7 dye. The observed signal-to-noise ratio of better than 100:1 demonstrates that the background signal from scattered pump light is low despite the compact size of the optical system and is adequate for system sensitivity requirements. (c) 1999 American Vacuum Society

Closed-form expressions for integrals of MKdV and sine-Gordon maps

International Nuclear Information System (INIS)

Kamp, Peter H van der; Rojas, O; Quispel, G R W

2007-01-01

We present closed-form expressions for approximately N integrals of 2N-dimensional maps. The maps are obtained by travelling wave reductions of the modified Korteweg-de Vries equation and of the sine-Gordon equation, respectively. We provide the integrating factors corresponding to the integrals. Moreover we show how the integrals and the integrating factors relate to the staircase method

Mining disease genes using integrated protein-protein interaction and gene-gene co-regulation information.

Science.gov (United States)

Li, Jin; Wang, Limei; Guo, Maozu; Zhang, Ruijie; Dai, Qiguo; Liu, Xiaoyan; Wang, Chunyu; Teng, Zhixia; Xuan, Ping; Zhang, Mingming

2015-01-01

In humans, despite the rapid increase in disease-associated gene discovery, a large proportion of disease-associated genes are still unknown. Many network-based approaches have been used to prioritize disease genes. Many networks, such as the protein-protein interaction (PPI), KEGG, and gene co-expression networks, have been used. Expression quantitative trait loci (eQTLs) have been successfully applied for the determination of genes associated with several diseases. In this study, we constructed an eQTL-based gene-gene co-regulation network (GGCRN) and used it to mine for disease genes. We adopted the random walk with restart (RWR) algorithm to mine for genes associated with Alzheimer disease. Compared to the Human Protein Reference Database (HPRD) PPI network alone, the integrated HPRD PPI and GGCRN networks provided faster convergence and revealed new disease-related genes. Therefore, using the RWR algorithm for integrated PPI and GGCRN is an effective method for disease-associated gene mining.

Application-specific integrated circuit design for a typical pressurized water reactor pressure channel trip

International Nuclear Information System (INIS)

Battle, R.E.; Manges, W.W.; Emery, M.S.; Vendermolen, R.I.; Bhatt, S.

1994-01-01

This article discusses the use of application-specific integrated circuits (ASICs) in nuclear plant safety systems. ASICs have certain advantages over software-based systems because they can be simple enough to be thoroughly tested, and they can be tailored to replace existing equipment. An architecture to replace a pressurized water reactor pressure channel trip is presented. Methods of implementing digital algorithms are also discussed

Trafficking regulates the subcellular distribution of voltage-gated sodium channels in primary sensory neurons.

Science.gov (United States)

Bao, Lan

2015-09-30

Voltage-gated sodium channels (Navs) comprise at least nine pore-forming α subunits. Of these, Nav1.6, Nav1.7, Nav1.8 and Nav1.9 are the most frequently studied in primary sensory neurons located in the dorsal root ganglion and are mainly localized to the cytoplasm. A large pool of intracellular Navs raises the possibility that changes in Nav trafficking could alter channel function. The molecular mediators of Nav trafficking mainly consist of signals within the Navs themselves, interacting proteins and extracellular factors. The surface expression of Navs is achieved by escape from the endoplasmic reticulum and proteasome degradation, forward trafficking and plasma membrane anchoring, and it is also regulated by channel phosphorylation and ubiquitination in primary sensory neurons. Axonal transport and localization of Navs in afferent fibers involves the motor protein KIF5B and scaffold proteins, including contactin and PDZ domain containing 2. Localization of Nav1.6 to the nodes of Ranvier in myelinated fibers of primary sensory neurons requires node formation and the submembrane cytoskeletal protein complex. These findings inform our understanding of the molecular and cellular mechanisms underlying Nav trafficking in primary sensory neurons.

Integrated synchronous receiver channel for optical instrumentation applications

Science.gov (United States)

Benten, Harold G. P. H.; Ruotsalainen, Tarmo; Maekynen, Anssi J.; Rahkonen, Timo E.; Kopola, Harri K.

1997-09-01

A two-channel synchronous receiver circuit for optical instrumentation applications has been designed and implemented. Each receiver channel comprises a.o. transimpedance preamplifier, voltage amplifiers, programmable feedback networks, and a synchronous detector. The function of the channel is to extract the slowly varying information carrying signal from a modulated carrier which is accompanied by relatively high levels of noise. As a whole, the channel can be characterized as a narrow band filter around the frequency of interest. Medical applications include arterial oxygen saturation (SaO2) measurement and dental pulp vitality measurement. In both cases, two optical signals with different frequencies are received by a single photodiode. The measured performance of the optical receiver shows its suitability for the above mentioned applications. Therefore the circuit will be used in a small sized, battery-operated sensor prototype to test the sensing method in a clinical environment. Other applications include the signal processing of optical position-sensitive detectors. A summary of measured receiver channel performance: input reduced noise current spectral density between 0.20 and 0.30 pA/(root)Hz at all relevant frequencies, total programmable channel transimpedance between 7 M(Omega) and 500 M(Omega) , lower -3 dB frequency of at least 50 Hz, upper -3 dB frequency of 40 kHz, maximum voltage swing at the demodulator output of 2.4 V.

Atomic force microscopy imaging reveals the formation of ASIC/ENaC cross-clade ion channels

International Nuclear Information System (INIS)

Jeggle, Pia; Smith, Ewan St. J.; Stewart, Andrew P.; Haerteis, Silke; Korbmacher, Christoph; Edwardson, J. Michael

2015-01-01

ASIC and ENaC are co-expressed in various cell types, and there is evidence for a close association between them. Here, we used atomic force microscopy (AFM) to determine whether ASIC1a and ENaC subunits are able to form cross-clade hybrid ion channels. ASIC1a and ENaC could be co-isolated from detergent extracts of tsA 201 cells co-expressing the two subunits. Isolated proteins were incubated with antibodies against ENaC and Fab fragments against ASIC1a. AFM imaging revealed proteins that were decorated by both an antibody and a Fab fragment with an angle of ∼120° between them, indicating the formation of ASIC1a/ENaC heterotrimers. - Highlights: • There is evidence for a close association between ASIC and ENaC. • We used AFM to test whether ASIC1a and ENaC subunits form cross-clade ion channels. • Isolated proteins were incubated with subunit-specific antibodies and Fab fragments. • Some proteins were doubly decorated at ∼120° by an antibody and a Fab fragment. • Our results indicate the formation of ASIC1a/ENaC heterotrimers

Atomic force microscopy imaging reveals the formation of ASIC/ENaC cross-clade ion channels

Energy Technology Data Exchange (ETDEWEB)

Jeggle, Pia; Smith, Ewan St. J.; Stewart, Andrew P. [Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1PD (United Kingdom); Haerteis, Silke; Korbmacher, Christoph [Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstrasse 6, 91054 Erlangen (Germany); Edwardson, J. Michael, E-mail: jme1000@cam.ac.uk [Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1PD (United Kingdom)

2015-08-14

ASIC and ENaC are co-expressed in various cell types, and there is evidence for a close association between them. Here, we used atomic force microscopy (AFM) to determine whether ASIC1a and ENaC subunits are able to form cross-clade hybrid ion channels. ASIC1a and ENaC could be co-isolated from detergent extracts of tsA 201 cells co-expressing the two subunits. Isolated proteins were incubated with antibodies against ENaC and Fab fragments against ASIC1a. AFM imaging revealed proteins that were decorated by both an antibody and a Fab fragment with an angle of ∼120° between them, indicating the formation of ASIC1a/ENaC heterotrimers. - Highlights: • There is evidence for a close association between ASIC and ENaC. • We used AFM to test whether ASIC1a and ENaC subunits form cross-clade ion channels. • Isolated proteins were incubated with subunit-specific antibodies and Fab fragments. • Some proteins were doubly decorated at ∼120° by an antibody and a Fab fragment. • Our results indicate the formation of ASIC1a/ENaC heterotrimers.

A computational model of the LGI1 protein suggests a common binding site for ADAM proteins.

Directory of Open Access Journals (Sweden)

Emanuela Leonardi

Full Text Available Mutations of human leucine-rich glioma inactivated (LGI1 gene encoding the epitempin protein cause autosomal dominant temporal lateral epilepsy (ADTLE, a rare familial partial epileptic syndrome. The LGI1 gene seems to have a role on the transmission of neuronal messages but the exact molecular mechanism remains unclear. In contrast to other genes involved in epileptic disorders, epitempin shows no homology with known ion channel genes but contains two domains, composed of repeated structural units, known to mediate protein-protein interactions.A three dimensional in silico model of the two epitempin domains was built to predict the structure-function relationship and propose a functional model integrating previous experimental findings. Conserved and electrostatic charged regions of the model surface suggest a possible arrangement between the two domains and identifies a possible ADAM protein binding site in the β-propeller domain and another protein binding site in the leucine-rich repeat domain. The functional model indicates that epitempin could mediate the interaction between proteins localized to different synaptic sides in a static way, by forming a dimer, or in a dynamic way, by binding proteins at different times.The model was also used to predict effects of known disease-causing missense mutations. Most of the variants are predicted to alter protein folding while several other map to functional surface regions. In agreement with experimental evidence, this suggests that non-secreted LGI1 mutants could be retained within the cell by quality control mechanisms or by altering interactions required for the secretion process.

Regulation of the Water Channel Aquaporin-2 via 14-3-3θ and -ζ

DEFF Research Database (Denmark)

Moeller, Hanne B; Slengerik-Hansen, Joachim; Aroankins, Takwa

2016-01-01

The 14-3-3 family of proteins are multifunctional proteins that interact with many of their cellular targets in a phosphorylation-dependent manner. Here, we determined that 14-3-3 proteins interact with phosphorylated forms of the water channel aquaporin-2 (AQP2) and modulate its function. With t...... levels. In conclusion, this study demonstrates phosphorylation-dependent interactions of AQP2 with 14-3-3 θ and ζ. These interactions play divergent roles in modulating AQP2 trafficking, phosphorylation, ubiquitylation and degradation....

Heterologous Expression of Tulip Petal Plasma Membrane Aquaporins in Pichia pastoris for Water Channel Analysis▿

Science.gov (United States)

Azad, Abul Kalam; Sawa, Yoshihiro; Ishikawa, Takahiro; Shibata, Hitoshi

2009-01-01

Water channels formed by aquaporins (AQPs) play an important role in the control of water homeostasis in individual cells and in multicellular organisms. Plasma membrane intrinsic proteins (PIPs) constitute a subclass of plant AQPs. TgPIP2;1 and TgPIP2;2 from tulip petals are members of the PIP family. In this study, we overexpressed TgPIP2;1 and TgPIP2;2 in Pichia pastoris and monitored their water channel activity (WCA) either by an in vivo spheroplast-bursting assay performed after hypo-osmotic shock or by growth assay. Osmolarity, pH, and inhibitors of AQPs, protein kinases (PKs), and protein phosphatases (PPs) affect the WCA of heterologous AQPs in this expression system. The WCA of TgPIP2;2-expressing spheroplasts was affected by inhibitors of PKs and PPs, which indicates that the water channel of this homologue is regulated by phosphorylation in P. pastoris. From the results reported herein, we suggest that P. pastoris can be employed as a heterologous expression system to assay the WCA of PIPs and to monitor the AQP-mediated channel gating mechanism, and it can be developed to screen inhibitors/effectors of PIPs. PMID:19251885

Heterologous expression of tulip petal plasma membrane aquaporins in Pichia pastoris for water channel analysis.

Science.gov (United States)

Azad, Abul Kalam; Sawa, Yoshihiro; Ishikawa, Takahiro; Shibata, Hitoshi

2009-05-01

Water channels formed by aquaporins (AQPs) play an important role in the control of water homeostasis in individual cells and in multicellular organisms. Plasma membrane intrinsic proteins (PIPs) constitute a subclass of plant AQPs. TgPIP2;1 and TgPIP2;2 from tulip petals are members of the PIP family. In this study, we overexpressed TgPIP2;1 and TgPIP2;2 in Pichia pastoris and monitored their water channel activity (WCA) either by an in vivo spheroplast-bursting assay performed after hypo-osmotic shock or by growth assay. Osmolarity, pH, and inhibitors of AQPs, protein kinases (PKs), and protein phosphatases (PPs) affect the WCA of heterologous AQPs in this expression system. The WCA of TgPIP2;2-expressing spheroplasts was affected by inhibitors of PKs and PPs, which indicates that the water channel of this homologue is regulated by phosphorylation in P. pastoris. From the results reported herein, we suggest that P. pastoris can be employed as a heterologous expression system to assay the WCA of PIPs and to monitor the AQP-mediated channel gating mechanism, and it can be developed to screen inhibitors/effectors of PIPs.

Effect of the I(to) activator NS5806 on cloned K(V)4 channels depends on the accessory protein KChIP2

DEFF Research Database (Denmark)

Lundby, Alicia; Jespersen, Thomas; Schmitt, Nicole

2010-01-01

The compound NS5806 increases the transient outward current (I(to)) in canine ventricular cardiomyocytes and slows current decay. In human and canine ventricle, I(to) is thought to be mediated by K(V)4.3 and various ancillary proteins, yet, the exact subunit composition of I(to) channels is still...... debated. Here we characterize the effect of NS5806 on heterologously expressed putative I(to) channel subunits and other potassium channels....

Traveling Wave-Guide Channels of a New Coupled Integrable Dispersionless System

International Nuclear Information System (INIS)

Souleymanou, Abbagari; Kuetche, Victor K.; Bouetou, Thomas B.; Kofane, Timoleon C.

2012-01-01

In the wake of the recent investigation of new coupled integrable dispersionless equations by means of the Darboux transformation [Zhaqilao, et al., Chin. Phys. B 18 (2009) 1780], we carry out the initial value analysis of the previous system using the fourth-order Runge-Kutta's computational scheme. As a result, while depicting its phase portraits accordingly, we show that the above dispersionless system actually supports two kinds of solutions amongst which the localized traveling wave-guide channels. In addition, paying particular interests to such localized structures, we construct the bilinear transformation of the current system from which scattering amongst the above waves can be deeply studied. (general)

Dynamic nuclear polarization of membrane proteins: covalently bound spin-labels at protein–protein interfaces

International Nuclear Information System (INIS)

Wylie, Benjamin J.; Dzikovski, Boris G.; Pawsey, Shane; Caporini, Marc; Rosay, Melanie; Freed, Jack H.; McDermott, Ann E.

2015-01-01

We demonstrate that dynamic nuclear polarization of membrane proteins in lipid bilayers may be achieved using a novel polarizing agent: pairs of spin labels covalently bound to a protein of interest interacting at an intermolecular interaction surface. For gramicidin A, nitroxide tags attached to the N-terminal intermolecular interface region become proximal only when bimolecular channels forms in the membrane. We obtained signal enhancements of sixfold for the dimeric protein. The enhancement effect was comparable to that of a doubly tagged sample of gramicidin C, with intramolecular spin pairs. This approach could be a powerful and selective means for signal enhancement in membrane proteins, and for recognizing intermolecular interfaces

Na(v)1.8 channelopathy in mutant mice deficient for myelin protein zero is detrimental to motor axons

DEFF Research Database (Denmark)

Moldovan, Mihai; Alvarez Herrero, Susana; Pinchenko, Volodymyr

2011-01-01

Myelin protein zero mutations were found to produce Charcot-Marie-Tooth disease phenotypes with various degrees of myelin impairment and axonal loss, ranging from the mild 'demyelinating' adult form to severe and early onset forms. Protein zero deficient homozygous mice ( ) show a severe and prog......Myelin protein zero mutations were found to produce Charcot-Marie-Tooth disease phenotypes with various degrees of myelin impairment and axonal loss, ranging from the mild 'demyelinating' adult form to severe and early onset forms. Protein zero deficient homozygous mice ( ) show a severe...... and progressive dysmyelinating neuropathy from birth with compromised myelin compaction, hypomyelination and distal axonal degeneration. A previous study using immunofluorescence showed that motor nerves deficient of myelin protein zero upregulate the Na(V)1.8 voltage gated sodium channel isoform, which...... is normally present only in restricted populations of sensory axons. The aim of this study was to investigate the function of motor axons in protein zero-deficient mice with particular emphasis on ectopic Na(V)1.8 voltage gated sodium channel. We combined 'threshold tracking' excitability studies...

Integrating complex functions: coordination of nuclear pore complex assembly and membrane expansion of the nuclear envelope requires a family of integral membrane proteins.

Science.gov (United States)

Schneiter, Roger; Cole, Charles N

2010-01-01

The nuclear envelope harbors numerous large proteinaceous channels, the nuclear pore complexes (NPCs), through which macromolecular exchange between the cytosol and the nucleoplasm occurs. This double-membrane nuclear envelope is continuous with the endoplasmic reticulum and thus functionally connected to such diverse processes as vesicular transport, protein maturation and lipid synthesis. Recent results obtained from studies in Saccharomyces cerevisiae indicate that assembly of the nuclear pore complex is functionally dependent upon maintenance of lipid homeostasis of the ER membrane. Previous work from one of our laboratories has revealed that an integral membrane protein Apq12 is important for the assembly of functional nuclear pores. Cells lacking APQ12 are viable but cannot grow at low temperatures, have aberrant NPCs and a defect in mRNA export. Remarkably, these defects in NPC assembly can be overcome by supplementing cells with a membrane fluidizing agent, benzyl alcohol, suggesting that Apq12 impacts the flexibility of the nuclear membrane, possibly by adjusting its lipid composition when cells are shifted to a reduced temperature. Our new study now expands these findings and reveals that an essential membrane protein, Brr6, shares at least partially overlapping functions with Apq12 and is also required for assembly of functional NPCs. A third nuclear envelope membrane protein, Brl1, is related to Brr6, and is also required for NPC assembly. Because maintenance of membrane homeostasis is essential for cellular survival, the fact that these three proteins are conserved in fungi that undergo closed mitoses, but are not found in metazoans or plants, may indicate that their functions are performed by proteins unrelated at the primary sequence level to Brr6, Brl1 and Apq12 in cells that disassemble their nuclear envelopes during mitosis.

Electrostatic tuning of permeation and selectivity in aquaporin water channels

DEFF Research Database (Denmark)

Jensen, Mogens O Stibius; Tajkhorshid, E.; Schulten, K.

2003-01-01

Water permeation and electrostatic interactions between water and channel are investigated in the Escherichia coli glycerol uptake facilitator GlpF, a member of the aquaporin water channel family, by molecular dynamics simulations. A tetrameric model of the channel embedded in a 16:0/ 18:1c9...... with the protein electrostatic fields enforce a bipolar water configuration inside the channel with dipole inversion at the NPA motifs. At the NPA motifs water-protein electrostatic interactions facilitate this inversion. Furthermore, water-water electrostatic interactions are in all regions inside the channel...... stronger than water-protein interactions, except near a conserved, positively charged Arg residue. We find that variations of the protein electrostatic field through the channel, owing to preserved structural features, completely explain the bipolar orientation of water. This orientation persists despite...

Kv10.1 potassium channel: from the brain to the tumors.

Science.gov (United States)

Cázares-Ordoñez, V; Pardo, L A

2017-10-01

The KCNH1 gene encodes the Kv10.1 (Eag1) ion channel, a member of the EAG (ether-à-go-go) family of voltage-gated potassium channels. Recent studies have demonstrated that KCHN1 mutations are implicated in Temple-Baraitser and Zimmermann-Laband syndromes and other forms of developmental deficits that all present with mental retardation and epilepsy, suggesting that Kv10.1 might be important for cognitive development in humans. Although the Kv10.1 channel is mainly expressed in the mammalian brain, its ectopic expression occurs in 70% of human cancers. Cancer cells and tumors expressing Kv10.1 acquire selective advantages that favor cancer progression through molecular mechanisms that involve several cellular pathways, indicating that protein-protein interactions may be important for Kv10.1 influence in cell proliferation and tumorigenesis. Several studies on transcriptional and post-transcriptional regulation of Kv10.1 expression have shown interesting mechanistic insights about Kv10.1 role in oncogenesis, increasing the importance of identifying the cellular factors that regulate Kv10.1 expression in tumors.

Volume Regulated Channels

DEFF Research Database (Denmark)

Klausen, Thomas Kjær

of volume perturbations evolution have developed system of channels and transporters to tightly control volume homeostasis. In the past decades evidence has been mounting, that the importance of these volume regulated channels and transporters are not restricted to the defense of cellular volume...... but are also essential for a number of physiological processes such as proliferation, controlled cell death, migration and endocrinology. The thesis have been focusing on two Channels, namely the swelling activated Cl- channel (ICl, swell) and the transient receptor potential Vanilloid (TRPV4) channel. I: Cl......- serves a multitude of functions in the mammalian cell, regulating the membrane potential (Em), cell volume, protein activity and the driving force for facilitated transporters giving Cl- and Cl- channels a major potential of regulating cellular function. These functions include control of the cell cycle...

The quadratic-form identity for constructing the Hamiltonian structure of integrable systems

International Nuclear Information System (INIS)

Guo Fukui; Zhang Yufeng

2005-01-01

A usual loop algebra, not necessarily the matrix form of the loop algebra A-tilde n-1 , is also made use of for constructing linear isospectral problems, whose compatibility conditions exhibit a zero-curvature equation from which integrable systems are derived. In order to look for the Hamiltonian structure of such integrable systems, a quadratic-form identity is created in the present paper whose special case is just the trace identity; that is, when taking the loop algebra A-tilde 1 , the quadratic-form identity presented in this paper is completely consistent with the trace identity

Entropic potential field formed for a linear-motor protein near a filament: Statistical-mechanical analyses using simple models.

Science.gov (United States)

Amano, Ken-Ichi; Yoshidome, Takashi; Iwaki, Mitsuhiro; Suzuki, Makoto; Kinoshita, Masahiro

2010-07-28

We report a new progress in elucidating the mechanism of the unidirectional movement of a linear-motor protein (e.g., myosin) along a filament (e.g., F-actin). The basic concept emphasized here is that a potential field is entropically formed for the protein on the filament immersed in solvent due to the effect of the translational displacement of solvent molecules. The entropic potential field is strongly dependent on geometric features of the protein and the filament, their overall shapes as well as details of the polyatomic structures. The features and the corresponding field are judiciously adjusted by the binding of adenosine triphosphate (ATP) to the protein, hydrolysis of ATP into adenosine diphosphate (ADP)+Pi, and release of Pi and ADP. As the first step, we propose the following physical picture: The potential field formed along the filament for the protein without the binding of ATP or ADP+Pi to it is largely different from that for the protein with the binding, and the directed movement is realized by repeated switches from one of the fields to the other. To illustrate the picture, we analyze the spatial distribution of the entropic potential between a large solute and a large body using the three-dimensional integral equation theory. The solute is modeled as a large hard sphere. Two model filaments are considered as the body: model 1 is a set of one-dimensionally connected large hard spheres and model 2 is a double helical structure formed by two sets of connected large hard spheres. The solute and the filament are immersed in small hard spheres forming the solvent. The major findings are as follows. The solute is strongly confined within a narrow space in contact with the filament. Within the space there are locations with sharply deep local potential minima along the filament, and the distance between two adjacent locations is equal to the diameter of the large spheres constituting the filament. The potential minima form a ringlike domain in model 1

A numerical model for meltwater channel evolution in glaciers

Directory of Open Access Journals (Sweden)

A. H. Jarosch

2012-04-01

Full Text Available Meltwater channels form an integral part of the hydrological system of a glacier. Better understanding of how meltwater channels develop and evolve is required to fully comprehend supraglacial and englacial meltwater drainage. Incision of supraglacial stream channels and subsequent roof closure by ice deformation has been proposed in recent literature as a possible englacial conduit formation process. Field evidence for supraglacial stream incision has been found in Svalbard and Nepal. In Iceland, where volcanic activity provides meltwater with temperatures above 0 °C, rapid enlargement of supraglacial channels has been observed. Supraglacial channels provide meltwater through englacial passages to the subglacial hydrological systems of big ice sheets, which in turn affects ice sheet motion and their contribution to eustatic sea level change. By coupling, for the first time, a numerical ice dynamic model to a hydraulic model which includes heat transfer, we investigate the evolution of meltwater channels and their incision behaviour. We present results for different, constant meltwater fluxes, different channel slopes, different meltwater temperatures, different melt rate distributions in the channel as well as temporal variations in meltwater flux. The key parameters governing incision rate and depth are channel slope, meltwater temperature loss to the ice and meltwater flux. Channel width and geometry are controlled by melt rate distribution along the channel wall. Calculated Nusselt numbers suggest that turbulent mixing is the main heat transfer mechanism in the meltwater channels studied.

The effectiveness of styrene-maleic acid (SMA) copolymers for solubilisation of integral membrane proteins from SMA-accessible and SMA-resistant membranes.

Science.gov (United States)

Swainsbury, David J K; Scheidelaar, Stefan; Foster, Nicholas; van Grondelle, Rienk; Killian, J Antoinette; Jones, Michael R

2017-10-01

Solubilisation of biological lipid bilayer membranes for analysis of their protein complement has traditionally been carried out using detergents, but there is increasing interest in the use of amphiphilic copolymers such as styrene maleic acid (SMA) for the solubilisation, purification and characterisation of integral membrane proteins in the form of protein/lipid nanodiscs. Here we survey the effectiveness of various commercially-available formulations of the SMA copolymer in solubilising Rhodobacter sphaeroides reaction centres (RCs) from photosynthetic membranes. We find that formulations of SMA with a 2:1 or 3:1 ratio of styrene to maleic acid are almost as effective as detergent in solubilising RCs, with the best solubilisation by short chain variants (membranes was uniformly low, but could be increased through a variety of treatments to increase the lipid:protein ratio. However, proteins isolated from such membranes comprised clusters of complexes in small membrane patches rather than individual proteins. We conclude that short-chain 2:1 and 3:1 formulations of SMA are the most effective in solubilising integral membrane proteins, but that solubilisation efficiencies are strongly influenced by the size of the target protein and the density of packing of proteins in the membrane. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

[Synopsis about the hypothesis of "information channel" of channel-collateral system].

Science.gov (United States)

Chang, Xi-Lang

2008-10-01

The author of the present paper founded a theorem about the "incompleteness of single channel structure" (nerve, blood vessel, lymphatic, interspace, aperture, etc.) through quantitative and qualitative analysis about the economic information channel in the human body, which eliminates the probability of single channel structure in the information channel of channel (meridian)-collateral system. After comprehensive analysis on the current researches, the author puts forward a neodoxy, i.e., the body "information channel" structure of the channel-collateral system, mainly follows the distribution regularity of systemic statistics, and is not a single specific entity; various layers of the information channel in the main stems of the channel-collaterals are composed of optimized structure tissues. Hence, the structure of this information channel of channel-collateral system is an overall-optimized, sequential and compatible systemic structure. From this neodoxy, the author brings forward a working principle of channel-collaterals, which is supported theoretically by bio-auxology. The longitudinal distribution of the main stems of meridian-collaterals is considered to result from that in the process of the animal evolution, in the animals moving forward, the microscopic complicated movement of intracorporeal information and energy molecules is related to the forward macroscopic and non-uniform movement of organism in trans-measure. Its impulse and kinetic momentum forms a main vector in the longitudinal direction of the body (the direction of the main stem of channel-collaterals). In order to adapt to and utilize natural regularities, the main stems of the channel-collaterals gradually differentiate and evolve in the living organism, forming a whole system. The "hypothesis of biological origin of channel-collateral system" and "that of information channel of the channel-collaterals in the body" constitute a relatively complete theoretical system framework.

LRRK2 regulates voltage-gated calcium channel function.

Directory of Open Access Journals (Sweden)

Cade eBedford

2016-05-01

Full Text Available Voltage-gated Ca2+ (CaV channels enable Ca2+ influx in response to membrane depolarization. CaV2.1 channels are localized to the presynaptic membrane of many types of neurons where they are involved in triggering neurotransmitter release. Several signaling proteins have been identified as important CaV2.1 regulators including protein kinases, G-proteins and Ca2+ binding proteins. Recently, we discovered that leucine rich repeat kinase 2 (LRRK2, a protein associated with inherited Parkinson’s disease, interacts with specific synaptic proteins and influences synaptic transmission. Since synaptic proteins functionally interact with CaV2.1 channels and synaptic transmission is triggered by Ca2+ entry via CaV2.1, we investigated whether LRRK2 could impact CaV2.1 channel function. CaV2.1 channel properties were measured using whole cell patch clamp electrophysiology in HEK293 cells transfected with CaV2.1 subunits and various LRRK2 constructs. Our results demonstrate that both wild type LRRK2 and the G2019S LRRK2 mutant caused a significant increase in whole cell Ca2+ current density compared to cells expressing only the CaV2.1 channel complex. In addition, LRRK2 expression caused a significant hyperpolarizing shift in voltage-dependent activation while having no significant effect on inactivation properties. These functional changes in CaV2.1 activity are likely due to a direct action of LRRK2 as we detected a physical interaction between LRRK2 and the β3 CaV channel subunit via coimmunoprecipitation. Furthermore, effects on CaV2.1 channel function are dependent on LRRK2 kinase activity as these could be reversed via treatment with a LRRK2 inhibitor. Interestingly, LRRK2 also augmented endogenous voltage-gated Ca2+ channel function in PC12 cells suggesting other CaV channels could also be regulated by LRRK2. Overall, our findings support a novel physiological role for LRRK2 in regulating CaV2.1 function that could have implications for how

Modulation of nucleotide sensitivity of ATP-sensitive potassium channels by phosphatidylinositol-4-phosphate 5-kinase.

Science.gov (United States)

Shyng, S L; Barbieri, A; Gumusboga, A; Cukras, C; Pike, L; Davis, J N; Stahl, P D; Nichols, C G

2000-01-18

ATP-sensitive potassium channels (K(ATP) channels) regulate cell excitability in response to metabolic changes. K(ATP) channels are formed as a complex of a sulfonylurea receptor (SURx), a member of the ATP-binding cassette protein family, and an inward rectifier K(+) channel subunit (Kir6.x). Membrane phospholipids, in particular phosphatidylinositol (PI) 4,5-bisphosphate (PIP(2)), activate K(ATP) channels and antagonize ATP inhibition of K(ATP) channels when applied to inside-out membrane patches. To examine the physiological relevance of this regulatory mechanism, we manipulated membrane PIP(2) levels by expressing either the wild-type or an inactive form of PI-4-phosphate 5-kinase (PIP5K) in COSm6 cells and examined the ATP sensitivity of coexpressed K(ATP) channels. Channels from cells expressing the wild-type PIP5K have a 6-fold lower ATP sensitivity (K(1/2), the half maximal inhibitory concentration, approximately 60 microM) than the sensitivities from control cells (K(1/2) approximately 10 microM). An inactive form of the PIP5K had little effect on the K(1/2) of wild-type channels but increased the ATP-sensitivity of a mutant K(ATP) channel that has an intrinsically lower ATP sensitivity (from K(1/2) approximately 450 microM to K(1/2) approximately 100 microM), suggesting a decrease in membrane PIP(2) levels as a consequence of a dominant-negative effect of the inactive PIP5K. These results show that PIP5K activity, which regulates PIP(2) and PI-3,4,5-P(3) levels, is a significant determinant of the physiological nucleotide sensitivity of K(ATP) channels.

Direct effects of ionizing radiation on integral membrane proteins. Noncovalent energy transfer requires specific interpeptide interactions

International Nuclear Information System (INIS)

Jhun, E.; Jhun, B.H.; Jones, L.R.; Jung, C.Y.

1991-01-01

The 12 transmembrane alpha helices (TMHs) of human erythrocyte glucose transporter were individually cut by pepsin digestion as membrane-bound 2.5-3.5-kDa peptide fragments. Radiation-induced chemical degradation of these fragments showed an average target size of 34 kDa. This is 10-12 x larger than the average size of an individual TMH, demonstrating that a significant energy transfer occurs among these TMHs in the absence of covalent linkage. Heating this TMH preparation at 100 degree C for 15 min reduced the target size to 5 kDa or less, suggesting that the noncovalent energy transfer requires specific helix-helix interactions. Purified phospholamban, a small (6-kDa) integral membrane protein containing a single TMH, formed a pentameric assembly in sodium dodecyl sulfate. The chemical degradation target size of this phospholamban pentamer was 5-6 kDa, illustrating that not all integral membrane protein assemblies permit intersubunit energy transfer. These findings together with other published observations suggest strongly that significant noncovalent energy transfer can occur within the tertiary and quaternary structure of membrane proteins and that as yet undefined proper molecular interactions are required for such covalent energy transfer. Our results with pepsin-digested glucose transporter also illustrate the importance of the interhelical interaction as a predominating force in maintaining the tertiary structure of a transmembrane protein

Numerical studies of the polymer melt flow in the extruder screw channel and the forming tool

Science.gov (United States)

Ershov, S. V.; Trufanova, N. M.

2017-06-01

To date, polymer compositions based on polyethylene or PVC is widely used as insulating materials. These materials processing conjugate with a number of problems during selection of the rational extrusion regimes. To minimize the time and cost when determining the technological regime uses mathematical modeling techniques. The paper discusses heat and mass transfer processes in the extruder screw channel, output adapter and the cable head. During the study were determined coefficients for three rheological models based on obtained viscosity vs. shear rate experimental data. Also a comparative analysis of this viscosimetric laws application possibility for studying polymer melt flow during its processing on the extrusion equipment was held. As a result of numerical study the temperature, viscosity and shear rate fields in the extruder screw channel and forming tool were obtained.

Aggregated forms of bull seminal plasma proteins and their heparin-binding activity

Czech Academy of Sciences Publication Activity Database

Jelínková, Petra; Ryšlavá, H.; Liberda, J.; Jonáková, Věra; Tichá, M.

2004-01-01

Roč. 69, - (2004), s. 616-630 ISSN 0010-0765 R&D Projects: GA ČR GA303/02/0433; GA ČR GP303/02/P069; GA MZd NJ7463 Institutional research plan: CEZ:AV0Z5052915; CEZ:MSM 113100001 Keywords : bull seminal plasma proteins * heparin-binding proteins * aggregated forms of proteins Subject RIV: CE - Biochemistry Impact factor: 1.062, year: 2004

Why do alpha-beta parallel proteins, like flavodoxins, form misfolded off-pathway intermediates?

NARCIS (Netherlands)

Nabuurs, S.M.

2009-01-01

The question: “Why do α-β parallel proteins, like flavodoxins, form misfolded off-pathway
intermediates?" is the main subject of this thesis. A. vinelandii apoflavodoxin is chosen as protein
of interest as it is a representative of α-β parallel proteins, which are widely prevalent in

Hole mobility enhancement of p-MOSFETs using global and local Ge-channel technologies

International Nuclear Information System (INIS)

Takagi, Shinichi; Tezuka, T.; Irisawa, T.; Nakaharai, S.; Maeda, T.; Numata, T.; Ikeda, K.; Sugiyama, N.

2006-01-01

Mobility enhancement technologies have currently been recognized as mandatory for future scaled MOSFETs. In this paper, we review our recent results on high hole mobility p-MOSFETs using global/local SiGe or Ge channels. There are two directions for introducing SiGe or Ge channels into Si CMOS platform. One is to use SiGe or Ge global substrates and the other is to form SiGe or Ge-channel regions locally on Si wafers. In both cases, the Ge condensation technique, where Ge-channel layers are formed by oxidizing SiGe films on SOI substrates, are effectively utilized. As for the global technologies, ultrathin GOI substrates are prepared and used to fabricate high mobility GOI p-MOSFETs. As for the local technologies, SGOI or GOI channels are formed locally in the active area of p-MOSFETs on SOI wafers. It is shown that the hole mobility enhancement factor of as high as 10 is obtained in locally fabricated p-MOSFETs through the effects of high-Ge content and the compressive strain. Furthermore, the local Ge-channel technologies are combined with global SiGe or Ge substrates for pursuing the optimal and individual design of n-MOSFETs and p-MOSFETs on a single Si wafer. The CMOS device composed of strained-Si n-MOSFETs and SGOI p-MOSFETs is successfully integrated on a same wafer, which is a promising CMOS structure under deep sub 100 nm technology nodes

Intron retention in mRNA encoding ancillary subunit of insect voltage-gated sodium channel modulates channel expression, gating regulation and drug sensitivity.

Directory of Open Access Journals (Sweden)

Céline M Bourdin

Full Text Available Insect voltage-gated sodium (Nav channels are formed by a well-known pore-forming α-subunit encoded by para-like gene and ancillary subunits related to TipE from the mutation "temperature-induced-paralysis locus E." The role of these ancillary subunits in the modulation of biophysical and pharmacological properties of Na(+ currents are not enough documented. The unique neuronal ancillary subunit TipE-homologous protein 1 of Drosophila melanogaster (DmTEH1 strongly enhances the expression of insect Nav channels when heterologously expressed in Xenopus oocytes. Here we report the cloning and functional expression of two neuronal DmTEH1-homologs of the cockroach, Periplaneta americana, PaTEH1A and PaTEH1B, encoded by a single bicistronic gene. In PaTEH1B, the second exon encoding the last 11-amino-acid residues of PaTEH1A is shifted to 3'UTR by the retention of a 96-bp intron-containing coding-message, thus generating a new C-terminal end. We investigated the gating and pharmacological properties of the Drosophila Nav channel variant (DmNav1-1 co-expressed with DmTEH1, PaTEH1A, PaTEH1B or a truncated mutant PaTEH1Δ(270-280 in Xenopus oocytes. PaTEH1B caused a 2.2-fold current density decrease, concomitant with an equivalent α-subunit incorporation decrease in the plasma membrane, compared to PaTEH1A and PaTEH1Δ(270-280. PaTEH1B positively shifted the voltage-dependences of activation and slow inactivation of DmNav1-1 channels to more positive potentials compared to PaTEH1A, suggesting that the C-terminal end of both proteins may influence the function of the voltage-sensor and the pore of Nav channel. Interestingly, our findings showed that the sensitivity of DmNav1-1 channels to lidocaine and to the pyrazoline-type insecticide metabolite DCJW depends on associated TEH1-like subunits. In conclusion, our work demonstrates for the first time that density, gating and pharmacological properties of Nav channels expressed in Xenopus oocytes can be

Dynamics of ceramide channels detected using a microfluidic system.

Directory of Open Access Journals (Sweden)

Chenren Shao

Full Text Available Ceramide, a proapoptotic sphingolipid, has been shown to form channels, in mitochondrial outer membranes, large enough to translocate proteins. In phospholipid membranes, electrophysiological studies and electron microscopic visualization both report that these channels form in a range of sizes with a modal value of 10 nm in diameter. A hydrogen bonded barrel-like structure consisting of hundreds of ceramide molecules has been proposed for the structure of the channel and this is supported by electrophysiological studies and molecular dynamic simulations. To our knowledge, the mechanical strength and deformability of such a large diameter but extremely thin cylindrical structure has never been reported. Here we present evidence for a reversible mechanical distortion of the cylinder following the addition of La(3+. A microfluidic system was used to repeatedly lower and then restore the conductance by alternatively perfusing La(3+ and EDTA. Although aspects of the kinetics of conductance drop and recovery are consistent with a disassembly/diffusion/reassembly model, others are inconsistent with the expected time scale of lateral diffusion of disassembled channel fragments in the membrane. The presence of a residual conductance following La(3+ treatment and the relationship between the residual conductance and the initial conductance were both indicative of a distortion/recovery process in analogy with a pressure-induced distortion of a flexible cylinder.

IRaPPA: information retrieval based integration of biophysical models for protein assembly selection.

Science.gov (United States)

Moal, Iain H; Barradas-Bautista, Didier; Jiménez-García, Brian; Torchala, Mieczyslaw; van der Velde, Arjan; Vreven, Thom; Weng, Zhiping; Bates, Paul A; Fernández-Recio, Juan

2017-06-15

In order to function, proteins frequently bind to one another and form 3D assemblies. Knowledge of the atomic details of these structures helps our understanding of how proteins work together, how mutations can lead to disease, and facilitates the designing of drugs which prevent or mimic the interaction. Atomic modeling of protein-protein interactions requires the selection of near-native structures from a set of docked poses based on their calculable properties. By considering this as an information retrieval problem, we have adapted methods developed for Internet search ranking and electoral voting into IRaPPA, a pipeline integrating biophysical properties. The approach enhances the identification of near-native structures when applied to four docking methods, resulting in a near-native appearing in the top 10 solutions for up to 50% of complexes benchmarked, and up to 70% in the top 100. IRaPPA has been implemented in the SwarmDock server ( http://bmm.crick.ac.uk/∼SwarmDock/ ), pyDock server ( http://life.bsc.es/pid/pydockrescoring/ ) and ZDOCK server ( http://zdock.umassmed.edu/ ), with code available on request. moal@ebi.ac.uk. Supplementary data are available at Bioinformatics online. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com

Substrate channel in nitrogenase revealed by a molecular dynamics approach.

Science.gov (United States)

Smith, Dayle; Danyal, Karamatullah; Raugei, Simone; Seefeldt, Lance C

2014-04-15

Mo-dependent nitrogenase catalyzes the biological reduction of N2 to two NH3 molecules at FeMo-cofactor buried deep inside the MoFe protein. Access of substrates, such as N2, to the active site is likely restricted by the surrounding protein, requiring substrate channels that lead from the surface to the active site. Earlier studies on crystallographic structures of the MoFe protein have suggested three putative substrate channels. Here, we have utilized submicrosecond atomistic molecular dynamics simulations to allow the nitrogenase MoFe protein to explore its conformational space in an aqueous solution at physiological ionic strength, revealing a putative substrate channel. The viability of this observed channel was tested by examining the free energy of passage of N2 from the surface through the channel to FeMo-cofactor, resulting in the discovery of a very low energy barrier. These studies point to a viable substrate channel in nitrogenase that appears during thermal motions of the protein in an aqueous environment and that approaches a face of FeMo-cofactor earlier implicated in substrate binding.

integrating indigenous and exogenous communication channels

African Journals Online (AJOL)

Prof. Adipala Ekwamu

1Ministry of Agriculture and Food Security, Pesticide Control Board, Malawi. 2Ministry of ... noticed by the authors that local communication channels and folk media play an important role in CBAF, ... traditionally generate, store, share,.

A nonlinear model of flow in meandering submarine and subaerial channels

Science.gov (United States)

Imran, Jasim; Parker, Gary; Pirmez, Carlos

1999-12-01

A generalized model of flow in meandering subaqueous and subaerial channels is developed. The conservation equations of mass and momentum are depth/layer integrated, normalized, and represented as deviations from a straight base state. This allows the determination of integrable forms which can be solved at both linear and nonlinear levels. The effects of various flow and geometric parameters on the flow dynamics are studied. Although the model is not limited to any specific planform, this study focuses on sine-generated curves. In analysing the flow patterns, the turbidity current of the subaqueous case is simplified to a conservative density flow with water entrainment from above neglected. The subaqueous model thus formally corresponds to a subcritical or only mildly supercritical mud-rich turbidity current. By extension, however the analysis can be applied to a depositional or erosional current carrying sand that is changing only slowly in the streamwise direction. By bringing the subaqueous and subaerial cases into a common form, flow behaviour in the two environments can be compared under similar geometric and boundary conditions. A major difference between the two cases is the degree of superelevation of channel flow around bends, which is modest in the subaerial case but substantial in the subaqueous case. Another difference concerns Coriolis effects: some of the largest subaqueous meandering systems are so large that Coriolis effects can become important. The model is applied to meander bends on the youngest channel in the mid-fan region of the Amazon Fan and a mildly sinuous bend of the North-West Atlantic Mid-Ocean Channel. In the absence of specific data on the turbid flows that created the channel, the model can be used to make inferences about the flow, and in particular the range of values of flow velocity and sediment concentration that would allow the growth and downfan migration of meander bends.

Structure, diversity and evolution of protein toxins from spore-forming entomopathogenic bacteria

NARCIS (Netherlands)

Maagd, de R.A.; Bravo, A.; Berry, C.; Crickmore, N.; Schnepf, H.E.

2003-01-01

Gram-positive spore-forming entomopathogenic bacteria can utilize a large variety of protein toxins to help them invade, infect, and finally kill their hosts, through their action on the insect midgut. These toxins belong to a number of homology groups containing a diversity of protein structures

Pannexin-1 channels show distinct morphology and no gap junction characteristics in mammalian cells.

Science.gov (United States)

Beckmann, Anja; Grissmer, Alexander; Krause, Elmar; Tschernig, Thomas; Meier, Carola

2016-03-01

Pannexins (Panx) are proteins with a similar membrane topology to connexins, the integral membrane protein of gap junctions. Panx1 channels are generally of major importance in a large number of system and cellular processes and their function has been thoroughly characterized. In contrast, little is known about channel structure and subcellular distribution. We therefore determine the subcellular localization of Panx1 channels in cultured cells and aim at the identification of channel morphology in vitro. Using freeze-fracture replica immunolabeling on EYFP-Panx1-overexpressing HEK 293 cells, large particles were identified in plasma membranes, which were immunogold-labeled using either GFP or Panx1 antibodies. There was no labeling or particles in the nuclear membranes of these cells, pointing to plasma membrane localization of Panx1-EYFP channels. The assembly of particles was irregular, this being in contrast to the regular pattern of gap junctions. The fact that no counterparts were identified on apposing cells, which would have been indicative of intercellular signaling, supported the idea of Panx1 channels within one membrane. Control cells (transfected with EYFP only, non-transfected) were devoid of both particles and immunogold labeling. Altogether, this study provides the first demonstration of Panx1 channel morphology and assembly in intact cells. The identification of Panx1 channels as large particles within the plasma membrane provides the knowledge required to enable recognition of Panx1 channels in tissues in future studies. Thus, these results open up new avenues for the detailed analysis of the subcellular localization of Panx1 and of its nearest neighbors such as purinergic receptors in vivo.

Voltage-gated lipid ion channels

DEFF Research Database (Denmark)

Blicher, Andreas; Heimburg, Thomas Rainer

2013-01-01

Synthetic lipid membranes can display channel-like ion conduction events even in the absence of proteins. We show here that these events are voltage-gated with a quadratic voltage dependence as expected from electrostatic theory of capacitors. To this end, we recorded channel traces and current...... histograms in patch-experiments on lipid membranes. We derived a theoretical current-voltage relationship for pores in lipid membranes that describes the experimental data very well when assuming an asymmetric membrane. We determined the equilibrium constant between closed and open state and the open...... probability as a function of voltage. The voltage-dependence of the lipid pores is found comparable to that of protein channels. Lifetime distributions of open and closed events indicate that the channel open distribution does not follow exponential statistics but rather power law behavior for long open times...

Gating of human ClC-2 chloride channels and regulation by carboxy-terminal domains.

Science.gov (United States)

Garcia-Olivares, Jennie; Alekov, Alexi; Boroumand, Mohammad Reza; Begemann, Birgit; Hidalgo, Patricia; Fahlke, Christoph

2008-11-15

Eukaryotic ClC channels are dimeric proteins with each subunit forming an individual protopore. Single protopores are gated by a fast gate, whereas the slow gate is assumed to control both protopores through a cooperative movement of the two carboxy-terminal domains. We here study the role of the carboxy-terminal domain in modulating fast and slow gating of human ClC-2 channels, a ubiquitously expressed ClC-type chloride channel involved in transepithelial solute transport and in neuronal chloride homeostasis. Partial truncation of the carboxy-terminus abolishes function of ClC-2 by locking the channel in a closed position. However, unlike other isoforms, its complete removal preserves function of ClC-2. ClC-2 channels without the carboxy-terminus exhibit fast and slow gates that activate and deactivate significantly faster than in WT channels. In contrast to the prevalent view, a single carboxy-terminus suffices for normal slow gating, whereas both domains regulate fast gating of individual protopores. Our findings demonstrate that the carboxy-terminus is not strictly required for slow gating and that the cooperative gating resides in other regions of the channel protein. ClC-2 is expressed in neurons and believed to open at negative potentials and increased internal chloride concentrations after intense synaptic activity. We propose that the function of the ClC-2 carboxy-terminus is to slow down the time course of channel activation in order to stabilize neuronal excitability.

Maternal protein restriction induces alterations in insulin signaling and ATP sensitive potassium channel protein in hypothalami of intrauterine growth restriction fetal rats.

Science.gov (United States)

Liu, Xiaomei; Qi, Ying; Gao, Hong; Jiao, Yisheng; Gu, Hui; Miao, Jianing; Yuan, Zhengwei

2013-01-01

It is well recognized that intrauterine growth restriction leads to the development of insulin resistance and type 2 diabetes mellitus in adulthood. To investigate the mechanisms behind this "metabolic imprinting" phenomenon, we examined the impact of maternal undernutrition on insulin signaling pathway and the ATP sensitive potassium channel expression in the hypothalamus of intrauterine growth restriction fetus. Intrauterine growth restriction rat model was developed through maternal low protein diet. The expression and activated levels of insulin signaling molecules and K(ATP) protein in the hypothalami which were dissected at 20 days of gestation, were analyzed by western blot and real time PCR. The tyrosine phosphorylation levels of the insulin receptor substrate 2 and phosphatidylinositol 3'-kinase p85α in the hypothalami of intrauterine growth restriction fetus were markedly reduced. There was also a downregulation of the hypothalamic ATP sensitive potassium channel subunit, sulfonylurea receptor 1, which conveys the insulin signaling. Moreover, the abundances of gluconeogenesis enzymes were increased in the intrauterine growth restriction livers, though no correlation was observed between sulfonylurea receptor 1 and gluconeogenesis enzymes. Our data suggested that aberrant intrauterine milieu impaired insulin signaling in the hypothalamus, and these alterations early in life might contribute to the predisposition of the intrauterine growth restriction fetus toward the adult metabolic disorders.

Integrated Structural Biology for α-Helical Membrane Protein Structure Determination.

Science.gov (United States)

Xia, Yan; Fischer, Axel W; Teixeira, Pedro; Weiner, Brian; Meiler, Jens

2018-04-03

While great progress has been made, only 10% of the nearly 1,000 integral, α-helical, multi-span membrane protein families are represented by at least one experimentally determined structure in the PDB. Previously, we developed the algorithm BCL::MP-Fold, which samples the large conformational space of membrane proteins de novo by assembling predicted secondary structure elements guided by knowledge-based potentials. Here, we present a case study of rhodopsin fold determination by integrating sparse and/or low-resolution restraints from multiple experimental techniques including electron microscopy, electron paramagnetic resonance spectroscopy, and nuclear magnetic resonance spectroscopy. Simultaneous incorporation of orthogonal experimental restraints not only significantly improved the sampling accuracy but also allowed identification of the correct fold, which is demonstrated by a protein size-normalized transmembrane root-mean-square deviation as low as 1.2 Å. The protocol developed in this case study can be used for the determination of unknown membrane protein folds when limited experimental restraints are available. Copyright © 2018 Elsevier Ltd. All rights reserved.

Signature and Pathophysiology of Non-canonical Pores in Voltage-Dependent Cation Channels.

Science.gov (United States)

Held, Katharina; Voets, Thomas; Vriens, Joris

2016-01-01

Opening and closing of voltage-gated cation channels allows the regulated flow of cations such as Na(+), K(+), and Ca(2+) across cell membranes, which steers essential physiological processes including shaping of action potentials and triggering Ca(2+)-dependent processes. Classical textbooks describe the voltage-gated cation channels as membrane proteins with a single, central aqueous pore. In recent years, however, evidence has accumulated for the existence of additional ion permeation pathways in this group of cation channels, distinct from the central pore, which here we collectively name non-canonical pores. Whereas the first non-canonical pores were unveiled only after making specific point mutations in the voltage-sensor region of voltage-gated Na(+) and K(+) channels, recent evidence indicates that they may also be functional in non-mutated channels. Moreover, several channelopathies have been linked to mutations that cause the appearance of a non-canonical ion permeation pathway as a new pathological mechanism. This review provides an integrated overview of the biophysical properties of non-canonical pores described in voltage-dependent cation channels (KV, NaV, Cav, Hv1, and TRPM3) and of the (patho)physiological impact of opening of such pores.

Free-energy relationships in ion channels activated by voltage and ligand

Science.gov (United States)

Chowdhury, Sandipan

2013-01-01

Many ion channels are modulated by multiple stimuli, which allow them to integrate a variety of cellular signals and precisely respond to physiological needs. Understanding how these different signaling pathways interact has been a challenge in part because of the complexity of underlying models. In this study, we analyzed the energetic relationships in polymodal ion channels using linkage principles. We first show that in proteins dually modulated by voltage and ligand, the net free-energy change can be obtained by measuring the charge-voltage (Q-V) relationship in zero ligand condition and the ligand binding curve at highly depolarizing membrane voltages. Next, we show that the voltage-dependent changes in ligand occupancy of the protein can be directly obtained by measuring the Q-V curves at multiple ligand concentrations. When a single reference ligand binding curve is available, this relationship allows us to reconstruct ligand binding curves at different voltages. More significantly, we establish that the shift of the Q-V curve between zero and saturating ligand concentration is a direct estimate of the interaction energy between the ligand- and voltage-dependent pathway. These free-energy relationships were tested by numerical simulations of a detailed gating model of the BK channel. Furthermore, as a proof of principle, we estimate the interaction energy between the ligand binding and voltage-dependent pathways for HCN2 channels whose ligand binding curves at various voltages are available. These emerging principles will be useful for high-throughput mutagenesis studies aimed at identifying interaction pathways between various regulatory domains in a polymodal ion channel. PMID:23250866

Semantic integration to identify overlapping functional modules in protein interaction networks

Directory of Open Access Journals (Sweden)

Ramanathan Murali

2007-07-01

Full Text Available Abstract Background The systematic analysis of protein-protein interactions can enable a better understanding of cellular organization, processes and functions. Functional modules can be identified from the protein interaction networks derived from experimental data sets. However, these analyses are challenging because of the presence of unreliable interactions and the complex connectivity of the network. The integration of protein-protein interactions with the data from other sources can be leveraged for improving the effectiveness of functional module detection algorithms. Results We have developed novel metrics, called semantic similarity and semantic interactivity, which use Gene Ontology (GO annotations to measure the reliability of protein-protein interactions. The protein interaction networks can be converted into a weighted graph representation by assigning the reliability values to each interaction as a weight. We presented a flow-based modularization algorithm to efficiently identify overlapping modules in the weighted interaction networks. The experimental results show that the semantic similarity and semantic interactivity of interacting pairs were positively correlated with functional co-occurrence. The effectiveness of the algorithm for identifying modules was evaluated using functional categories from the MIPS database. We demonstrated that our algorithm had higher accuracy compared to other competing approaches. Conclusion The integration of protein interaction networks with GO annotation data and the capability of detecting overlapping modules substantially improve the accuracy of module identification.

Cholesterol-Binding Sites in GIRK Channels: The Devil is in the Details.

Science.gov (United States)

Rosenhouse-Dantsker, Avia

2018-01-01

In recent years, it has become evident that cholesterol plays a direct role in the modulation of a variety of ion channels. In most cases, cholesterol downregulates channel activity. In contrast, our earlier studies have demonstrated that atrial G protein inwardly rectifying potassium (GIRK) channels are upregulated by cholesterol. Recently, we have shown that hippocampal GIRK currents are also upregulated by cholesterol. A combined computational-experimental approach pointed to putative cholesterol-binding sites in the transmembrane domain of the GIRK2 channel, the primary subunit in hippocampal GIRK channels. In particular, the principal cholesterol-binding site was located in the center of the transmembrane domain in between the inner and outer α-helices of 2 adjacent subunits. Further studies pointed to a similar cholesterol-binding site in GIRK4, a major subunit in atrial GIRK channels. However, a close look at a sequence alignment of the transmembrane helices of the 2 channels reveals surprising differences among the residues that interact with the cholesterol molecule in these 2 channels. Here, we compare the residues that form putative cholesterol-binding sites in GIRK2 and GIRK4 and discuss the similarities and differences among them.

Monitoring voltage-dependent charge displacement of Shaker B-IR K+ ion channels using radio frequency interrogation.

Directory of Open Access Journals (Sweden)

Sameera Dharia

2011-02-01

Full Text Available Here we introduce a new technique that probes voltage-dependent charge displacements of excitable membrane-bound proteins using extracellularly applied radio frequency (RF, 500 kHz electric fields. Xenopus oocytes were used as a model cell for these experiments, and were injected with cRNA encoding Shaker B-IR (ShB-IR K(+ ion channels to express large densities of this protein in the oocyte membranes. Two-electrode voltage clamp (TEVC was applied to command whole-cell membrane potential and to measure channel-dependent membrane currents. Simultaneously, RF electric fields were applied to perturb the membrane potential about the TEVC level and to measure voltage-dependent RF displacement currents. ShB-IR expressing oocytes showed significantly larger changes in RF displacement currents upon membrane depolarization than control oocytes. Voltage-dependent changes in RF displacement currents further increased in ShB-IR expressing oocytes after ∼120 µM Cu(2+ addition to the external bath. Cu(2+ is known to bind to the ShB-IR ion channel and inhibit Shaker K(+ conductance, indicating that changes in the RF displacement current reported here were associated with RF vibration of the Cu(2+-linked mobile domain of the ShB-IR protein. Results demonstrate the use of extracellular RF electrodes to interrogate voltage-dependent movement of charged mobile protein domains--capabilities that might enable detection of small changes in charge distribution associated with integral membrane protein conformation and/or drug-protein interactions.

Monitoring voltage-dependent charge displacement of Shaker B-IR K+ ion channels using radio frequency interrogation.

Science.gov (United States)

Dharia, Sameera; Rabbitt, Richard D

2011-02-28

Here we introduce a new technique that probes voltage-dependent charge displacements of excitable membrane-bound proteins using extracellularly applied radio frequency (RF, 500 kHz) electric fields. Xenopus oocytes were used as a model cell for these experiments, and were injected with cRNA encoding Shaker B-IR (ShB-IR) K(+) ion channels to express large densities of this protein in the oocyte membranes. Two-electrode voltage clamp (TEVC) was applied to command whole-cell membrane potential and to measure channel-dependent membrane currents. Simultaneously, RF electric fields were applied to perturb the membrane potential about the TEVC level and to measure voltage-dependent RF displacement currents. ShB-IR expressing oocytes showed significantly larger changes in RF displacement currents upon membrane depolarization than control oocytes. Voltage-dependent changes in RF displacement currents further increased in ShB-IR expressing oocytes after ∼120 µM Cu(2+) addition to the external bath. Cu(2+) is known to bind to the ShB-IR ion channel and inhibit Shaker K(+) conductance, indicating that changes in the RF displacement current reported here were associated with RF vibration of the Cu(2+)-linked mobile domain of the ShB-IR protein. Results demonstrate the use of extracellular RF electrodes to interrogate voltage-dependent movement of charged mobile protein domains--capabilities that might enable detection of small changes in charge distribution associated with integral membrane protein conformation and/or drug-protein interactions.

Integral membrane protein structure determination using pseudocontact shifts

Energy Technology Data Exchange (ETDEWEB)

Crick, Duncan J.; Wang, Jue X. [University of Cambridge, Department of Biochemistry (United Kingdom); Graham, Bim; Swarbrick, James D. [Monash University, Monash Institute of Pharmaceutical Sciences (Australia); Mott, Helen R.; Nietlispach, Daniel, E-mail: dn206@cam.ac.uk [University of Cambridge, Department of Biochemistry (United Kingdom)

2015-04-15

Obtaining enough experimental restraints can be a limiting factor in the NMR structure determination of larger proteins. This is particularly the case for large assemblies such as membrane proteins that have been solubilized in a membrane-mimicking environment. Whilst in such cases extensive deuteration strategies are regularly utilised with the aim to improve the spectral quality, these schemes often limit the number of NOEs obtainable, making complementary strategies highly beneficial for successful structure elucidation. Recently, lanthanide-induced pseudocontact shifts (PCSs) have been established as a structural tool for globular proteins. Here, we demonstrate that a PCS-based approach can be successfully applied for the structure determination of integral membrane proteins. Using the 7TM α-helical microbial receptor pSRII, we show that PCS-derived restraints from lanthanide binding tags attached to four different positions of the protein facilitate the backbone structure determination when combined with a limited set of NOEs. In contrast, the same set of NOEs fails to determine the correct 3D fold. The latter situation is frequently encountered in polytopical α-helical membrane proteins and a PCS approach is thus suitable even for this particularly challenging class of membrane proteins. The ease of measuring PCSs makes this an attractive route for structure determination of large membrane proteins in general.

Identification of a novel protein complex containing ASIC1a and GABAA receptors and their interregulation.

Directory of Open Access Journals (Sweden)

Dongbo Zhao

Full Text Available Acid-sensing ion channels (ASICs belong to the family of the epithelial sodium channel/degenerin (ENaC/DEG and are activated by extracellular protons. They are widely distributed within both the central and peripheral nervous systems. ASICs were modified by the activation of γ-aminobutyric acid receptors (GABAA, a ligand-gated chloride channels, in hippocampal neurons. In contrast, the activity of GABAA receptors were also modulated by extracellular pH. However so far, the mechanisms underlying this intermodulation remain obscure. We hypothesized that these two receptors-GABAA receptors and ASICs channels might form a novel protein complex and functionally interact with each other. In the study reported here, we found that ASICs were modified by the activation of GABAA receptors either in HEK293 cells following transient co-transfection of GABAA and ASIC1a or in primary cultured dorsal root ganglia (DRG neurons. Conversely, activation of ASIC1a also modifies the GABAA receptor-channel kinetics. Immunoassays showed that both GABAA and ASIC1a proteins were co-immunoprecipitated mutually either in HEK293 cells co-transfected with GABAA and ASIC1a or in primary cultured DRG neurons. Our results indicate that putative GABAA and ASIC1a channels functionally interact with each other, possibly via an inter-molecular association by forming a novel protein complex.

The effects of information technology based retail channels integration on retail stores performance: mediating role of organizational Ambidexterity (case study: Rasht stores

Directory of Open Access Journals (Sweden)

Mohamadrahim Ramazanian

2015-12-01

Full Text Available Information technology is a critical tool for companies to achieve the competitive advantage and organizational innovation. IT capability provides an appropriate opportunity for retailers to improve their relationships with customers and progress firms’ performance. Comes with advances in technology, retail industry by using Information technology has changed its business process from traditional to online channels. This paper, investigates the effects of IT based retail channel integration on retail stores performance, furthermore the mediating role of organizational ambidexterity as organizational capability in exploitation and exploration of growth opportunities has been examined. Research data has been collected from the retailer sales chains in Rasht city. Data was collected through questionnaires and analyzed by structural equation modeling and partial least squares algorithm. Findings show that retail channel integration based on information technology by mediated organizational ambidexterity influence on performance.

Growing functional modules from a seed protein via integration of protein interaction and gene expression data

Directory of Open Access Journals (Sweden)

Dimitrakopoulou Konstantina

2007-10-01

Full Text Available Abstract Background Nowadays modern biology aims at unravelling the strands of complex biological structures such as the protein-protein interaction (PPI networks. A key concept in the organization of PPI networks is the existence of dense subnetworks (functional modules in them. In recent approaches clustering algorithms were applied at these networks and the resulting subnetworks were evaluated by estimating the coverage of well-established protein complexes they contained. However, most of these algorithms elaborate on an unweighted graph structure which in turn fails to elevate those interactions that would contribute to the construction of biologically more valid and coherent functional modules. Results In the current study, we present a method that corroborates the integration of protein interaction and microarray data via the discovery of biologically valid functional modules. Initially the gene expression information is overlaid as weights onto the PPI network and the enriched PPI graph allows us to exploit its topological aspects, while simultaneously highlights enhanced functional association in specific pairs of proteins. Then we present an algorithm that unveils the functional modules of the weighted graph by expanding a kernel protein set, which originates from a given 'seed' protein used as starting-point. Conclusion The integrated data and the concept of our approach provide reliable functional modules. We give proofs based on yeast data that our method manages to give accurate results in terms both of structural coherency, as well as functional consistency.

Purification and subunit structure of a putative K+-channel protein identified by its binding properties for dendrotoxin I