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At least 199 records · Page 11

Life Finds A Way, the Dynamics of E. Coli Evolution in Microgravity

Investigating the evolutionary dynamics of Escherichia coli in microgravity offers a unique opportunity to understand microbial adaptation to extreme environments. Here, we explored the effects of simulated microgravity (SµG) on gene expression and genome evolution of Escherichia coli REL606, a strain continuously evolved and documented terrestrially for 35 years. We used transcriptomic profiling over a 24-hour growth cycle to examine how short-term exposure to SµG under glucose-limiting and glucose-replete conditions may influence the genetic adaptations in microbial populations. Pathway analyses of differentially expressed genes suggest that SµG may alter cell membrane structure and function across all conditions, while changes to protein synthesis machinery were uniquely observed in glucose-replete samples. Furthermore, altered expression of several prophage genes across conditions in SµG samples and upregulation of general stress response factors hints at the potential for stress-induced mutagenesis in response to microgravity. We further investigated the impact of long-term exposure to SµG on genome evolution and observed a more rapid accumulation of base substitutions and deletions in SµG sample genomes across time. Specifically, mutations in the mraZ and elyC genes suggest a mechanism for increased production of peptidoglycan in the cell membrane. These findings offer insights into bacterial adaptations in long-term microgravity environments and pave the way for further detailed investigations.

Brittney Lozzi↗

Some Comments on the Status of Shell Theory at the End of the 20th Century: Complaints and Correctives

This review is divided into complaints and correctives. Complaints are directed at: sloppy refereeing and editing; authors who fail to read or acknowledge what others have done; the mis-naming or mis-crediting of results; the misunderstanding and misuse of the Kirchhoff hypothesis; inflated claims of accuracy based on overly-simplified benchmark problems; the failure to appreciate the inherent errors in various shell models; the failure to appreciate that the physical response and mathematical structure of shell theory are fundamentally different from 3-dimensional elasticity; and the irrelevance of Cosserat-type theories. Correctives include a simple, straight-forward derivation of a general nonlinear dynamic shell theory with the following features: (1) the equations of motion and kinematics (and those of thermodynamics, if desired) are exact consequences of their 3-dimensional counterparts; (2) there are no asymptotic or series expansions through the thickness; (3) all approximations (including the Kirchhoff Hypothesis) occur in the constitutive relations; (4) in static problems, there is a mixed form of the governing equations involving a mixed-energy density and exhibiting remnants of the well-known static-geometric duality of linear theory which is numerically robust because the limiting cases of nonlinear membrane theory and inextensional bending theory fall out naturally. (These latter two special cases are known to produce numerical nightmares unless treated with great care); and (5) all equations may be expressed in coordinate-free form (although, sometimes, a hybrid form is shown to be superior).

Simmonds, James G.↗

Stoichiometry of the large conductance bacterial mechanosensitive channel of E. coli. A biochemical study

MscL, a 15 kDa transmembrane protein, is the only component involved in the formation of a 3 nS channel in the inner membrane of Escherichia coli that opens in response to mechanical or osmotic stress. While previous data had suggested that the functional MscL complex might be a hexamer, a recent crystallographic study of the MscL homologue from M. tuberculosis reveals a pentameric structure. The present work further examines the stoichiometry of the E. coli MscL using a variety of biochemical approaches. Detergent-purified 6His-MscL in solution and MscL in the membrane could be chemically crosslinked with the products displaying ladderlike patterns on SDS gels. Three crosslinking agents (EDC, DMS, and DMA) used at saturating concentrations invariably generated pentamers as the largest product. DSS produced additional bands corresponding to larger complexes although the pentamer band appeared to be the predominant product at high levels of crosslinker. It is not clear whether these extra bands reflect a difference in the crosslinking chemistry of DSS or whether its spacer arm is the longest of those used, or a combination of both facts. For the detergent-solubilized 6His-MscL both sedimentation equilibrium and gel chromatography showed the presence of multiple species. Thus the longer spacer arm could permit both intra- and intercomplex linkages. Nonetheless, the patterns obtained with all agents are consistent with and strongly suggest a pentameric organization for the MscL channel. Expression of MscL as genetically engineered double or triple subunit tandems yields low numbers of functional channels as compared to expressed monomers. The double-tandem assemblies must have an even number of subunits and crosslinking in the membrane confirmed hexamerization. Gel chromatography clearly demonstrated that the channels formed from the double tandems were larger than those formed from WT MscL, consistent with the native channel being pentameric. The observation that both double and triple tandems form channels of normal conductance implies that the pentameric assembly is to some degree independent of the number of subunit repeats in the polypeptide precursor. The channel is thus a pentameric core with the 'extra' subunits left out of the functional complex. From sedimentation equilibrium and size-exclusion chromatography, we also conclude that MscL complexes are not in a dynamic equilibrium with monomers, but are pre-assembled; and thus, their gating properties must result from changes in the conformation of the entire complex induced by the mechanical stress.

NASA Discipline Cell Biology↗

NASA Tech Briefs, March 2009

Topics covered include: Improved Instrument for Detecting Water and Ice in Soil; Real-Time Detection of Dust Devils from Pressure Readings; Determining Surface Roughness in Urban Areas Using Lidar Data; DSN Data Visualization Suite; Hamming and Accumulator Codes Concatenated with MPSK or QAM; Wide-Angle-Scanning Reflectarray Antennas Actuated by MEMS; Biasable Subharmonic Membrane Mixer for 520 to 600 GHz; Hardware Implementation of Serially Concatenated PPM Decoder; Symbolic Processing Combined with Model-Based Reasoning; Presentation Extensions of the SOAP; Spreadsheets for Analyzing and Optimizing Space Missions; Processing Ocean Images to Detect Large Drift Nets; Alternative Packaging for Back-Illuminated Imagers; Diamond Machining of an Off-Axis Biconic Aspherical Mirror; Laser Ablation Increases PEM/Catalyst Interfacial Area; Damage Detection and Self-Repair in Inflatable/Deployable Structures; Polyimide/Glass Composite High-Temperature Insulation; Nanocomposite Strain Gauges Having Small TCRs; Quick-Connect Windowed Non-Stick Penetrator Tips for Rapid Sampling; Modeling Unsteady Cavitation and Dynamic Loads in Turbopumps; Continuous-Flow System Produces Medical-Grade Water; Discrimination of Spore-Forming Bacilli Using spoIVA; nBn Infrared Detector Containing Graded Absorption Layer; Atomic References for Measuring Small Accelerations; Ultra-Broad-Band Optical Parametric Amplifier or Oscillator; Particle-Image Velocimeter Having Large Depth of Field; Enhancing SERS by Means of Supramolecular Charge Transfer; Improving 3D Wavelet-Based Compression of Hyperspectral Images; Improved Signal Chains for Readout of CMOS Imagers; SOI CMOS Imager with Suppression of Cross-Talk; Error-Rate Bounds for Coded PPM on a Poisson Channel; Biomorphic Multi-Agent Architecture for Persistent Computing; and Using Covariance Analysis to Assess Pointing Performance.

Source record↗

Small, Fast TES Microcalorimeters with Unprecedented X-ray Spectral Performance

Driven initially by the desire for X-ray microcalorimeter arrays suitable for imaging the dynamic solar corona, we have developed a transition-edge-sensor (TES) microcalorimeter optimization that exhibits a unique combination of high spectral resolving power and a wide X-ray bandpass. These devices have achieved spectral performance of dE approximately 1.3 eV FWHM at 1.5 keV, 1.6 eV at 6 keV, and 2.0 eV at 8 keV, using small TESs (e.g., approximately 35 micron x 35 micron) that operate in a regime in which the superconducting transition is highly current dependent. In order to accommodate high X-ray count rates, the devices sit directly on a solid substrate instead of on membranes, and we use an embedded heatsinking layer to reduce pixel-to-pixel crosstalk. We will present results from devices with a range of TES and absorber sizes, and from device wafers with varied embedded heatsink materials. This contribution will focus on count-rate capabilities, including a discussion of the trade-off between count rate and energy resolution, and the heatsinking design. We will also present preliminary tests of array readout using a code-division multiplexed SQUID readout scheme, which may be necessary to enable large arrays of these fast devices.

Eckart, M. E.↗

Observations of dynamical phenomena in sunspots

A preliminary report of the results of one observing run based on data from one spectral line, the photospheric magnetic line Fe 6303, is presented as part of a series of observations of dynamical phenomena in sunspots using photographic spectra with the SPO vacuum tower telescope and echelle spectrograph. The ejection of a magnetic feature from the outer edge of the penumbra was observed. The initial total field strength of the feature was about 1000 gauss, which appeared to decrease as the feature moved away from the sunspot. The proper motion was about 2 km/s, and the velocity field measured in the V profile showed a downflow of 400 m/s on the spotward side of the moving magnetic feature. Umbral oscillations at the photospheric level with a herringbone structure characteristic of horizontally propagating waves, suggesting some overtone mode of membrane oscillation in the umbra, were seen. The peak amplitude of the oscillation was about 200 m/s, and the mean power spectrum had several clear peaks.

Nye, A. H.↗

Amyloplast sedimentation dynamics in maize columella cells support a new model for the gravity-sensing apparatus of roots

Quantitative analysis of statolith sedimentation behavior was accomplished using videomicroscopy of living columella cells of corn (Zea mays) roots, which displayed no systematic cytoplasmic streaming. Following 90 degrees rotation of the root, the statoliths moved downward along the distal wall and then spread out along the bottom with an average velocity of 1.7 microm min(-1). When statolith trajectories traversed the complete width or length of the cell, they initially moved horizontally toward channel-initiation sites and then moved vertically through the channels to the lower side of the reoriented cell where they again dispersed. These statoliths exhibited a significantly lower average velocity than those sedimenting on distal-to-side trajectories. In addition, although statoliths undergoing distal-to-side sedimentation began at their highest velocity and slowed monotonically as they approached the lower cell membrane, statoliths crossing the cell's central region remained slow initially and accelerated to maximum speed once they reached a channel. The statoliths accelerated sooner, and the channeling effect was less pronounced in roots treated with cytochalasin D. Parallel ultrastructural studies of high-pressure frozen-freeze-substituted columella cells suggest that the low-resistance statolith pathway in the cell periphery corresponds to the sharp interface between the endoplasmic reticulum (ER)-rich cortical and the ER-devoid central region of these cells. The central region is also shown to contain an actin-based cytoskeletal network in which the individual, straight, actin-like filaments are randomly distributed. To explain these findings as well as the results of physical simulation experiments, we have formulated a new, tensegrity-based model of gravity sensing in columella cells. This model envisages the cytoplasm as pervaded by an actin-based cytoskeletal network that is denser in the ER-devoid central region than in the ER-rich cell cortex and is linked to stretch receptors in the plasma membrane. Sedimenting statoliths are postulated to produce a directional signal by locally disrupting the network and thereby altering the balance of forces acting on the receptors in different plasma membrane regions.

NASA Program Fundamental Space Biology↗

New Modeling Approaches to Investigate Cell Signaling in Radiation Response

Ionizing radiation damages individual cells and tissues leading to harmful biological effects. Among many radiation-induced lesions, DNA double-strand breaks (DSB) are considered the key precursors of most early and late effects [1] leading to direct mutation or aberrant signal transduction processes. In response to damage, a flow of information is communicated to cells not directly hit by the radiation through signal transduction pathways [2]. Non-targeted effects (NTE), which includes bystander effects and genomic instability in the progeny of irradiated cells and tissues, may be particularly important for space radiation risk assessment [1], because astronauts are exposed to a low fluence of heavy ions and only a small fraction of cells are traversed by an ion. NTE may also have important consequences clinical radiotherapy [3]. In the recent years, new simulation tools and modeling approaches have become available to study the tissue response to radiation. The simulation of signal transduction pathways require many elements such as detailed track structure calculations, a tissue or cell culture model, knowledge of biochemical pathways and Brownian Dynamics (BD) propagators of the signaling molecules in their micro-environment. Recently, the Monte-Carlo simulation code of radiation track structure RITRACKS was used for micro and nano-dosimetry calculations [4]. RITRACKS will be used to calculate the fraction of cells traversed by an ion and delta-rays and the energy deposited in cells in a tissue model. RITRACKS also simulates the formation of chemical species by the radiolysis of water [5], notably the .OH radical. This molecule is implicated in DNA damage and in the activation of the transforming growth factor beta (TGF), a signaling molecule involved in NTE. BD algorithms for a particle near a membrane comprising receptors were also developed and will be used to simulate trajectories of signaling molecules in the micro-environment and characterize autocrine and paracrine cell communication and signal transduction.

Plante, Ianik↗

Mechanical behavior in living cells consistent with the tensegrity model

Alternative models of cell mechanics depict the living cell as a simple mechanical continuum, porous filament gel, tensed cortical membrane, or tensegrity network that maintains a stabilizing prestress through incorporation of discrete structural elements that bear compression. Real-time microscopic analysis of cells containing GFP-labeled microtubules and associated mitochondria revealed that living cells behave like discrete structures composed of an interconnected network of actin microfilaments and microtubules when mechanical stresses are applied to cell surface integrin receptors. Quantitation of cell tractional forces and cellular prestress by using traction force microscopy confirmed that microtubules bear compression and are responsible for a significant portion of the cytoskeletal prestress that determines cell shape stability under conditions in which myosin light chain phosphorylation and intracellular calcium remained unchanged. Quantitative measurements of both static and dynamic mechanical behaviors in cells also were consistent with specific a priori predictions of the tensegrity model. These findings suggest that tensegrity represents a unified model of cell mechanics that may help to explain how mechanical behaviors emerge through collective interactions among different cytoskeletal filaments and extracellular adhesions in living cells.

NASA Discipline Cell Biology↗

NASA Tech Briefs, January 2012

Contents of this issue are: (1) Energy-Based Tetrahedron Sensor for High-Temperature, High-Pressure Environments (2) Handheld Universal Diagnostic Sensor (3) Large-Area Vacuum Ultraviolet Sensors (4) Fiber Bragg Grating Sensor System for Monitoring Smart Composite Aerospace Structures (5) Health-Enabled Smart Sensor Fusion Technology (6) Extended-Range Passive RFID and Sensor Tags (7) Hybrid Collaborative Learning for Classification and Clustering in Sensor Networks (8) Self-Healing, Inflatable, Rigidizable Shelter (9) Improvements in Cold-Plate Fabrication (10) Technique for Radiometer and Antenna Array Calibration - TRAAC (11) Real-Time Cognitive Computing Architecture for Data Fusion in a Dynamic Environment (12) Programmable Digital Controller (13) Use of CCSDS Packets Over SpaceWire to Control Hardware (14) Key Decision Record Creation and Approval Module (15) Enhanced Graphics for Extended Scale Range (16) Debris Examination Using Ballistic and Radar Integrated Software (17) Data Distribution System (DDS) and Solar Dynamic Observatory Ground Station (SDOGS) (18) Integration Manager (19) Eclipse-Free-Time Assessment Tool for IRIS (20) Automated and Manual Rocket Crater Measurement Software (21) MATLAB Stability and Control Toolbox Trim and Static Stability Module (22) Patched Conic Trajectory Code (23) Ring Image Analyzer (24) SureTrak Probability of Impact Display (25) Implementation of a Non-Metallic Barrier in an Electric Motor (26) Multi-Mission Radioisotope Thermoelectric Generator Heat Exchangers for the Mars Science Laboratory Rover (27) Uniform Dust Distributor for Testing Radiative Emittance of Dust-Coated Surfaces (28) MicroProbe Small Unmanned Aerial System (29) Highly Stable and Active Catalyst for Sabatier Reactions (30) Better Proton-Conducting Polymers for Fuel-Cell Membranes (31) CCD Camera Lens Interface for Real-Time Theodolite Alignment (32) Peregrine 100-km Sounding Rocket Project (33) SOFIA Closed- and Open-Door Aerodynamic Analyses (34) Sonic Thermometer for High-Altitude Balloons (35) Near-Infrared Photon-Counting Camera for High-Sensitivity Observations (36) Integrated Optics Achromatic Nuller for Stellar Interferometry (37) High-Speed Digital Interferometry (38) Ultra-Miniature Lidar Scanner for Launch Range Data Collection (39) Shape and Color Features for Object Recognition Search (40) Explanation Capabilities for Behavior-Based Robot Control (41) A DNA-Inspired Encryption Methodology for Secure, Mobile Ad Hoc Networks (42) Quality Control Method for a Micro-Nano-Channel Microfabricated Device (43) Corner-Cube Retroreflector Instrument for Advanced Lunar Laser Ranging (44) Electrospray Collection of Lunar Dust (45) Fabrication of a Kilopixel Array of Superconducting Microcalorimeters with Microstripline Wiring Spacecraft Attitude Tracking and Maneuver Using Combined Magnetic Actuators (46) Coherent Detector for Near-Angle Scattering and Polarization Characterization of Telescope Mirror Coatings

Source record↗

A study of structural concepts for ultralightweight spacecraft

Structural concepts for ultralightweight spacecraft were studied. Concepts for ultralightweight space structures were identified and the validity of heir potential application in advanced spacecraft was assessed. The following topics were investigated: (1) membrane wrinkling under pretensioning; (2) load-carrying capability of pressurized tubes; (3) equilibrium of a precompressed rim; (4) design of an inflated reflector spacecraft; (5) general instability of a rim; and (6) structural analysis of a pressurized isotensoid column. The design approaches for a paraboloidal reflector spacecraft included a spin-stiffened design, both inflated and truss central columns, and to include both deep truss and rim-stiffened geodesic designs. The spinning spacecraft analysis is included, and the two truss designs are covered. The performances of four different approaches to the structural design of a paraboloidal reflector spacecraft are compared. The spinning and inflated configurations result in very low total masses and some concerns about their performance due to unresolved questions about dynamic stability and lifetimes, respectively.

Miller, R. K.↗

Linking Archaeal Molecular Diversity and Lipid Biomarker Composition in a Hypersaline Microbial Mat Community

Lipid biomarkers for discrete microbial groups are a valuable tool for establishing links to ancient microbial ecosystems. Lipid biomarkers can establish organism source and function in contemporary microbial ecosystems (membrane lipids) and by analogy, potential relevance to the fossilized carbon skeletons (geolipids) extracted from ancient sedimentary rock. The Mars Exploration Rovers have provided clear evidence for an early wet Mars and the presence of hypersaline evaporitic basins. Ongoing work on an early Earth analog, the hypersaline benthic mats in Guerrero Negro, Baja California Sur, may provide clues to what may have evolved and flourished on an early wet Mars, if only for a short period. Cyanobacterial mats are a pertinent early Earth analog for consideration of evolutionary and microbial processes within the aerobic photosynthetic and adjacent anoxic layers. Fluctuations in physio-chemical parameters associated with spatial and temporal scales are expressed through vast microbial metabolic diversity. Our recent work hopes to establish the dynamic of archaeal diversity, particularly as it relates to methane production in this high sulfate environment, through the use of lipid biomarker and phylogenetic analyses. Archaeal 16s rRNA and mcrA gene assemblages, demonstrated distinct spatial separation over the 130 mm core of at least three distinct genera within the order Methanosarcinales, as well as an abundance of uncultured members of the Thermoplasmales and Crenarchaeota. Ether-bound lipid analysis identified abundant 0-alkyl and 0-isopranyl chains throughout the core, and the presence of sn-2 hydroxyarchaeol, a biomarker for methylotrophic methanogens. A unique ether isoprenoid chain, a C30:1 , possibly related to the geolipid squalane, a paleobiomarker associated with hypersaline environments, was most abundant within the oxic-anoxic transition zone.

Jahnke, Linda↗

Epidermal Homeostasis and Radiation Responses in a Multiscale Tissue Modeling Framework

The surface of skin is lined with several thin layers of epithelial cells that are maintained throughout life time by a small population of stem cells. High dose radiation exposures could injure and deplete the underlying proliferative cells and induce cutaneous radiation syndrome. In this work we propose a multiscale computational model for skin epidermal dynamics that links phenomena occurring at the subcellular, cellular, and tissue levels of organization, to simulate the experimental data of the radiation response of swine epidermis, which is closely similar to human epidermis. Incorporating experimentally measured histological and cell kinetic parameters, we obtain results of population kinetics and proliferation indexes comparable to observations in unirradiated and acutely irradiated swine experiments. At the sub-cellular level, several recently published Wnt signaling controlled cell-cycle models are applied and the roles of key components and parameters are analyzed. Based on our simulation results, we demonstrate that a moderate increase of proliferation rate for the survival proliferative cells is sufficient to fully repopulate the area denuded by high dose radiation, as long as the integrity of underlying basement membrane is maintained. Our work highlights the importance of considering proliferation kinetics as well as the spatial organization of tissues when conducting in vivo investigations of radiation responses. This integrated model allow us to test the validity of several basic biological rules at the cellular level and sub-cellular mechanisms by qualitatively comparing simulation results with published research, and enhance our understanding of the pathophysiological effects of ionizing radiation on skin.

Hu, Shaowen↗

Evaluation of Microbolometer-Based Thermography for Gossamer Space Structures

In August 2003, NASA's In-Space Propulsion Program contracted with our team to develop a prototype on-board Optical Diagnostics System (ODS) for solar sail flight tests. The ODS is intended to monitor sail deployment as well as structural and thermal behavior, and to validate computational models for use in designing future solar sail missions. This paper focuses on the thermography aspects of the ODS. A thermal model was developed to predict local sail temperature variations as a function of sail tilt to the sun, billow depth, and spectral optical properties of front and back sail surfaces. Temperature variations as small as 0.5 C can induce significant thermal strains that compare in magnitude to mechanical strains. These thermally induced strains may result in changes in shape and dynamics. The model also gave insight into the range and sensitivity required for in-flight thermal measurements and supported the development of an ABAQUS-coupled thermo-structural model. The paper also discusses three kinds of tests conducted to 1) determine the optical properties of candidate materials; 2) evaluate uncooled microbolometer-type infrared imagers; and 3) operate a prototype imager with the ODS baseline configuration. (Uncooled bolometers are less sensitive than cooled ones, but may be necessary because of restrictive ODS mass and power limits.) The team measured the spectral properties of several coated polymer samples at various angles of incidence. Two commercially available uncooled microbolometer imagers were compared, and it was found that reliable temperature measurements are feasible for both coated and uncoated sides of typical sail membrane materials.

Miles, Jonathan J.↗

Mathematical model of diffusion-limited gas bubble dynamics in unstirred tissue with finite volume

Models of gas bubble dynamics for studying decompression sickness have been developed by considering the bubble to be immersed in an extravascular tissue with diffusion-limited gas exchange between the bubble and the surrounding unstirred tissue. In previous versions of this two-region model, the tissue volume must be theoretically infinite, which renders the model inapplicable to analysis of bubble growth in a finite-sized tissue. We herein present a new two-region model that is applicable to problems involving finite tissue volumes. By introducing radial deviations to gas tension in the diffusion region surrounding the bubble, the concentration gradient can be zero at a finite distance from the bubble, thus limiting the tissue volume that participates in bubble–tissue gas exchange. It is shown that these deviations account for the effects of heterogeneous perfusion on gas bubble dynamics, and are required for the tissue volume to be finite. The bubble growth results from a difference between the bubble gas pressure and an average gas tension in the surrounding diffusion region that explicitly depends on gas uptake and release by the bubble. For any given decompression, the diffusion region volume must stay above a certain minimum in order to sustain bubble growth.

NASA Discipline Environmental Health↗

Dynamic and thermal response finite element models of multi-body space structural configurations

Presented is structural dynamics modeling of two multibody space structural configurations. The first configuration is a generic space station model of a cylindrical habitation module, two solar array panels, radiator panel, and central connecting tube. The second is a 15-m hoop-column antenna. Discussed is the special joint elimination sequence used for these large finite element models, so that eigenvalues could be extracted. The generic space station model aided test configuration design and analysis/test data correlation. The model consisted of six finite element models, one of each substructure and one of all substructures as a system. Static analysis and tests at the substructure level fine-tuned the finite element models. The 15-m hoop-column antenna is a truss column and structural ring interconnected with tension stabilizing cables. To the cables, pretensioned mesh membrane elements were attached to form four parabolic shaped antennae, one per quadrant. Imposing thermal preloads in the cables and mesh elements produced pretension in the finite element model. Thermal preload variation in the 96 control cables was adjusted to maintain antenna shape within the required tolerance and to give pointing accuracy.

Edighoffer, Harold H.↗

Technical advance: identification of plant actin-binding proteins by F-actin affinity chromatography

Proteins that interact with the actin cytoskeleton often modulate the dynamics or organization of the cytoskeleton or use the cytoskeleton to control their localization. In plants, very few actin-binding proteins have been identified and most are thought to modulate cytoskeleton function. To identify actin-binding proteins that are unique to plants, the development of new biochemical procedures will be critical. Affinity columns using actin monomers (globular actin, G-actin) or actin filaments (filamentous actin, F-actin) have been used to identify actin-binding proteins from a wide variety of organisms. Monomeric actin from zucchini (Cucurbita pepo L.) hypocotyl tissue was purified to electrophoretic homogeneity and shown to be native and competent for polymerization to actin filaments. G-actin, F-actin and bovine serum albumin affinity columns were prepared and used to separate samples enriched in either soluble or membrane-associated actin-binding proteins. Extracts of soluble actin-binding proteins yield distinct patterns when eluted from the G-actin and F-actin columns, respectively, leading to the identification of a putative F-actin-binding protein of approximately 40 kDa. When plasma membrane-associated proteins were applied to these columns, two abundant polypeptides eluted selectively from the F-actin column and cross-reacted with antiserum against pea annexins. Additionally, a protein that binds auxin transport inhibitors, the naphthylphthalamic acid binding protein, which has been previously suggested to associate with the actin cytoskeleton, was eluted in a single peak from the F-actin column. These experiments provide a new approach that may help to identify novel actin-binding proteins from plants.

Non-NASA Center↗

Small Ion Channel Linking Molecular Simulations and Electrophysiology

Ion channels are pore-forming protein assemblies that mediate the transport of small ions across cell membranes. Otherwise, membrane bilayers would be almost impermeable to ions incapable to traverse the low dielectric constant, hydrophobic membrane core. Ion channels are ubiquitous to all life forms. In humans and other higher organisms they play the central role in conducting nerve impulses, cardiac functions, muscle contraction and apoptosis. On the other extreme of biological complexity, viral ion channels (viroporins) influence many stages of the virus infection cycle either through regulating virus replication, such as entry, assembly and release or modulating the electrochemical balance in the subcellular compartments of host cells. Ion channels were crucial components of protocells. Their emergence facilitated adaptation of nascent life to different environmental conditions. The earliest ion channels must have been much simpler than most of their modern ancestors. Viral channels are among only a few naturally occurring models to study the structure, function and evolution of primordial channels. Experimental studies of these properties are difficult and often unreliable. In principle, computational methods, and molecular dynamics (MD) simulations in particular, can aid in providing information about both the structure and the function of ion channels. However, MD suffers from its own problems, such as inability to access sufficiently long time scales or limited accuracy of force fields. It is, therefore, essential to determine the reliability of MD simulations. We propose to do so on the basis of two criteria. One is channel stability on time scales that extend for several microseconds or longer. The other is the ability to reproduce the measured ionic conductance as a function of applied voltage. If both the stability and the calculated ionic conductance are satisfactory it will greatly increase our confidence that the structure and the function of a channel are described sufficiently accurately. To our knowledge, long time scale stability (approx.10 micro-sec) and the correct electrophysiology have been shown so far for only one channel - the synthetic LS3 hexamer). In this presentation, this approach will be discussed in application to two viral channels - Vpu, encoded by the HIV-1 genome and p7 of hepatitis C.

Pohorille, Andrzej↗