The N-Terminus of Apolipoprotein A-V Adopts a Helix-Bundle Molecular Architecture

Previous studies of recombinant full-length human apolipoprotein A-V (apoA-V) provided evidence of the presence of two independently folded structural domains. Computer-assisted sequence analysis and limited proteolysis studies identified an N-terminal fragment as a candidate for one of the domains. C-Terminal truncation variants in this size range, apoA-V(1-146) and apoA-V(1-169), were expressed in Escherichia coli and isolated. Unlike full-length apoA-V or apoA-V(1-169), apoA-V(1-146) was soluble in neutral-pH buffer in the absence of lipid. Sedimentation equilibrium analysis yielded a weight-average molecular weight of 18811, indicating apoA-V(1-146) exists as a monomer in solution. Guanidine HCl denaturation experiments at pH 3.0 yielded a one-step native to unfolded transition that corresponds directly with the more stable component of the two-stage denaturation profile exhibited by full-length apoA-V. On the other hand, denaturation experiments conducted at pH 7.0 revealed a less stable structure. In a manner similar to that of known helix bundle apolipoproteins, apoA-V(1-146) induced a relatively small enhancement in 8-anilino-1-naphthalenesulfonic acid fluorescence intensity. Quenching studies with single-Trp apoA-V(1-146) variants revealed that a unique site predicted to reside on the nonpolar face of an amphipathic R-helix was protected from quenching by KI. Taken together, the data suggest the 146 N-terminal residues of human apoA-V adopt a helix bundle molecular architecture in the absence of lipid and, thus, likely exist as an independently folded structural domain within the context of the intact protein

Forced triboelectrification of fine powders in particle wall collisions

Triboelectric separation as an inexpensive and environmentally friendly technique could contribute to material-specific sorting. However, the application as a widespread method is limited due to the complexity of the process. In particle wall collisions, various parameters like collision energy and angle, work function of the contact partners, humidity, surface roughness, etc. influence the particle charging in a hardly predictable way. This study investigates the possibilities of forced triboelectric particle charging by applying an electrical potential to the metal contact partner (copper/steel pipe). The variations included different pipe lengths (0.5 m–3 m), particle materials, and particle sizes for limestone. A distinction is made between the net charge of the particles and the positive, negative, and neutral mass fractions. The work functions of the investigated materials vary from about 3.2 eV to >8.5 eV for glass, limestone, artificial slag, and lithium aluminate particles. With the applied high-voltage potential, the particle net charge can be shifted linearly. For limestone, it is shown that the neutral fraction is highest at the Point of Zero Net Charge (PZNC). This observation may identify an approach for the material selective separation of one target component from a multi-material mixture

Development of a model for the separation characteristics of a deflector wheel classifier including particle collision and rebound behavior

Deflector wheel classifiers are widespread in industry for the separation of powders into fine and coarse powders. Even though this separation process has been known for quite some time, it is not yet fully understood, and existing models fail to precisely predict the separation characteristics. Due to the high throughput of deflector wheel classifiers, it is greatly beneficial to estimate the separation characteristics before the experiment. Here, the developed model critically examines the usual assumptions, such as ideal airflow, neglection of particle–wall and particle–particle interactions, or spherically-shaped particles. First, the investigation of the air flow using a Particle Image Velocimetry (PIV) system showed significant differences to the assumed ideal flow field, then particle sphericity and its influence on the interaction between the particles and the paddles of the deflector wheel was investigated and compared with particle rebound behavior on a static wall. Surprisingly, comminuted glass behaves similarly to comminuted limestone in multiple aspects and not like glass beads. To determine the number of particle–particle collisions, Discrete Element Method (DEM) simulations were performed. The aforementioned aspects found application in the model and the separation behavior was well-estimated

Amphotericin B induced interdigitation of apolipoprotein stabilized nanodisk bilayers

Amphotericin B nanodisks (AMB-ND) are ternary complexes of AMB, phospholipid (PL) and apolipoprotein organized as discrete nanometer scale disk-shaped bilayers. In gel filtration chromatography experiments, empty ND lacking AMB elute as a single population of particles with a molecular weight in the range of 200 kDa. AMB-ND formulated at a 4:1 PL:AMB weight ratio, separated into two peaks. Peak 1 eluted at the position of control ND lacking AMB while the second peak, containing all of the AMB present in the original sample, eluted in the void volume. When ND prepared with increased AMB (1:1 phospholipid:AMB molar ratio) were subjected to gel filtration chromatography, an increased proportion of phospholipid and apolipoprotein were recovered in the void volume with the AMB. Prior to gel filtration the AMB-ND sample could be passed through a 0.22 {micro}m filter without loss of AMB while the voided material was lost. Native gel electrophoresis studies corroborated the gel permeation chromatography data. Far UV circular dichroism analyses revealed that apoA-I associated with AMB-ND denatures at a lower guanidine HCl concentration than apoA-I associated with ND lacking AMB. Atomic force microscopy revealed that AMB induces compression of the ND bilayer thickness consistent with bilayer interdigitation, a phenomenon that is likely related to the ability of AMB to induce pore formation in susceptible membranes

Apolipoprotein A–I binding to anionic vesicles and lipopolysaccharides: Role for lysine residues in antimicrobial properties

AbstractHuman apolipoprotein A–I (apoA–I) is a 28kDa protein and a major component of high-density lipoproteins, mediating several essential metabolic functions related to heart disease. In the present study the potential protective role against bacterial pathogens was explored. ApoA–I suppressed bacterial growth of Escherichia coli and Klebsiella pneumoniae. The protein was able to bind lipopolysaccharides and showed a strong preference for bilayer vesicles made of phosphatidylglycerol over phosphatidylcholine. Lysine side chains of apoA–I were acetylated to evaluate the importance of electrostatic forces in the binding interaction with both membrane components. Electrophoresis properties, dot blot analysis, circular dichroism, and fluorescence spectroscopy to probe for changes in protein structure indicated that the acetylated protein displayed a strongly reduced lipopolysaccharide and phosphatidylglycerol binding. A mutant containing only the N-terminal domain of apoA–I also showed a reduced ability to interact with the membrane components, although to a lesser extent. These results indicate the potential for apoA–I to function as an antimicrobial protein and exerts this function through lysine residues

Measurements of NO, NOy, N2O, and O3 during SPURT: implications for transport and chemistry in the lowermost stratosphere

We present measurements of NO, NOy, O3, and N2O within the lowermost stratosphere (LMS) over Europe obtained during the SPURT project. The measurements cover all seasons between November 2001 and July 2003. They span a broad band of latitudes from 30° N to 75° N and a potential temperature range from 290 to 380 K. The measurements represent a comprehensive data set of these tracers and reveal atmospheric transport processes that influence tracer distributions in the LMS. Median mixing ratios of stratospheric tracers in equivalent latitude-potential temperature coordinates show a clear seasonal cycle related to the Brewer-Dobson circulation, with highest values in spring and lowest values in autumn. Vertical tracer profiles show strong gradients at the extratropical tropopause, suggesting that vertical (cross-isentropic) mixing is reduced above the tropopause. Pronounced meridional gradients in the tracer mixing ratios are found on potential temperature surfaces in the LMS. This suggests strongly reduced mixing along isentropes. Concurrent large gradients in static stability in the vertical direction, and of PV in the meridional direction, suggest the presence of a mixing barrier. Seasonal cycles were found in the correlation slopes ΔO3/ΔN2O and ΔNOy/ΔN2O well above the tropopause. Absolute slope values are smallest in spring indicating chemically aged stratospheric air originating from high altitudes and latitudes. Larger values were measured in summer and autumn suggesting that a substantial fraction of air takes a "short-cut" from the tropical tropopause region into the extratropical LMS. The seasonal change in the composition of the LMS has direct implications for the ozone chemistry in this region. Comparisons of measured NO with the critical NO value at which net ozone production changes from negative to positive, imply ozone production up to 20 K above the local tropopause in spring, up to 30 K in summer, and up to 40 K in autumn. Above these heights, and in winter, net ozone production is negative

Tracing troposphere-to-stratosphere transport above a mid-latitude deep convective system

Within the project SPURT (trace gas measurements in the tropopause region) a variety of trace gases have been measured in situ in order to investigate the role of dynamical and chemical processes in the extra-tropical tropopause region. In this paper we report on a flight on 10 November 2001 leading from Hohn, Germany (52�N) to Faro, Portugal (37�N) through a strongly developed deep stratospheric intrusion. This streamer was associated with a large convective system over the western Mediterranean with potentially significant troposphere-to-stratosphere transport. Along major parts of the flight we measured unexpectedly high NOy mixing ratios. Also H2O mixing ratios were significantly higher than stratospheric background levels confirming the extraordinary chemical signature of the probed air masses in the interior of the streamer. Backward trajectories encompassing the streamer enable to analyze the origin and physical characteristics of the air masses and to trace troposphere-to-stratosphere transport. Near the western flank of the streamer features caused by long range transport, such as tropospheric filaments characterized by sudden drops in the O3 and NOy mixing ratios and enhanced CO and H2O can be reconstructed in great detail using the reverse domain filling technique. These filaments indicate a high potential for subsequent mixing with the stratospheric air. At the south-western edge of the streamer a strong gradient in the NOy and the O3 mixing ratios coincides very well with a sharp gradient in potential vorticity in the ECMWF fields. In contrast, in the interior of the streamer the observed highly elevated NOy and H2O mixing ratios up to a potential temperature level of 365K and potential vorticity values of maximum 10 PVU cannot be explained in terms of resolved troposphere-to-stratosphere transport along the backward trajectories. Also mesoscale simulations with a High Resolution Model reveal no direct evidence for convective H2O injection up to this level. Elevated H2O mixing ratios in the ECMWF and HRM are seen only up to about tropopause height at 340 hPa and 270 hPa, respectively, well below flight altitude of about 200 hPa. However, forward tracing of the convective influence as identified by satellite brightness temperature measurements and counts of lightning strokes shows that during this part of the flight the aircraft was closely following the border of an air mass which was heavily impacted by convective activity over Spain and Algeria. This is evidence that deep convection at mid-latitudes may have a large impact on the tracer distribution of the lowermost stratosphere reaching well above the thunderstorms anvils as claimed by recent studies using cloud-resolving models

Development of a Model for the Separation Characteristics of a Deflector Wheel Classifier Including Particle Collision and Rebound Behavior

Deflector wheel classifiers are widespread in industry for the separation of powders into fine and coarse powders. Even though this separation process has been known for quite some time, it is not yet fully understood, and existing models fail to precisely predict the separation characteristics. Due to the high throughput of deflector wheel classifiers, it is greatly beneficial to estimate the separation characteristics before the experiment. Here, the developed model critically examines the usual assumptions, such as ideal airflow, neglection of particle–wall and particle–particle interactions, or spherically-shaped particles. First, the investigation of the air flow using a Particle Image Velocimetry (PIV) system showed significant differences to the assumed ideal flow field, then particle sphericity and its influence on the interaction between the particles and the paddles of the deflector wheel was investigated and compared with particle rebound behavior on a static wall. Surprisingly, comminuted glass behaves similarly to comminuted limestone in multiple aspects and not like glass beads. To determine the number of particle–particle collisions, Discrete Element Method (DEM) simulations were performed. The aforementioned aspects found application in the model and the separation behavior was well-estimated.DFG, 313858373, SPP 2045: Hochspezifische mehrdimensionale Fraktionierung von technischen Feinstpartikelsysteme

The x-ray microcalorimeter spectrometer onboard Athena

Trabajo presentado a la conferencia: "Space Telescopes and Instrumentation: Ultraviolet to Gamma Ray" celebrada en Amsterdam (Holanda) el 1 de julio de 2012.-- et al.One of the instruments on the Advanced Telescope for High-Energy Astrophysics (Athena) which was one of the three missions under study as one of the L-class missions of ESA, is the X-ray Microcalorimeter Spectrometer (XMS). This instrument, which will provide high-spectral resolution images, is based on X-ray micro-calorimeters with Transition Edge Sensor (TES) and absorbers that consist of metal and semi-metal layers and a multiplexed SQUID readout. The array (32 x 32 pixels) provides an energy resolution of < 3 eV. Due to the large collection area of the Athena optics, the XMS instrument must be capable of processing high counting rates, while maintaining the spectral resolution and a low deadtime. In addition, an anti-coincidence detector is required to suppress the particle-induced background. Compared to the requirements for the same instrument on IXO, the performance requirements have been relaxed to fit into the much more restricted boundary conditions of Athena. In this paper we illustrate some of the science achievable with the instrument. We describe the results of design studies for the focal plane assembly and the cooling systems. Also, the system and its required spacecraft resources will be given. © (2012) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE). Downloading of the abstract is permitted for personal use only.Peer Reviewe