Localization of trehalose in partially hydrated DOPC bilayers: insights into cryoprotective mechanisms

Trehalose, a natural disaccharide with bioprotective properties, is widely recognized for its ability to preserve biological membranes during freezing and dehydration events. Despite debate over the molecular mechanisms by which this is achieved, and that different mechanisms imply quite different distributions of trehalose molecules with respect to the bilayer, there are no direct experimental data describing the location of trehalose within lipid bilayer membrane systems during dehydration. Here, we use neutron membrane diffraction to conclusively show that the trehalose distribution in a dioleoylphosphatidylcholine (DOPC) system follows a Gaussian profile centred in the water layer between bilayers. The absence of any preference for localizing near the lipid headgroups of the bilayers indicates that the bioprotective effects of trehalose at physiologically relevant concentrations are the result of non-specific mechanisms that do not rely on direct interactions with the lipid headgroups.     

Supramolecular Assembly of Aminoethylene‐Lipopeptide PMO Conjugates into RNA Splice‐Switching Nanomicelles

Phosphorodiamidate morpholino oligomers (PMOs) are oligonucleotide analogs that can be used for therapeutic modulation of pre‐mRNA splicing. Similar to other classes of nucleic acid‐based therapeutics, PMOs require delivery systems for efficient transport to the intracellular target sites. Here, artificial peptides based on the oligo(ethylenamino) acid succinyl‐tetraethylenpentamine (Stp), hydrophobic modifications, and an azide group are presented, which are used for strain‐promoted azide‐alkyne cycloaddition conjugation with splice‐switching PMOs. By systematically varying the lead structure and formulation, it is determined that the type of contained fatty acid and supramolecular assembly have a critical impact on the delivery efficacy. A compound containing linolenic acid with three cis double bonds exhibits the highest splice‐switching activity and significantly increases functional protein expression in pLuc/705 reporter cells in vitro and after local administration in vivo. Structural and mechanistic studies reveal that the lipopeptide PMO conjugates form nanoparticles, which accelerate cellular uptake and that the content of unsaturated fatty acids enhances endosomal escape. In an in vitro Duchenne muscular dystrophy exon skipping model using H2K‐mdx52 dystrophic skeletal myotubes, the highly potent PMO conjugates mediate significant splice‐switching at very low nanomolar concentrations. The presented aminoethylene‐lipopeptides are thus a promising platform for the generation of PMO‐therapeutics with a favorable activity/toxicity profile.     

Photoluminescence of spray pyrolysis deposited ZnO nanorods

Photoluminescence of highly structured ZnO layers comprising well-shaped hexagonal rods is presented. The ZnO rods (length 500-1,000 nm, diameter 100-300 nm) were grown in air onto a preheated soda-lime glass (SGL) or ITO/SGL substrate by low-cost chemical spray pyrolysis method using zinc chloride precursor solutions and growth temperatures in the range of 450-550°C. We report the effect of the variation in deposition parameters (substrate type, growth temperature, spray rate, solvent type) on the photoluminescence properties of the spray-deposited ZnO nanorods. A dominant near band edge (NBE) emission is observed at 300 K and at 10 K. High-resolution photoluminescence measurements at 10 K reveal fine structure of the NBE band with the dominant peaks related to the bound exciton transitions. It is found that all studied technological parameters affect the excitonic photoluminescence in ZnO nanorods.PACS: 78.55.Et, 81.15.Rs, 61.46.Km.     

The effect of damming on biological quality according to macroinvertebrates in some Estonian streams,Central—Baltic Europe: A pilot study

The effect of damming on the structure of the macroinvertebrate community and biological quality was studied in nine (the 3–6th order) lowland streams of Estonia, Central—Baltic ecoregion of Europe. Four habitats—reservoirs with accumulated fine sediments, reservoirs with hard bottom, and two corresponding below‐dam areas (both fast‐flowing)—were compared to study whether and how significantly the bottom substrata in dammed areas affected macroinvertebrates and biological quality downstream of dams. The standard kick‐net samples (1.25 m², complemented with qualitative sample) were collected in autumn 2005—spring 2006. The multimetric biological quality, based on five macroinvertebrate indices (total taxa richness, EPT taxa richness, Average Score Per Taxon, Danish Stream Fauna Index, Shannon diversity) was estimated and compared with reference values. Biological quality in reservoirs with hard bottom and their downstream reaches corresponded to good, or even high quality. Conversely, damming affected biological quality significantly and negatively, above the dam if fine sediments were accumulated. The effect was the strongest within muddy reservoirs themselves (revealing moderate quality only). However, some harmful consequences of mud were observed also downstream of dams. The results also demonstrated that the indices of estimation of organic pollution and/or general quality were able to reflect significant changes in stream flow. Copyright © 2010 John Wiley & Sons, Ltd.     

Identifying Untestable Faults in Sequential Circuits Using Test Path Constraints

The paper proposes a hierarchical untestable stuck-at fault identification method for non-scan synchronous sequential circuits. The method is based on deriving, minimizing and solving test path constraints for modules embedded into Register-Transfer Level (RTL) designs. First, an RTL test pattern generator is applied in order to extract the set of all possible test path constraints for a module under test. Then, the constraints are minimized using an SMT solver Z3 and a logic minimization tool ESPRESSO. Finally, a constraint-driven deterministic test pattern generator is run providing hierarchical test generation and untestability proof in sequential circuits. We show by experiments that the method is capable of quickly proving a large number of untestable faults obtaining higher fault efficiency than achievable by a state-of-the-art commercial ATPG. As a side effect, our study shows that traditional bottom-up test generation based on symbolic test environment generation at RTL is too optimistic due to the fact that propagation constraints are ignored.     

Mixed hierarchical-functional fault models for targeting sequential cores

Current work presents a set of fault models allowing high coverage for sequential cores in systems-on-a-chip. We propose a novel approach combining a hierarchical fault model for functional blocks, a functional fault model for multiplexers and a mixed hierarchical-functional fault model for comparison operators, respectively. The fault models are integrated into a fast high-level decision diagram based test path activation tool. According to the experiments, the proposed method significantly outperforms state-of-the-art test pattern generation tools. The main new contribution of this paper is a formal definition of high-level decision diagram representations and the combination of the three fault models in order to target high gate-level stuck-at fault coverage for sequential cores.     

Underwater confined space mapping by resource‐constrained autonomous vehicle

For marine industrial inspection, archaeology, and geological formation study, the ability to map unknown underwater enclosed and confined spaces is desirable and well suited for robotic vehicles. To date, there are few solutions thoroughly tested in the field designed to perform this specific task, none of which operate autonomously. With a small, low‐cost biomimetic platform known as the U‐CAT, we developed a mapping‐mission software architecture in which the vehicle executes three key sensor‐based reactive stages: entering, exploring, and exiting. Encapsulated in the exploring stage are several state‐defined navigation strategies, called patterns, which were designed and initially tested in simulation. The results of simulation work informed the selection of two patterns that were executed in field trials at a submerged building in Rummu Quarry Lake, Estonia, as part of several full mapping missions. Over the course of these trials, the vehicle was capable of observing the majority (78–97%) of 49.9 explorable square meters within 7 minutes. Based on these results, we demonstrate the capability of a low‐cost and resource‐constrained vehicle to perform confined space mapping under sensor uncertainty. Further, the observations made by the vehicle are shown to be suitable for a target site reconstruction and analysis in postprocessing, which is the intended outcome of this type of mission in practical applications.     

Identification of hydrophobic tags for the degradation of stabilized proteins

New HyTs are a knockout: we previously reported that labeling HaloTag proteins with low molecular weight hydrophobic tags (HyTs) leads to targeted degradation of HaloTag fusion proteins. In this report, we employed a chemical approach to extend this hydrophobic tagging methodology to highly stabilized proteins by synthesizing and evaluating a library of HyTs, which led to the identification of HyT36.