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The social amoeba Dictyostelium discoideum is a widely used model organism for studying basic functions of protozoan and metazoan cells, such as osmoregulation and cell motility. There is evidence from other species that cellular water channels, aquaporins (AQP), are central to both processes. Yet, data on D. discoideum AQPs is almost absent. Despite cloning of two putative D. discoideum AQPs, WacA, and AqpA, water permeability has not been shown. Further, WacA and AqpA are expressed at the late multicellular stage and in spores but not in amoebae. We cloned a novel AQP, AqpB, from amoeboidal D. discoideum cells. Wild-type AqpB was impermeable to water, glycerol, and urea when expressed in Xenopus laevis oocytes. Neither stepwise truncation of the N terminus nor selected point mutations activated the water channel. However, mutational truncation by 12 amino acids of an extraordinary long intracellular loop induced water permeability of AqpB, hinting at a novel gating mechanism. This AqpB mutant was inhibited by mercuric chloride, confirming the presence of a cysteine residue in the selectivity filter as predicted by our structure model. We detected AqpB by Western blot analysis in a glycosylated and a non-glycosylated form throughout all developmental stages. When expressed in D. discoideum amoebae, AqpB-GFP fusion constructs localized to vacuolar structures, to the plasma membrane, and to lamellipodia-like membrane protrusions. We conclude that the localization pattern in conjunction with channel gating may be indicative of AqpB functions in osmoregulation as well as cell motility of D. discoideum.
The 2[4Fe-4S] ferredoxin from Chromatium vinosum arises as one prominent member of a recently defined family of proteins found in very diverse bacteria. The potentiometric circular dichroism titrations of the protein and of several molecular variants generated by site-directed mutagenesis have established that the reduction potentials of the two clusters differ widely by almost 200 mV. This large difference has been confirmed by electrochemical methods, and each redox transition has been assigned to one of the clusters. The unusually low potential center is surprisingly the one that displays a conventional CX1X2CX3X4C (Xn, variable amino acid) binding motif and a structural environment similar to that of clusters having less negative potentials. A comparison with other ferredoxins has highlighted factors contributing to the reduction potential of [4Fe-4S] clusters in proteins. (i) The loop between the coordinating cysteines 40 and 49 and the C terminus alpha-helix of C. vinosum ferredoxin cause a negative, but relatively moderate, shift of approximately 60 mV for the nearby cluster. (ii) Very negative potentials, below -600 mV, correlate with the presence of a bulky side chain in position X4 of the coordinating triad of cysteines. These findings set the framework in which previous observations on ferredoxins can be better understood. They also shed light onto the possible occurrence and properties of very low potential [4Fe-4S] clusters in less well characterized proteins.
Atovaquone is a substituted 2-hydroxynaphthoquinone that is used therapeutically to treat Plasmodium falciparum malaria, Pneumocystis carinii pneumonia, and Toxoplasma gondii toxoplasmosis. It is thought to act on these organisms by inhibiting the cytochrome bc1 complex. We have examined the interaction of atovaquone with the bc1 complex isolated from Saccharomyces cerevisiae, a surrogate, nonpathogenic fungus. Atovaquone inhibits the bc1 complex competitively with apparent Ki = 9 nm, raises the midpoint potential of the Rieske iron-sulfur protein from 285 to 385 mV, and shifts the g values in the EPR spectrum of the Rieske center. These results indicate that atovaquone binds to the ubiquinol oxidation pocket of the bc1 complex, where it interacts with the Rieske iron-sulfur protein. A computed energy-minimized structure for atovaquone liganded to the yeast bc1 complex suggests that a phenylalanine at position 275 of cytochrome b in the bovine bc1 complex, as opposed to leucine at the equivalent position in the yeast enzyme, is responsible for the decreased sensitivity of the bovine bc1 complex (Ki = 80 nm) to atovaquone. When a L275F mutation was introduced into the yeast cytochrome b, the sensitivity of the yeast enzyme to atovaquone decreased (Ki = 100 nm) with no loss in activity, confirming that the L275F exchange contributes to the differential sensitivity of these two species to atovaquone. These results provide the first molecular description of how atovaquone binds to the bc1 complex and explain the differential inhibition of the fungal versus mammalian enzymes.
The crystal structure of the bovine Rieske iron-sulfur protein indicates a sulfur atom (S-1) of the iron-sulfur cluster and the sulfur atom (Sgamma) of a cysteine residue that coordinates one of the iron atoms form hydrogen bonds with the hydroxyl groups of Ser-163 and Tyr-165, respectively. We have altered the equivalent Ser-183 and Tyr-185 in the Saccharomyces cerevisiae Rieske iron-sulfur protein by site-directed mutagenesis of the iron-sulfur protein gene to examine how these hydrogen bonds affect the midpoint potential of the iron-sulfur cluster and how changes in the midpoint potential affect the activity of the enzyme. Eliminating the hydrogen bond from the hydroxyl group of Ser-183 to S-1 of the cluster lowers the midpoint potential of the cluster by 130 mV, and eliminating the hydrogen bond from the hydroxyl group of Tyr-185 to Sgamma of Cys-159 lowers the midpoint potential by 65 mV. Eliminating both hydrogen bonds has an approximately additive effect, lowering the midpoint potential by 180 mV. Thus, these hydrogen bonds contribute significantly to the positive midpoint potential of the cluster but are not essential for its assembly. The activity of the bc1 complex decreases with the decrease in midpoint potential, confirming that oxidation of ubiquinol by the iron-sulfur protein is the rate-limiting partial reaction in the bc1 complex, and that the rate of this reaction is extensively influenced by the midpoint potential of the iron-sulfur cluster.
The prize-collecting vehicle routing problem with single and multiple depots and non-linear cost
(2013)
In this paper, we propose a new routing problem to model a highly relevant planning task in small package shipping. We consider the Prize-Collecting Vehicle Routing Problem with Non-Linear cost in its single and multi-depot version, which integrates the option of outsourcing customers to subcontractors instead of serving them with the private fleet. Thereby, a lower bound on the total customer demand to be served by the private fleet guarantees a high utilization of the fleet capacity. To represent the practical situation, where a discount is given by a subcontractor if larger amounts of packages are outsourced, subcontracting costs follow a non-linear function. The considered problem is NP-hard and we propose an Adaptive Variable Neighborhood Search algorithm to solve instances of realistic size. We propose new benchmark sets for the single and the multi-depot problem, which are adapted from test instances of the capacitated VRP and the closely related Multi-Depot VRP with Private fleet and Common carrier. In numerical studies, we investigate the performance of our algorithm on the newly generated test instances and on standard benchmark problems of related problems. Moreover, we study the effect of different cost functions and different values of the minimal demand to be served by the private fleet on the routing solutions obtained.
Background/Objectives: Agility and cognitive abilities are typically assessed separately by different motor and cognitive tests. While many agility tests lack a reactive decision-making component, cognitive assessments are still mainly based on computer-based or paper-pencil tests with low ecological validity. This study is the first to validate the novel SKILLCOURT technology as an integrated assessment tool for agility and cognitive-motor performance.
Methods: Thirty-two healthy adults performed agility (Star Run), reactive agility (Random Star Run) and cognitive-motor (executive function test, 1-back decision making) performance assessments on the SKILLCOURT. Cognitive-motor tests included lower limb responses in a standing position to increase the ecological validity when compared to computer-based tests. Test results were compared to established motor and agility tests (countermovement jump, 10 m linear sprint, T-agility tests) as well as computer-based cognitive assessments (choice-reaction, Go-NoGo, task switching, memory span). Correlation and multiple regression analyses quantified the relation between SKILLCOURT performance and motor and cognitive outcomes.
Results: Star Run and Random Star Run tests were best predicted by linear sprint (r = 0.68, p < 0.001) and T-agility performance (r = 0.77, p < 0.001), respectively. The executive function test performance was well explained by computer-based assessments on choice reaction speed and cognitive flexibility (r = 0.64, p < 0.001). The 1-back test on the SKILLCOURT revealed moderate but significant correlations with the computer-based assessments (r = 0.47, p = 0.007).
Conclusion: The results support the validity of the SKILLCOURT technology for agility and cognitive assessments in more ecologically valid cognitive-motor tasks. This technology provides a promising alternative to existing performance assessment tools.
Perceptual expectations influence perception, attention and the perceptual decision bias during visuospatial orienting, which is impaired in individuals with Autism Spectrum Disorder (ASD). In this study, we investigated whether during visuospatial orienting, perceptual expectations in ASD differentially influence perception, attention and the perceptual decision bias relative to neurotypical controls (NT). Twenty-three children and adolescents with ASD and 23 NT completed a visuospatial orienting task, which compared the effect of a valid relative to an invalid perceptual expectation on target detection (cue validity effect). Group differences were calculated regarding the cue validity effect on neural correlates of processing gain (N1a amplitude) and attention (N1pc amplitude), the perceptual decision bias and mean reaction time (RT). In ASD relative to NT, findings showed a reduced processing gain for validly relative to invalidly cued targets and increased attentional response following invalidly relative to validly cued targets. Increased attention correlated with faster performance across groups. Increased processing correlated with a higher perceptual decision bias and faster mean RT in NT, but not in ASD. Results suggest that during visuospatial orienting, perceptual expectations in ASD may drive changes in sensory processing and stimulus-driven attention, which may differentially guide behavioural responses.
Two new endemic genera of Therevinae are described from Madagascar. Rinhatiana gen. nov. contains three new species (R. arctifestuca gen. et sp. nov., R. cracentis gen. et sp. nov. and R. latifestuca gen. et sp. nov.) as well as R. distincta (Lyneborg, 1976) comb. nov., which is transferred from Stenopomyia Lyneborg, 1976. Tianarinha gen. nov. is described containing two new species, T. goodmani gen. et sp. nov. and T. micet gen. et sp. nov. All species are diagnosed and figured along with distribution data.
Decline in physical activity in the weeks preceding sustained ventricular arrhythmia in women
(2020)
Background: Heightened risk of cardiac arrest following physical exertion has been reported. Among patients with an implantable defibrillator, an appropriate shock for sustained ventricular arrhythmia was preceded by a retrospective self-report of engaging in mild-to-moderate physical activity. Previous studies evaluating the relationship between activity and sudden cardiac arrest lacked an objective measure of physical activity and women were often underrepresented.
Objective: To determine the relationship between physical activity, recorded by accelerometer in a wearable cardioverter-defibrillator (WCD), and sustained ventricular arrhythmia among female patients.
Methods: A dataset of female adult patients prescribed a WCD for a diagnosis of myocardial infarction or dilated cardiomyopathy was compiled from a commercial database. Curve estimation, to include linear and nonlinear interpolation, was applied to physical activity as a function of time (days before arrhythmia).
Results: Among women who received an appropriate WCD shock for sustained ventricular arrhythmia (N = 120), a quadratic relationship between time and activity was present prior to shock. Physical activity increased starting at the beginning of the 30-day period up until day -16 (16 days before the ventricular arrhythmia) when activity begins to decline.
Conclusion: For patients who received treatment for sustained ventricular arrhythmia, a decline in physical activity was found during the 2 weeks preceding the arrhythmic event. Device monitoring for a sustained decline in physical activity may be useful to identify patients at near-term risk of a cardiac arrest.
Homeownership rates differ widely across European countries. We document that part of this variation is driven by differences in the fraction of adults co-residing with their parents. Comparing Germany and Italy, we show that in contrast to homeownership rates per household, homeownership rates per individual are very similar during the first part of the life cycle. To understand these patterns, we build an overlapping-generations model where individuals face uninsurable income risk and make consumption-saving and housing tenure decisions. We embed an explicit intergenerational link between children and parents to capture the three-way trade-off between owning, renting, and co-residing. Calibrating the model to Germany we explore the role of income profiles, housing policies, and the taste for independence and show that a combination of these factors goes a long way in explaining the differential life-cycle patterns of living arrangements between the two countries.