Crystal structure of a monomeric form of SARS-CoV endonuclease nsp15 suggests a role for hexamerization as an allosteric switch

Mature nonstructural protein-15 (nsp15) from the severe acute respiratory syndrome coronavirus (SARS-CoV) contains a novel uridylate-specific Mn2+-dependent endoribonuclease (NendoU). Structure studies of the full-length form of the obligate hexameric enzyme from two CoVs, SARS-CoV and murine hepatitis virus, and its monomeric homologue, XendoU from Xenopus laevis, combined with mutagenesis studies have implicated several residues in enzymatic activity and the N-terminal domain as the major determinant of hexamerization. However, the tight link between hexamerization and enzyme activity in NendoUs has remained an enigma. Here, we report the structure of a trimmed, monomeric form of SARS-CoV nsp15 (residues 28 to 335) determined to a resolution of 2.9 A. The catalytic loop (residues 234 to 249) with its two reactive histidines (His 234 and His 249) is dramatically flipped by approximately 120 degrees into the active site cleft. Furthermore, the catalytic nucleophile Lys 289 points in a diametrically opposite direction, a consequence of an outward displacement of the supporting loop (residues 276 to 295). In the full-length hexameric forms, these two loops are packed against each other and are stabilized by intimate intersubunit interactions. Our results support the hypothesis that absence of an adjacent monomer due to deletion of the hexamerization domain is the most likely cause for disruption of the active site, offering a structural basis for why only the hexameric form of this enzyme is active.

Direct observation by time-resolved infrared spectroscopy of the bright and the dark excited states of the [Ru(phen)2(dppz)]2+ light-switch compound in solution and when bound to DNA

The [Ru(phen)2(dppz)]2+ complex (1) is non-emissive in water but is highly luminescent in organic solvents or when bound to DNA, making it a useful probe for DNA binding. To date, a complete mechanistic explanation for this “light-switch” effect is still lacking. With this in mind we have undertaken an ultrafast time resolved infrared (TRIR) study of 1 and directly observe marker bands between 1280–1450 cm-1, which characterise both the emissive “bright” and the non-emissive “dark” excited states of the complex, in CD3CN and D2O respectively. These characteristic spectral features are present in the [Ru(dppz)3]2+ solvent light-switch complex but absent in [Ru(phen)3]2+, which is luminescent in both solvents. DFT calculations show that the vibrational modes responsible for these characteristic bands are predominantly localised on the dppz ligand. Moreover, they reveal that certain vibrational modes of the “dark” excited state couple with vibrational modes of two coordinating water molecules, and through these to the bulk solvent, thus providing a new insight into the mechanism of the light-switch effect. We also demonstrate that the marker bands for the “bright” state are observed for both L- and D enantiomers of 1 when bound to DNA and that photo-excitation of the complex induces perturbation of the guanine and cytosine carbonyl bands. This perturbation is shown to be stronger for the L enantiomer, demonstrating the different binding site properties of the two enantiomers and the ability of this technique to determine the identity and nature of the binding site of such intercalators.

Bone Remodeling Adjacent to Morse Cone-Connection Implants with Platform Switch: A Fluorescence Study in the Dog Mandible

Purpose: The aim of this study was to evaluate, through fluorescence analysis, the effect that different interimplant distances, after prosthetic restoration, will have on bone remodeling in submerged and nonsubmerged implants restored with a ""platform switch."" Materials and Methods: Fifty-six Ankylos implants were placed 1.5 mm subcrestally in seven dogs. The implants were placed so that two fixed prostheses, with three interimplant contacts separated by 1-mm, 2-mm, and 3-mm distances, could be fabricated for each side of the mandible. The sides and the positions of the groups were selected randomly. To better evaluate bone remodeling, calcein green was injected 3 days before placement of the prostheses at 12 weeks postimplantation. At 3 days before sacrifice (8 weeks postloading), alizarin red was injected. The amounts of remodeled bone within the different interimplant areas were compared statistically before and after loading in submerged and nonsubmerged implants. Results: Statistically significant differences existed in the percentage of remodeled bone seen in the different regions. Mean percentages of remodeled bone in the submerged and nonsubmerged groups, respectively, were as follows: for the 1-mm distance, 23.0% +/- 0.05% and 23.1% +/- 0.03% preloading and 27.0% +/- 0.03% and 25.2% +/- 0.04% postloading, for the 2-mm distance, 18.2% +/- 0.05% and 18.1% +/- 0.04% preloading and 21.3% +/- 0.07% and 19.9% +/- 0.03% postloading, for the 3-mm distance, 18.3% +/- 0.03% and 18.3% +/- 0.03% preloading and 18.8% +/- 0.04% and 19.8% +/- 0.04% postloading, for distal-extension regions, 16.6% +/- 0.02% and 17.4% +/- 0.04% preloading and 17.0% +/- 0.04% and 18.4% +/- 0.04% postloading. Conclusions: Based upon this animal study, loading increases bone formation for submerged or nonsubmerged implants, and the interimplant distance of 1 mm appears to result in more pronounced bone remodeling than the 2-mm or 3-mm distances in implants with a ""platform switch."" INT J ORAL MAXILLOFAC IMPLANTS 2009;24:257-266

A Single Mutation at the Sheet Switch Region Results in Conformational Changes Favoring lambda 6 Light-Chain Fibrillogenesis

Systemic amyloid light-chain (LC) amyloidosis is a disease process characterized by the pathological deposition of monoclonal LCs in tissue. All LC subtypes are capable of fibril formation although lambda chains, particularly those belonging to the lambda 6 type, are overrepresented. Here, we report the thermodynamic and in vitro fibrillogenic properties of several mutants of the lambda 6 protein 6aJL2 in which Pro7 and/or His8 was substituted by Ser or Pro. The H8P and H8S mutants were almost as stable as the wildtype protein and were poorly fibrillogenic. In contrast, the P7S mutation decreased the thermodynamic stability of 6aJL2 and greatly enhanced its capacity to form amyloid-like fibrils in vitro. The crystal structure of the P7S mutant showed that the substitution induced both local and long-distance effects, such as the rearrangement of the V(L) (variable region of the light chain)-V(L) interface. This mutant crystallized in two orthorhombic polymorphs, P2(1)2(1)2(1) and C222(1). In the latter, a monomer that was not arranged in the typical Bence-Jones dimer was observed for the first time. Crystal-packing analysis of the C222(1) lattice showed the establishment of intermolecular beta-beta interactions that involved the N-terminus and beta-strand B and that these could be relevant in the mechanism of LC fibril formation. Our results strongly suggest that Pro7 is a key residue in the conformation of the N-terminal sheet switch motif and, through long-distance interactions, is also critically involved in the contacts that stabilized the V(L) interface in lambda 6 LCs. (C) 2009 Elsevier Ltd. All rights reserved.

Load balancing solution and evaluation of F5 content switch equipment

The Thesis focused on hardware based Load balancing solution of web traffic through a load balancer F5 content switch. In this project, the implemented scenario for distributing HTTPtraffic load is based on different CPU usages (processing speed) of multiple member servers.Two widely used load balancing algorithms Round Robin (RR) and Ratio model (weighted Round Robin) are implemented through F5 load balancer. For evaluating the performance of F5 content switch, some experimental tests has been taken on implemented scenarios using RR and Ratio model load balancing algorithms. The performance is examined in terms of throughput (bits/sec) and Response time of member servers in a load balancing pool. From these experiments we have observed that Ratio Model load balancing algorithm is most suitable in the environment of load balancing servers with different CPU usages as it allows assigning the weight according to CPU usage both in static and dynamic load balancing of servers.

Biodistribution of the Radiopharmaceutical Sodium Pertechnetate after Biliopancreatic Bypass with a Duodenal Switch

Study with the purpose to examine the effects of duodenal switch (DS), regularly performed in morbidly obese patients, on biodistribution of sodium pertechnetate in several organs of rats. There was no early or late mortality in either rats groups. The values of percent radioactivity per gram of tissue (%ATI/g), showed no significant difference in liver, stomach, small bowel, duodenum, kidney, heart, bladder, bone and brain, when compared the DS rats with sham and controls rats. A postoperative significant increase (p<0.05) in mean %ATI/g levels was observed in spleen, pancreas and muscle in group DS rats, as compared to group S and C rats. In the lung there was an increase and in thyroid a decrease in mean %ATI/g of DS rats, when compared to sham rats (p<0.05). In conclusion, the biliopancreatic diversion with duodenal switch in rats modified the biodistribution of sodium pertechnetate in thyroid, lung, pancreas, spleen and muscle

Biodistribuição do pertecnetato de sódio após cirurgia do Switch Duodenal

The biliopancretic diversion with duodenal switch is one of the mixing techniques used in the treatment of morbid obesity. The duodenal switch reduces the stomach capacity and leaves only 50-100 cm of small intestine for nutrition and absorption. The surgery produces hormonal, structural and biochemical changes, which can influence on the result of scintigraphic examinations in operated patients. With the objective of evaluate the postoperative biodistribution of sodium pertechnetate (Na99mTcO4) in brain, thyroid, heart, lung, liver, spleen, kidney, stomach, duodenum, pancreas, small intestine, bladder, muscle and bone of Wistar rats. The rats were randomly allocated into 3 groups of 7 rats each: the duodenal switch group (DS), the control group (C) and the sham group (S). They were operated under anesthesia and aseptic technique. In the tenth postoperative day, 0.1mL of sodium pertechnetate was injected via orbital plexus. After 30 min the animals were killed with overdose of anesthetic and samples of liver, spleen, pancreas, stomach, duodenum, small intestine, thyroid, lung, heart, kidney, bladder, muscle, bone and brain were harvested, washed with saline and weighed. The detention of radioactivity was made using the automatic Gamma Counter Wizard, PerkinElmer and the percentage of activity per gram of tissue (%ATI/g) was calculated. There was no early or late mortality in either rats groups. The values of percent radioactivity per gram of tissue (%ATI/g), showed no significant difference in liver, stomach, small bowel, duodenum, kidney, heart, bladder, bone and brain, when compared the DS rats with sham and controls rats. A postoperative significant increase in mean %ATI/g levels was observed in spleen, pancreas and muscle in group DS rats, as compared to group S and C rats (p<0.05). In the lung there was an increase and in thyroid a decrease in mean %ATI/g of DS rats, when compared to sham rats (p>0.05). In conclusion, the biliopancreatic diversion with duodenal switch in rats modified the biodistribution of sodium pertechnetate in thyroid, lung, pancreas, spleen and muscle. The study had the participation of the departments and laboratories researches, as Nucleus of Experimental Surgery, Department of Surgery, Laboratory of Radiobiology, Department of Pathology and Service of Nuclear Medicine, certifying the character of a multidisciplinary research

A unity power factor multiple isolated outputs switching mode power supply using a single switch

The authors present an offline switching power supply with multiple isolated outputs and unity power factor with the use of only one power processing stage, based on the DC-DC SEPIC (single ended primary inductance converter) modulated by variable hysteresis current control. The principle of operation, the theoretical analysis, the design procedure, an example, and simulation results are presented. A laboratory prototype, rated at 160 W, operating at a maximum switching frequency of 100 kHz, with isolated outputs rated at +5 V/15 A -5 V/1 A, +12 V/6 A and -12 V/1 A, has been built given an input power factor near unity.

Optimal switch allocation for automatic load transfer in distribution substations

This work proposes a methodology for optimized allocation of switches for automatic load transfer in distribution systems in order to improve the reliability indexes by restoring such systems which present voltage classes of 23 to 35 kV and radial topology. The automatic switches must be allocated on the system in order to transfer load remotely among the sources at the substations. The problem of switch allocation is formulated as nonlinear constrained mixed integer programming model subject to a set of economical and physical constraints. A dedicated Tabu Search (TS) algorithm is proposed to solve this model. The proposed methodology is tested for a large real-life distribution system. © 2011 IEEE.

An iterated sample construction with path relinking method: Application to switch allocation in electrical distribution networks

We present a metaheuristic approach which combines constructive heuristics and local searches based on sampling with path relinking. Its effectiveness is demonstrated by an application to the problem of allocating switches in electrical distribution networks to improve their reliability. Our approach also treats the service restoration problem, which has to be solved as a subproblem, to evaluate the reliability benefit of a given switch allocation proposal. Comparisons with other metaheuristics and with a branch-and-bound procedure evaluate its performance. © 2012 Published by Elsevier Ltd.

Switch allocation problems in power distribution systems

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Optimization of Amplifier Placements in Switch-Based Optical Networks

Wavelength division multiplexing (WDM) offers a solution to the problem of exploiting the large bandwidth on optical links; it is the current favorite multiplexing technology for optical communication networks. Due to the high cost of an optical amplifier, it is desirable to strategically place the amplifiers throughout the network in a way that guarantees that all the signals are adequately amplified while minimizing the total number amplifiers being used. Previous studies all consider a star-based network. This paper demonstrates an original approach for solving the problem in switch-based WDM optical network assuming the traffic matrix is always the permutation of the nodes. First we formulate the problem by choosing typical permutations which can maximize traffic load on individual links; then a GA (Genetic Algorithm) is used to search for feasible amplifier placements. Finally, by setting up all the lightpaths without violating the power constaints we confirm the feasibility of the solution.