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Despite the success of genomics in identifying new essential bacterial genes, there is a lack of sustainable leads in antibacterial drug discovery to address increasing multidrug resistance. Type IIA topoisomerases cleave and religate DNA to regulate DNA topology and are a major class of antibacterial and anticancer drug targets, yet there is no well developed structural basis for understanding drug action. Here we report the 2.1 Å crystal structure of a potent, new class, broad-spectrum antibacterial agent in complex with Staphylococcus aureus DNA gyrase and DNA, showing a new mode of inhibition that circumvents fluoroquinolone resistance in this clinically important drug target. The inhibitor ‘bridges’ the DNA and a transient non-catalytic pocket on the two-fold axis at the GyrA dimer interface, and is close to the active sites and fluoroquinolone binding sites. In the inhibitor complex the active site seems poised to cleave the DNA, with a single metal ion observed between the TOPRIM (topoisomerase/primase) domain and the scissile phosphate. This work provides new insights into the mechanism of topoisomerase action and a platform for structure-based drug design of a new class of antibacterial agents against a clinically proven, but conformationally flexible, enzyme class. Topoisomerase inhibition Enzymes that move along a DNA strand, such as DNA and RNA polymerases, tend to cause the build-up of supercoiling ahead of their motion. Unchecked, this would cause the DNA to become overwound, like a twisted rubber band. Topoisomerases relieve this stress by first cleaving and then re-ligating the DNA. Topoisomerase inhibitors are used as antibacterial and anticancer drugs — for example, antibacterials of the quinolone family have been in clinical use since 1962, but are now compromised by the emergence of multidrug-resistant bacteria. The crystal structure of a type II topoisomerase from Staphylococcus aureus , DNA gyrase, has now been determined in a complex with DNA and with the broad-spectrum antibiotic GSK299423. This is an example of a new class of antibiotics that interact with the same targets as fluoroquinolones, but are structurally and mechanistically distinct from them. The structure reveals a mechanism that circumvents fluoroquinolone resistance and opens up strategies of exploiting alternative inhibition mechanisms for clinically validated targets. Enzymes that move along DNA, such as DNA and RNA polymerases, cause the DNA ahead of them to become supercoiled. This would lead to the DNA becoming overwound, were the stress not relieved by topoisomerases. Topoisomerase inhibitors have been used as antibacterial and anticancer drugs, but the structural basis for their activity has been unclear. Here, the crystal structures are presented of a topoisomerase on DNA, either alone or in the presence of a new type of antibiotic.

As Pogo once said, \"We have met the enemy and he is us.\" In this book the author offers a scathing assessment of fiscal blunders in foreign lands, and details how economic repercussions are sure to be felt on American soil. Financial bubbles grew and burst, not only in the U.S. but in countries as diverse as Iceland, Germany, and Greece. Mixing humor with prescient insight, he depicts a precarious situation that demands attention. The tsunami of cheap credit that rolled across the planet between 2002 and 2008 was more than a simple financial phenomenon: it was temptation, offering entire societies the chance to reveal aspects of their characters they could not normally afford to indulge. Icelanders wanted to stop fishing and become investment bankers. The Greeks wanted to turn their country into a piنnata stuffed with cash and allow as many citizens as possible to take a whack at it. The Germans wanted to be even more German; the Irish wanted to stop being Irish. This investigation of bubbles beyond our shores is so sadly hilarious that it leads the American reader to a comfortable complacency: oh, those foolish foreigners. But when he turns a merciless eye on California and Washington, D.C., we see that the narrative is a trap baited with humor, and we understand the reckoning that awaits the greatest and greediest of debtor nations. - Publisher.

During low arousal states such as drowsiness and sleep, cortical neurons exhibit rhythmic slow wave activity associated with periods of neuronal silence. Slow waves are locally regulated, and local slow wave dynamics are important for memory, cognition, and behaviour. While several brainstem structures for controlling global sleep states have now been well characterized, a mechanism underlying fast and local modulation of cortical slow waves has not been identified. Here, using optogenetics and whole cortex electrophysiology, we show that local tonic activation of thalamic reticular nucleus (TRN) rapidly induces slow wave activity in a spatially restricted region of cortex. These slow waves resemble those seen in sleep, as cortical units undergo periods of silence phase-locked to the slow wave. Furthermore, animals exhibit behavioural changes consistent with a decrease in arousal state during TRN stimulation. We conclude that TRN can induce rapid modulation of local cortical state. We usually think of sleep as a global state: that the entire brain is either asleep or awake. However, recent evidence has suggested that smaller regions of the brain can show sleep-like activity while the rest of the brain remains awake. It is not clear why or how these sleep-like patterns of brain activity appear, and whether they are related to the drowsy behaviour that occurs when one is about to fall asleep. Lewis, Voigts et al. investigated how this process works in mice using a technique called optogenetics. This technique makes it possible to genetically engineer mice so that the activity of particular areas of the brain can be switched on or off by light. Lewis, Voigts et al. used light to stimulate different regions of the brain and tracked the resulting brain activity using tiny electrodes that are capable of detecting the activity of individual neurons. The experiments show that stimulating one part of a deep brain structure called the thalamic reticular nucleus causes just one small part of the brain to switch from being awake to producing sleep-like brain wave patterns. When a larger area is stimulated, the whole brain switches into this sleep-like pattern. Stimulation of the thalamic reticular nucleus also caused the animals to become drowsy and they were more likely to fall asleep, which suggests that sleep-like activity in small parts of the brain may contribute to drowsiness. Lewis, Voigts et al.’s findings identify a brain switch that can influence whether an animal is awake or asleep. Importantly, they show that sleep can be independently controlled in small brain regions, and that the thalamic reticular nucleus contains a ‘map’ that allows it to induce sleep in just one region, or across the whole brain. Memories are strengthened during sleep, so the next challenge is to study whether the thalamic reticular nucleus influences memory formation. The findings also suggest that further study of this brain region may be useful for understanding how the sleep and awake states are controlled by particular neurons.

An analysis of five financial upheavals in recent history includes coverage of the 1987 stock market crash, the 1998 Russian default (and the consequent collapse of U.S. hedge fund Long-Term Capital Management), the Asian currency crisis of 1999, the Internet bubble of 1995-2001, and the 2008 sub-prime mortgage crisis, in an anecdotal report that reveals how public knowledge differed from what was actually taking place.

The Escherichia coli DNA replication machinery has been used as a road map to uncover design rules that enable DNA duplication with high efficiency and fidelity. Although the enzymatic activities of the replicative DNA Pol III are well understood, its dynamics within the replisome are not. Here, we test the accepted view that the Pol III holoenzyme remains stably associated within the replisome. We use in vitro single-molecule assays with fluorescently labeled polymerases to demonstrate that the Pol III* complex (holoenzyme lacking the β2 sliding clamp), is rapidly exchanged during processive DNA replication. Nevertheless, the replisome is highly resistant to dilution in the absence of Pol III* in solution. We further show similar exchange in live cells containing labeled clamp loader and polymerase. These observations suggest a concentration-dependent exchange mechanism providing a balance between stability and plasticity, facilitating replacement of replisomal components dependent on their availability in the environment.

Protein disulfide isomerase (PDI) is a chaperone protein in the endoplasmic reticulum that is up-regulated in mouse models of, and brains of patients with, neurodegenerative diseases involving protein misfolding. PDI’s role in these diseases, however, is not fully understood. Here, we report the discovery of a reversible, neuroprotective lead optimized compound (LOC)14, that acts as a modulator of PDI. LOC14 was identified using a high-throughput screen of ∼10,000 lead-optimized compounds for potent rescue of viability of PC12 cells expressing mutant huntingtin protein, followed by an evaluation of compounds on PDI reductase activity in an in vitro screen. Isothermal titration calorimetry and fluorescence experiments revealed that binding to PDI was reversible with a K d of 62 nM, suggesting LOC14 to be the most potent PDI inhibitor reported to date. Using 2D heteronuclear single quantum correlation NMR experiments, we were able to map the binding site of LOC14 as being adjacent to the active site and to observe that binding of LOC14 forces PDI to adopt an oxidized conformation. Furthermore, we found that LOC14-induced oxidation of PDI has a neuroprotective effect not only in cell culture, but also in corticostriatal brain slice cultures. LOC14 exhibited high stability in mouse liver microsomes and blood plasma, low intrinsic microsome clearance, and low plasma-protein binding. These results suggest that LOC14 is a promising lead compound to evaluate the potential therapeutic effects of modulating PDI in animal models of disease. Significance Protein disulfide isomerase (PDI) is a chaperone protein in the endoplasmic reticulum. It is up-regulated in mouse models of, and brains of patients with, neurological protein folding diseases. Irreversible inhibition of PDI activity by the small molecule 16F16 results in protection in cell and organotypic brain slice culture models of Huntington disease. Here, we identified lead optimized compound (LOC)14 as a nanomolar, reversible inhibitor of PDI that protects PC12 cells and medium spiny neurons from the toxic mutant huntingtin protein. LOC14 has improved potency compared with 16F16 and displays favorable pharmaceutical properties, making it a suitable compound to evaluate the therapeutic potential of inhibiting PDI in multiple disease models.

The explosive story of how one group of ingenious oddballs and misfits set out to expose what was going on. It's the story of what it's like to declare war on some of the rishest and most powerful people in the world.

River systems once safeguarded from water development are being developed. This includes intermittent rivers that annually dry to a series of pools. Describing fish species relationships between abundance and pool depth can help managers set water-take rules that protect fish in dry-season pools. We sampled fish in main-channel and floodplain pools that spanned a gradient of depths and overcame sampling challenges by accounting for interacting effects of species mean length, environmental attributes, and sampling attributes on fish capture probabilities. Fish abundance-depth relationships varied systematically with species mean length, mesohabitat type (main channel, floodplain), water turbidity, and structural complexity, highlighting system complexity and the potential generality of abundance-depth relationships. Similarly, fish length moderated the effects of environmental attributes on capture probability for all sampling methods. We evaluated impacts of hypothetical water-take regulations on fish species’ distributions. Results suggested that water-take rules prohibiting draining of main-channel pools below 1.65 m and reducing floodplain pools by no more than 14% minimises impacts to species’ distributions, promoting conservation of the fish community. Additionally, our approach demonstrates the capacity of species length for predicting distributional and sampling patterns of fish species.