Allosteric drugs, which bind to proteins in regions other than their main ligand-binding or active sites, make it possible to target proteins considered "undruggable" and to develop new therapies ...that circumvent existing resistance. Despite growing interest in allosteric drug discovery, rational design is limited by a lack of sufficient structural information about alternative binding sites in proteins. Previously, we used Markov State Models (MSMs) to identify such "cryptic pockets," and here we describe a method for identifying compounds that bind in these cryptic pockets and modulate enzyme activity. Experimental tests validate our approach by revealing both an inhibitor and two activators of TEM β-lactamase (TEM). To identify hits, a library of compounds is first virtually screened against either the crystal structure of a known cryptic pocket or an ensemble of structures containing the same cryptic pocket that is extracted from an MSM. Hit compounds are then screened experimentally and characterized kinetically in individual assays. We identify three hits, one inhibitor and two activators, demonstrating that screening for binding to allosteric sites can result in both positive and negative modulation. The hit compounds have modest effects on TEM activity, but all have higher affinities than previously identified inhibitors, which bind the same cryptic pocket but were found, by chance, via a computational screen targeting the active site. Site-directed mutagenesis of key contact residues predicted by the docking models is used to confirm that the compounds bind in the cryptic pocket as intended. Because hit compounds are identified from docking against both the crystal structure and structures from the MSM, this platform should prove suitable for many proteins, particularly targets whose crystal structures lack obvious druggable pockets, and for identifying both inhibitory and activating small-molecule modulators.
TEM β-lactamase confers bacteria with resistance to many antibiotics and rapidly evolves activity against new drugs. However, functional changes are not easily explained by differences in crystal ...structures. We employ Markov state models to identify hidden conformations and explore their role in determining TEM's specificity. We integrate these models with existing drug-design tools to create a new technique, called Boltzmann docking, which better predicts TEM specificity by accounting for conformational heterogeneity. Using our MSMs, we identify hidden states whose populations correlate with activity against cefotaxime. To experimentally detect our predicted hidden states, we use rapid mass spectrometric footprinting and confirm our models' prediction that increased cefotaxime activity correlates with reduced Ω-loop flexibility. Finally, we design novel variants to stabilize the hidden cefotaximase states, and find their populations predict activity against cefotaxime in vitro and in vivo. Therefore, we expect this framework to have numerous applications in drug and protein design.
Globally, depression is a leading cause of disability and has remained so for decades. Antidepressant medications have suboptimal outcomes and are too frequently associated with side effects, ...highlighting the need for alternative treatment options. Although primarily known for its robust physical health benefits, exercise is increasingly recognized for its mental health and antidepressant benefits. Empirical evidence indicates that exercise is effective in treating individuals with depression; however, the mechanisms by which exercise exerts anti-depressant effects are not fully understood. Acute bouts of exercise have been shown to transiently modulate circulating levels of serotonin and norepinephrine, brain-derived neurotrophic factor, and a variety of immuno-inflammatory mechanisms in clinical cohorts with depression. However, exercise training has not been demonstrated to consistently modulate such mechanisms, and evidence linking these putative mechanisms and reductions in depression is lacking. The complexity of the biological underpinnings of depression coupled with the intricate molecular cascade induced by exercise are significant obstacles in the attempt to disentangle exercise's effects on depression. Notwithstanding our limited understanding of these effects, clinical evidence uniformly argues for the use of exercise to treat depression. Regrettably, exercise remains underutilized despite being an accessible, low-cost alternative/adjunctive intervention that can simultaneously reduce depression and improve overall health. To address the gaps in our understanding of the clinical and molecular effects of exercise on depression, we propose a model that leverages systems biology and multidisciplinary team science with a large-scale public health investment. Until the science matches the scale of complexity and burden posed by depression, our ability to advance knowledge and treatment will continue to be plagued by fragmented, irreproducible mechanistic findings and no guidelines for standards of care.
Neuromuscular electrical stimulation (NMES) is commonly used in clinical settings to activate skeletal muscle in an effort to mimic voluntary contractions and enhance the rehabilitation of human ...skeletal muscles. It is also used as a tool in research to assess muscle performance and/or neuromuscular activation levels. However, there are fundamental differences between voluntary- and artificial-activation of motor units that need to be appreciated before NMES protocol design can be most effective. The unique effects of NMES have been attributed to several mechanisms, most notably, a reversal of the voluntary recruitment pattern that is known to occur during voluntary muscle contractions. This review outlines the assertion that electrical stimulation recruits motor units in a nonselective, spatially fixed, and temporally synchronous pattern. Additionally, it synthesizes the evidence that supports the contention that this recruitment pattern contributes to increased muscle fatigue when compared with voluntary actions and provides some commentary on the parameters of electrical stimulation as well as emerging technologies being developed to facilitate NMES implementation. A greater understanding of how electrical stimulation recruits motor units, as well as the benefits and limitations of its use, is highly relevant when using this tool for testing and training in rehabilitation, exercise, and/or research.
The solid electrolyte interphase (SEI), a complex layer that forms over the surface of electrodes exposed to battery electrolyte, has a central influence on the structural evolution of the electrode ...during battery operation. For lithium metallic anodes, tailoring this SEI is regarded as one of the most effective avenues for ensuring consistent cycling behavior, and thus practical efficiencies. While fluoride‐rich interphases in particular seem beneficial, how they alter the structural dynamics of lithium plating and stripping to promote efficiency remains only partly understood. Here, operando liquid‐cell transmission electron microscopy is used to investigate the nanoscale structural evolution of lithium electrodeposition and dissolution at the electrode surface across fluoride‐poor and fluoride‐rich interphases. The in situ imaging of lithium cycling reveals that a fluoride‐rich SEI yields a denser Li structure that is particularly amenable to uniform stripping, thus suppressing lithium detachment and isolation. By combination with quantitative composition analysis via mass spectrometry, it is identified that the fluoride‐rich SEI suppresses overall lithium loss through drastically reducing the quantity of dead Li formation and preventing electrolyte decomposition. These findings highlight the importance of appropriately tailoring the SEI for facilitating consistent and uniform lithium dissolution, and its potent role in governing the plated lithium's structure.
The composition of the solid electrolyte interphase (SEI) layer in batteries can be manipulated to improve performance, yet remains challenging to understand. In this work, in situ transmission electron microscopy is used to directly image electrodes as they are cycled, revealing the role that a fluoride‐rich SEI layer has for promoting improved cycling performance.
Electromyostimulation (EMS) incorporates the use of electrical current to activate skeletal muscle and facilitate contraction. It is commonly used in clinical settings to mimic voluntary contractions ...and enhance the rehabilitation of human skeletal muscles. Although the beneficial effects of EMS are widely accepted, discrepancies concerning the specific responses to EMS versus voluntary actions exist. The unique effects of EMS have been attributed to several mechanisms, most notably a reversal of the recruitment pattern typically associated with voluntary muscle activation. This perspective outlines the authors' contention that electrical stimulation recruits motor units in a nonselective, spatially fixed, and temporally synchronous pattern. Furthermore, it synthesizes the evidence that supports the contention that this recruitment pattern contributes to increased muscle fatigue when compared with voluntary actions. The authors believe the majority of evidence suggests that EMS-induced motor unit recruitment is nonselective and that muscle fibers are recruited without obvious sequencing related to fiber types.
Background: Despite extensive study of the impact of stroke on muscle and functional performance, questions remain regarding the extent to which changes are due to the neurological injury vs. ...age-related loss of morphology and force production.
Objectives: To synthesize available evidence describing post-stroke changes in lower extremity muscle size and strength compared to healthy adults.
Methods: Scientific literature was searched up to April 2016 to identify studies that included lower extremity muscle size and strength measures in individuals with chronic stroke. Lower extremity muscle size and strength data from healthy controls were sought for comparison. Relative differences were calculated between paretic, nonparetic, and control limbs.
Results: Fifteen studies with 375 participants (61% male; age = 62 ± 5 years; time since stroke = 60 ± 42 months) were included. The paretic limb exhibited deficits of ~13% in thigh muscle size, ~5% in lower leg muscle size, and ~8% in lean leg mass compared to the nonparetic limb. Paretic plantarflexor and knee extensor strength were 52 and 36% lower, respectively, compared to the nonparetic limb. When compared to age-matched control data, both paretic and nonparetic limbs showed deficits in muscle size and strength.
Conclusions: Age-related differences support the impact of stroke-related sarcopenia as a contributor to hemiparetic muscle dysfunction. Understanding these muscular changes is necessary for designing appropriate exercise interventions aimed at restoring muscle function.
Global climate change is a major threat to biodiversity. Large-scale analyses have generally focused on the impacts of climate change on the geographic ranges of species and on phenology, the timing ...of ecological phenomena. We used long-term monitoring of the abundance of breeding birds across Europe and the United States to produce, for both regions, composite population indices for two groups of species: those for which climate suitability has been either improving or declining since 1980. The ratio of these composite indices, the climate impact indicator (CII), reflects the divergent fates of species favored or disadvantaged by climate change. The trend in CII is positive and similar in the two regions. On both continents, interspecific and spatial variation in population abundance trends are well predicted by climate suitability trends.
Neuromuscular electrical stimulation (NMES) has been shown to impart a number of health benefits and can be used to produce functional outcomes. However, one limitation of NMES is the onset of ...NMES-induced fatigue. Multi-channel asynchronous stimulation has been shown to reduce NMES-induced fatigue compared to conventional single-channel stimulation. However, in previous studies in man, the effect of stimulation frequency on the NMES-induced fatigue has not been examined for asynchronous stimulation. Low stimulation frequencies are known to reduce fatigue during conventional stimulation. Therefore, the aim of this study was to examine the fatigue characteristics of high- and low-frequency asynchronous stimulation as well as high- and low-frequency conventional stimulation. Experiments were performed in both able-bodied and spinal cord injured populations. Low frequency asynchronous stimulation is found to have significant fatigue benefits over high frequency asynchronous stimulation as well as high- and low-frequency conventional stimulation, motivating its use for rehabilitation and functional electrical stimulation (FES).
Electromechanical delay (EMD) is a biological artifact that arises due to a time lag between electrical excitation and tension development in a muscle. EMD is known to cause degraded performance and ...instability during neuromuscular electrical stimulation (NMES). Compensating for such input delay is complicated by the unknown nonlinear muscle force-length and muscle force-velocity relationships. This paper provides control development and a mathematical stability analysis of a NMES controller with a predictive term that actively accounts for EMD. The results are obtained through the development of a novel predictor-type method to address the delay in the voltage input to the muscle. Lyapunov-Krasovskii functionals are used within a Lyapunov-based stability analysis to prove semi-global uniformly ultimately bounded tracking. Experiments on able-bodied volunteers illustrate the performance and robustness of the developed controller during a leg extension trajectory following task.
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