Induction of flexibility through protein-protein interactions

The dimerization/docking (D/D) domain of the cyclic AMP-dependent protein kinase (PKA) holoenzyme mediates important protein-protein interactions that direct the subcellular localization of the enzyme. A kinase anchoring proteins (AKAPs) provide the molecular scaffold for the localization of PKA. The recent solution structures of two D/D AKAP complexes revealed that the AKAP binds to a surface-exposed, hydrophobic groove on the D/D. In the present study, we present an analysis of the changes in hydrogen/deuterium exchange protection and internal motions of the backbone of the D/D when free and bound to the prototype anchoring protein, Ht31(pep). We observe that formation of the complex results in significant, but small, increases in H/D exchange protection factors as well as increases in backbone flexibility, throughout the D/D, and in particular, in the hydrophobic binding groove. This unusual observation of increased backbone flexibility and marginal H/D exchange protection, despite high affinity protein-ligand interactions, may be a general effect observed for the stabilization of hydrophobic ligand/hydrophobic pocket interactions.     

Understanding protein flexibility through dimensionality reduction.

This work shows how to decrease the complexity of modeling flexibility in proteins by reducing the number of dimensions necessary to model important macromolecular motions such as the induced-fit process. Induced fit occurs during the binding of a protein to other proteins, nucleic acids, or small molecules (ligands) and is a critical part of protein function. It is now widely accepted that conformational changes of proteins can affect their ability to bind other molecules and that any progress in modeling protein motion and flexibility will contribute to the understanding of key biological functions. However, modeling protein flexibility has proven a very difficult task. Experimental laboratory methods, such as x-ray crystallography, produce rather limited information, while computational methods such as molecular dynamics are too slow for routine use with large systems. In this work, we show how to use the principal component analysis method, a dimensionality reduction technique, to transform the original high-dimensional representation of protein motion into a lower dimensional representation that captures the dominant modes of motions of proteins. For a medium-sized protein, this corresponds to reducing a problem with a few thousand degrees of freedom to one with less than fifty. Although there is inevitably some loss in accuracy, we show that we can obtain conformations that have been observed in laboratory experiments, starting from different initial conformations and working in a drastically reduced search space.     

Fine-tuning GPCR-mediated neuromodulation by biasing signaling through different G protein subunits

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Formins regulate actin filament flexibility through long range allosteric interactions

The members of the formin family nucleate actin polymerization and play essential roles in the regulation of the actin cytoskeleton during a wide range of cellular and developmental processes. In the present work, we describe the effects of mDia1-FH2 on the conformation of actin filaments by using a temperature-dependent fluorescence resonance energy transfer method. Our results revealed that actin filaments were more flexible in the presence than in the absence of formin. The effect strongly depends on the mDia1-FH2 concentration in a way that indicates that more than one mechanism is responsible for the formin effect. In accordance with the more flexible filament structure, the thermal stability of actin decreased and the rate of phosphate dissociation from actin filaments increased in the presence of formin. The interpretation of the results supports a model in which formin binding to barbed ends makes filaments more flexible through long range allosteric interactions, whereas binding of formin to the sides of the filaments stabilizes the protomer-protomer interactions. These results suggest that formins can regulate the conformation of actin filaments and may thus also modulate the affinity of actin-binding proteins to filaments nucleated/capped by formins.     

The intrinsic flexibility of the Kv voltage sensor and its implications for channel gating

Analysis of the crystal structures of the intact voltage-sensitive potassium channel KvAP (from Aeropyrum pernix) and Kv1.2 (from rat brain), along with the isolated voltage sensor (VS) domain from KvAP, raises the question of the exact nature of the voltage-sensing conformational change that triggers activation of Kv and related voltage-gated channels. Molecular dynamics simulations of the isolated VS of KvAP in a detergent micelle environment at two different temperatures (300 K and 368 K) have been used to probe the intrinsic flexibility of this domain on a tens-of-nanoseconds timescale. The VS contains a positively charged (S4) helix which is packed against a more hydrophobic S3 helix. The simulations at elevated temperature reveal an intrinsic flexibility/conformational instability of the S3a region (i.e., the C-terminus of the S3 helix). It is also evident that the S4 helix undergoes hinge bending and swiveling about its central I130 residue. The conformational instability of the S3a region facilitates the motion of the N-terminal segment of S4 (i.e., S4a). These simulations thus support a gating model in which, in response to depolarization, an S3b-S4a "paddle" may move relative to the rest of the VS domain. The flexible S3a region may in turn act to help restore the paddle to its initial conformation upon repolarization.     

Increasing Knowledge of HIV Infection Status through Opt-Out Testing

The diagnosis of HIV infection is the point of entry for treatment and prevention services, yet many infected persons in both developed and developing countries remain undiagnosed. To reduce the number of undiagnosed infections, a variety of expanded testing policies have been recommended, including opt-out testing. This testing model assumes that in populations of increased HIV prevalence, voluntary testing should be offered to all patients seen in healthcare settings and performed unless patients specifically decline. While this approach raises ethical issues concerning “voluntariness”, access to care, and stigma, the potential benefits of opt-out testing far outweigh its potential adverse effects.     

Serotonergic neuromodulation of peripheral nociceptors

Nociception, the encoding and processing of noxious environmental stimuli by sensory neurons, functions to protect an organism from bodily damage. Activation of the terminal endings of certain sensory neurons, termed nociceptors, triggers a train of impulses to neurons in the spinal cord. Signals are integrated and processed in the dorsal spinal cord and then projected to the brain where they elicit the perception of pain. A number of neuromodulators that can affect nociceptors are released in the periphery during the inflammation that follows an initial injury. Serotonin (5-HT) is a one such proinflammatory mediator. This review discusses our current understanding of the neuromodulatory role of 5-HT, and specifically how this monoamine activates and sensitizes nociceptors. Potential therapeutic targets to treat pain are described.     

Improved pKa calculations through flexibility based sampling of a water-dominated interaction scheme

Ionizable groups play critical roles in biological processes. Computation of pK(a)s is complicated by model approximations and multiple conformations. Calculated and experimental pK(a)s are compared for relatively inflexible active-site side chains, to develop an empirical model for hydration entropy changes upon charge burial. The modification is found to be generally small, but large for cysteine, consistent with small molecule ionization data and with partial charge distributions in ionized and neutral forms. The hydration model predicts significant entropic contributions for ionizable residue burial, demonstrated for components in the pyruvate dehydrogenase complex. Conformational relaxation in a pH-titration is estimated with a mean-field assessment of maximal side chain solvent accessibility. All ionizable residues interact within a low protein dielectric finite difference (FD) scheme, and more flexible groups also access water-mediated Debye-Hückel (DH) interactions. The DH method tends to match overall pH-dependent stability, while FD can be more accurate for active-site groups. Tolerance for side chain rotamer packing is varied, defining access to DH interactions, and the best fit with experimental pK(a)s obtained. The new (FD/DH) method provides a fast computational framework for making the distinction between buried and solvent-accessible groups that has been qualitatively apparent from previous work, and pK(a) calculations are significantly improved for a mixed set of ionizable residues. Its effectiveness is also demonstrated with computation of the pH-dependence of electrostatic energy, recovering favorable contributions to folded state stability and, in relation to structural genomics, with substantial improvement (reduction of false positives) in active-site identification by electrostatic strain.     

Uncertainty, neuromodulation, and attention

Uncertainty in various forms plagues our interactions with the environment. In a Bayesian statistical framework, optimal inference and prediction, based on unreliable observations in changing contexts, require the representation and manipulation of different forms of uncertainty. We propose that the neuromodulators acetylcholine and norepinephrine play a major role in the brain's implementation of these uncertainty computations. Acetylcholine signals expected uncertainty, coming from known unreliability of predictive cues within a context. Norepinephrine signals unexpected uncertainty, as when unsignaled context switches produce strongly unexpected observations. These uncertainty signals interact to enable optimal inference and learning in noisy and changeable environments. This formulation is consistent with a wealth of physiological, pharmacological, and behavioral data implicating acetylcholine and norepinephrine in specific aspects of a range of cognitive processes. Moreover, the model suggests a class of attentional cueing tasks that involve both neuromodulators and shows how their interactions may be part-antagonistic, part-synergistic.     

Increasing seaweed crop yields through organic fertilisation at the nursery stage

Extractive seaweed aquaculture is gaining attention in the western Baltic Sea and in particular the co-cultivation with other species for bioremediation or nutrient delivery. However, there are still limitations to viable seaweed production yields in a brackish habitat with a short production period for Saccharina latissima. This investigation presents the specific growth-enhancing effect of Mytilus edulis on the seaweed early nursery stages during the hatchery and during the grow out period at sea in a Baltic fjord. Gametogenesis and juvenile sporophyte development were evaluated with and without blue mussels during 9 weeks of seaweed hatchery. The presence of mussels resulted in a significantly higher abundance of large multicellular sporophytes. After the hatchery period, seedling lines were transferred into the field and installed both in the direct vicinity of and 25 m away from mussel culture ropes. The previously observed supporting effect of mussel co-culture on seaweed development during the hatchery period was still visible after 6 months at sea. Sporophytes were larger, had a higher biomass and had higher carbon content if previously combined with mussels in the hatchery. This investigation suggests that the co-cultivation of seaweed and mussels during seaweed hatchery can increase seaweed crop yields in the following grow out period at sea, with the possibility of being certified organic.     

Increasing rigor in NMR-based metabolomics through validated and open source tools

Opportunities and challenges ahead for NMR-based metabolomics.

     

Process flexibility and inventory flexibility via product substitution

This paper studies the optimal control of and interaction between two types of flexibility under Markov models of demand and production: process flexibility and inventory flexibility. In our model, process flexibility is generated by a multi-functional production facility that can produce two types of products, and inventory flexibility is manifested in firm-driven one-way product substitution. Both process flexibility and inventory flexibility are important drivers of supply chain performance and are strategic design considerations. To analyze the interaction between these two types of flexibility, we model a dynamically controlled two-product, make-to-stock system with stochastic processing times and stochastic demand. We characterize the complex joint optimal production and post-production policy for a special case and numerically show that a simply structured multi-threshold policy is a near-optimal heuristic policy for the general case. We gain further insight into the impact of system parameters on the value of process flexibility and inventory flexibility via a comprehensive numerical study. We find that for a wide range of capacity and cost parameters, process flexibility and inventory flexibility complement each other, so pursuing both forms of flexibility is effective.     

Defining the role of astrocytes in neuromodulation

Astrocytes undergo elevations in intracellular calcium following activation of metabotropic receptors, which may trigger glutamate secretion and excitation of surrounding neurons. In this issue of Neuron, Fiacco et al. use transgenic mice that express a foreign G(q)-coupled receptor in astrocytes to show that selective stimulation of astrocytes is not sufficient to induce the release of glutamate.