Toward Understanding the Molecular Bases of Stretch Activation: A Structural Comparison Of The Two Troponin C Isoforms Of Lethocerus

Muscles are usually activated by calcium binding to the calcium sensory protein troponin-C, which is one of the three components of the troponin complex. However, in cardiac and insect flight muscle activation is also produced by mechanical stress. Little is known about the molecular bases of this calcium-independent activation. In Lethocerus, a giant water bug often used as a model system because of its large muscle fibers, there are two troponin-C isoforms, called F1 and F2, that have distinct roles in activating the muscle. It has been suggested that this can be explained either by differences in structural features or by differences in the interactions with other proteins. Here we have compared the structural and dynamic properties of the two proteins and shown how they differ. We have also mapped the interactions of the F2 isoform with peptides spanning the sequence of its natural partner, troponin-I. Our data have allowed us to build a model of the troponin complex and may eventually help in understanding the specialized function of the F1 and F2 isoforms and the molecular mechanism of stretch activation

Towards Understanding the Molecular Bases of Stretch Activation: A Structural comparison of the Two Troponin C Isoforms of Lethocerus.

Muscles are usually activated by calcium binding to the calcium sensory protein troponin-C, which is one of the three components of the troponin complex. However, in cardiac and insect flight muscle activation is also produced by mechanical stress. Little is known about the molecular bases of this calcium-independent activation. In Lethocerus, a giant water bug often used as a model system because of its large muscle fibers, there are two troponin-C isoforms, called F1 and F2, that have distinct roles in activating the muscle. It has been suggested that this can be explained either by differences in structural features or by differences in the interactions with other proteins. Here we have compared the structural and dynamic properties of the two proteins and shown how they differ. We have also mapped the interactions of the F2 isoform with peptides spanning the sequence of its natural partner, troponin-I. Our data have allowed us to build a model of the troponin complex and may eventually help in understanding the specialized function of the F1 and F2 isoforms and the molecular mechanism of stretch activation

Geodemographic approaches to identifying U.S. furniture markets

Demand for household furniture is shaped by a multitude of dynamic forces, from the global economic situation to subtle changes in consumer preferences. Correspondingly, the academic disciplines employed to identify, analyze, and predict these forces range from macroeconomics to psychology, and include demography, sociology, geography, and several applied fields. The demographic characteristics of furniture consumers has been a particularly popular area of research, primarily because of the availability of secondary data from various public · agencies. Some of this research is reviewed in a subsequent section of this report

Central Bank Transparency under Model Uncertainty

This paper explores the effects of central bank transparency on the performance of optimal inflation targeting rules. I assume that both the central bank and the private sector face uncertainty about the correct model of the economy and have to learn. A transparent central bank can reduce one source of uncertainty for private agents by communicating its policy rule to the public. The paper shows that central bank transparency plays a crucial role in stabilizing the agents' learning process and expectations. By contrast, lack of transparency can lead to expectations-driven fluctuations that have destabilizing effects on the economy, even when the central bank has adopted optimal policie

Glutaminyl-tRNA Synthetase from Pseudomonas aeruginosa: Characterization, structure, and development as a screening platform

Pseudomonas aeruginosa has a high potential for developing resistance to multiple antibiotics. The gene (glnS) encoding glutaminyl-tRNA synthetase (GlnRS) from P. aeruginosa was cloned and the resulting protein characterized. GlnRS was kinetically evaluated and the KM and kcatobs , governing interactions with tRNA, were 1.0 μM and 0.15 s-1 , respectively. The crystal structure of the α2 form of P. aeruginosa GlnRS was solved to 1.9 Å resolution. The amino acid sequence and structure of P. aeruginosa GlnRS were analyzed and compared to that of GlnRS from Escherichia coli. Amino acids that interact with ATP, glutamine, and tRNA are well conserved and structure overlays indicate that both GlnRS proteins conform to a similar three-dimensional structure. GlnRS was developed into a screening platform using scintillation proximity assay technology and used to screen ~2,000 chemical compounds. Three inhibitory compounds were identified and analyzed for enzymatic inhibition as well as minimum inhibitory concentrations against clinically relevant bacterial strains. Two of the compounds, BM02E04 and BM04H03, were selected for further studies. These compounds displayed broad-spectrum antibacterial activity and exhibited moderate inhibitory activity against mutant efflux deficient strains of P. aeruginosa and E. coli. Growth of wild-type strains was unaffected, indicating that efflux was likely responsible for the lack of sensitivity. The global mode of action was determined using time-kill kinetics. BM04H03 did not inhibit the growth of human cell cultures at any concentration and BM02E04 only inhibit cultures at the highest concentration tested (400 μg/ml). In conclusion, GlnRS from P. aeruginosa is shown to have a structure similar to that of E. coli GlnRS and two natural product compounds were identified as inhibitors of P. aeruginosa GlnRS with the potential for utility as lead candidates in antibacterial drug development in a time of increased antibiotic resistance

Experimental observation and characterization of the magnetorotational instability

Differential rotation occurs in conducting flows in accretion disks and planetary cores. In such systems, the magnetorotational instability can arise from coupling Lorentz and centrifugal forces to cause large radial angular momentum fluxes. We present the first experimental observation of the magnetorotational instability. Our system consists of liquid sodium between differentially rotating spheres, with an imposed coaxial magnetic field. We characterize the observed patterns, dynamics and torque increases, and establish that this instability can occur from a hydrodynamic turbulent background.Comment: 4 pages, 5 figures. Final version, accepted for publication in Physical Review Letter

The Vaccinia Virus (VACV) B1 and Cellular VRK2 Kinases Promote VACV Replication Factory Formation through Phosphorylation-Dependent Inhibition of VACV B12

Comparative examination of viral and host protein homologs reveals novel mechanisms governing downstream signaling effectors of both cellular and viral origin. The vaccinia virus B1 protein kinase is involved in promoting multiple facets of the virus life cycle and is a homolog of three conserved cellular enzymes called vaccinia virus-related kinases (VRKs). Recent evidence indicates that B1 and VRK2 mediate a common pathway that is largely uncharacterized but appears independent of previous VRK substrates. Interestingly, separate studies described a novel role for B1 in inhibiting vaccinia virus protein B12, which otherwise impedes an early event in the viral lifecycle. Herein, we characterize the B1/VRK2 signaling axis to better understand their shared functions. First, we demonstrate that vaccinia virus uniquely requires VRK2 for viral replication in the absence of B1, unlike other DNA viruses. Employing loss-of-function analysis, we demonstrate that vaccinia virus’s dependence on VRK2 is only observed in the presence of B12, suggesting that B1 and VRK2 share a pathway controlling B12. Moreover, we substantiate a B1/VRK2/B12 signaling axis by examining coprecipitation of B12 by B1 and VRK2. Employing execution point analysis, we reveal that virus replication proceeds normally through early protein translation and uncoating but stalls at replication factory formation in the presence of B12 activity. Finally, structure/function analyses of B1 and VRK2 demonstrate that enzymatic activity is essential for B1 or VRK2 to inhibit B12. Together, these data provide novel insights into B1/VRK signaling coregulation and support a model in which these enzymes modulate B12 in a phosphorylation-dependent manner

MGMR: leveraging RNA-Seq population data to optimize expression estimation

<p>Abstract</p> <p>Background</p> <p>RNA-Seq is a technique that uses Next Generation Sequencing to identify transcripts and estimate transcription levels. When applying this technique for quantification, one must contend with reads that align to multiple positions in the genome (multireads). Previous efforts to resolve multireads have shown that RNA-Seq expression estimation can be improved using probabilistic allocation of reads to genes. These methods use a probabilistic generative model for data generation and resolve ambiguity using likelihood-based approaches. In many instances, RNA-seq experiments are performed in the context of a population. The generative models of current methods do not take into account such population information, and it is an open question whether this information can improve quantification of the individual samples</p> <p>Results</p> <p>In order to explore the contribution of population level information in RNA-seq quantification, we apply a hierarchical probabilistic generative model, which assumes that expression levels of different individuals are sampled from a Dirichlet distribution with parameters specific to the population, and reads are sampled from the distribution of expression levels. We introduce an optimization procedure for the estimation of the model parameters, and use HapMap data and simulated data to demonstrate that the model yields a significant improvement in the accuracy of expression levels of paralogous genes.</p> <p>Conclusions</p> <p>We provide a proof of principal of the benefit of drawing on population commonalities to estimate expression. The results of our experiments demonstrate this approach can be beneficial, primarily for estimation at the gene level.</p

Label-free electrochemical monitoring of DNA ligase activity

This study presents a simple, label-free electrochemical technique for the monitoring of DNA ligase activity. DNA ligases are enzymes that catalyze joining of breaks in the backbone of DNA and are of significant scientific interest due to their essential nature in DNA metabolism and their importance to a range of molecular biological methodologies. The electrochemical behavior of DNA at mercury and some amalgam electrodes is strongly influenced by its backbone structure, allowing a perfect discrimination between DNA molecules containing or lacking free ends. This variation in electrochemical behavior has been utilized previously for a sensitive detection of DNA damage involving the sugar-phosphate backbone breakage. Here we show that the same principle can be utilized for monitoring of a reverse process, i.e., the repair of strand breaks by action of the DNA ligases. We demonstrate applications of the electrochemical technique for a distinction between ligatable and unligatable breaks in plasmid DNA using T4 DNA ligase, as well as for studies of the DNA backbone-joining activity in recombinant fragments of E. coli DNA ligase