Mechanism of RNA Translocation by a Viral Packaging Motor

Molecular motors are proteins that convert chemical energy into mechanical work. The viral packaging ATPase P4 is a hexameric molecular motor that translocates RNA into preformed viral capsids. P4 belongs to the ubiquitous class of hexameric helicases. Although its structure is known, the mechanism of RNA translocation remains elusive. Here we present a detailed kinetic study of nucleotide binding, hydrolysis, and product release by P4. We propose a stochastic-sequential cooperative model to describe the coordination of ATP hydrolysis within the hexamer. In this model the apparent cooperativity is a result of hydrolysis stimulation by ATP and RNA binding to neighboring subunits rather than cooperative nucleotide binding. Simultaneous interaction of neighboring subunits with RNA makes the otherwise random hydrolysis sequential and processive. Further, we use hydrogen/deuterium exchange detected by high resolution mass spectrometry to visualize P4 conformational dynamics during the catalytic cycle. Concerted changes of exchange kinetics reveal a cooperative unit that dynamically links ATP binding sites and the central RNA binding channel. The cooperative unit is compatible with the structure-based model in which translocation is effected by conformational changes of a limited protein region. Deuterium labeling also discloses the transition state associated with RNA loading which proceeds via opening of the hexameric ring. Hydrogen/deuterium exchange is further used to delineate the interactions of the P4 hexamer with the viral procapsid. P4 associates with the procapsid via its C-terminal face. The interactions stabilize subunit interfaces within the hexamer. The conformation of the virus-bound hexamer is more stable than the hexamer in solution, which is prone to spontaneous ring openings. We propose that the stabilization within the viral capsid increases the packaging processivity and confers selectivity during RNA loading. Finally, we use single molecule techniques to characterize P4 translocation along RNA. While the P4 hexamer encloses RNA topologically within the central channel, it diffuses randomly along the RNA. In the presence of ATP, unidirectional net movement is discernible in addition to the stochastic motion. The diffusion is hindered by activation energy barriers that depend on the nucleotide binding state. The results suggest that P4 employs an electrostatic clutch instead of cycling through stable, discrete, RNA binding states during translocation. Conformational changes coupled to ATP hydrolysis modify the electrostatic potential inside the central channel, which in turn biases RNA motion in one direction. Implications of the P4 model for other hexameric molecular motors are discussed.

Kinetic characterization of rat brain type IIA sodium channel alpha-subunit stably expressed in a somatic cell line

1. The rat brain type IIA Na+ channel alpha-subunit was stably expressed in Chinese hamster ovary (CHO) cells. Current through the expressed Na+ channels was studied using the whole-cell configuration of the patch clamp technique. The transient Na+ current was sensitive to TTX and showed a bell-shaped peak current vs. membrane potential relation. 2. Na+ current inactivation was better described by the sum of two exponentials in the potential range -30 to +40 mV, with. a dominating fast component and a small slower component. 3. The steady-state inactivation, h(infinity), was related to potential by a Boltzmann distribution, underlying thr ee states of the inactivation gate. 4. Recovery of the channels from inactivation at different potentials in the range -70 to -120 mV were characterized by al? initial delay which decreased with hyperpolarization. The time course was well fitted by the sum of two exponentials. In this case the slower exponential was the major component, and both time constants decreased with hyperpolarization. 5. For a working description of the Na+ channel inactivation in this preparation, with a minimal deviation from the Hodgkin-Huxley model, a three-state scheme of the form O reversible arrow I-1 reversible arrow I-2 was proposed, replacing the original two-state scheme of the Hodgkin-Huxley model, and the rate constants are reported. 6. The instantaneous current-voltage relationship showed marked deviation from linearity and was satisfactorily fitted by the constant-field equation. 7. The time course of activation was described by an m(x) model. However, the best-fitted value of x varied with the membrane potential and had a mean value of 2. 8. Effective gating charge was determined to be 4.7e from the slope of the activation plot, plotted on a logarithmic scale. 9. The rate constants of activation, alpha(m) and beta(m), were determined. Their functional dependence on the membrane potential was investigated.

Continuous Distribution Kinetics for Photopolymerization of Alkyl Methacrylates

The photopolymerization of methyl,ethyl,butyl, and hexyl methacrylates in solution was studied. The effect of initial initiator and monomer concentrations on the time evolution of polymer concentration (M) over bar (n) and PDI was examined. The reversible chain addition and beta-scission, and primary radical termination steps were included in the mechanism along with the classical steps. The rate equations were derived using continuous distribution kinetics and solved numerically to fit the experimental data. The regressed rate coefficients compared well with the literature data. The model predicted the instantaneous increase in (M) over bar (n) and PDI to steady state values. The rate coefficients exhibited a linear increase with the size of alkyl chain of the alkyl methacrylates.

Identification of alpha- and beta-Hydroxy Acid Containing Cyclodepsipeptides in Natural Peptide Mixtures Using Negative Ion Mass Spectrometry

Natural peptide libraries often contain cyclodepsipeptides containing alpha or beta hydroxy residues. Extracts of fungal hyphae of Isaria yield a microheterogenous cyclodepsipeptide mixture in which two classes of molecules can be identified by mass spectral fragmentation of negative ions. In the case of isaridins, which contain an alpha-hydroxy residue and a beta-amino acid residue, a characteristic product ion corresponding to a neutral loss of 72 Da is obtained. hi addition, neutral loss of water followed by a 72 Da loss is also observed. Two distinct modes of fragmentation rationalize the observed product ion distribution. The neutral loss of 72 Da has also been obtained for a roseotoxin component, which is also an alpha-hydroxy residue containing cyclodepsipeptide. In the case of isariins, which contain a beta-hydroxy acid residue, ring opening and subsequent loss of the terminal residue as an unsaturated ketene fragment, rationalizes the observed product ion formation. Fragmentation of negative ions provide characteristic neutral losses, which are diagnostic of the presence of alpha-hydroxy or beta-hydroxy residues.

Cross-Layer Scheduling with Infrequent Channel and Queue Measurements

The downlink scheduling problem in multi-queue multi-server systems under channel uncertainty is considered. Two policies that make allocations based on predicted channel states are proposed. The first is an extension of the well-known dynamic backpressure policy to the uncertain channel case. The second is a variant that improves delay performance under light loads. The stability region of the system is characterised and the first policy is argued to be throughput optimal. A recently proposed policy of Kar et al [1] has lesser complexity, but is shown to be throughput suboptimal. Further, simulations demonstrate better delay and backlog properties for both our policies at light loads.

Multi-channel curve crossing problems: analytically solvable model

We have proposed a general method for finding the exact analytical solution for the multi-channel curve crossing problem in the presence of delta function couplings. We have analysed the case where aa potential energy curve couples to a continuum (in energy) of the potential energy curves.

Crystallization kinetics of SrBi2B2O7 glasses by non-isothermal methods

Transparent glasses of SrBi2B2O7 (SBBO) were fabricated via the conventional melt-quenching technique. The amorphous and the glassy nature of the as-quenched samples were, respectively, confirmed by X-ray powder diffraction (XRD) and differential scanning calorimetry (DSC). The glass transition (T (g)) and the crystallization parameters [crystallization activation energy (E (cr)) and Avrami exponent (n)] were evaluated under non-isothermal conditions using DSC. There was a close agreement between the activation energies for the crystallization process determined by Augis and Bennet and Kissinger methods. The variation of local activation energy [E (c)(x)] that was determined by Ozawa method, decreased with the fraction of crystallization (x). The Avrami exponent (n(x)) increased with the increase in fraction of crystallization (x) suggesting that there was a change over in the crystallization process from the surface to the bulk.

High-Rate Space-Time Coded Large-MIMO Systems: Low-Complexity Detection and Channel Estimation

In this paper, we present a low-complexity algorithm for detection in high-rate, non-orthogonal space-time block coded (STBC) large-multiple-input multiple-output (MIMO) systems that achieve high spectral efficiencies of the order of tens of bps/Hz. We also present a training-based iterative detection/channel estimation scheme for such large STBC MIMO systems. Our simulation results show that excellent bit error rate and nearness-to-capacity performance are achieved by the proposed multistage likelihood ascent search (M-LAS) detector in conjunction with the proposed iterative detection/channel estimation scheme at low complexities. The fact that we could show such good results for large STBCs like 16 X 16 and 32 X 32 STBCs from Cyclic Division Algebras (CDA) operating at spectral efficiencies in excess of 20 bps/Hz (even after accounting for the overheads meant for pilot based training for channel estimation and turbo coding) establishes the effectiveness of the proposed detector and channel estimator. We decode perfect codes of large dimensions using the proposed detector. With the feasibility of such a low-complexity detection/channel estimation scheme, large-MIMO systems with tens of antennas operating at several tens of bps/Hz spectral efficiencies can become practical, enabling interesting high data rate wireless applications.

A Joint Source-Channel Coding Scheme for Transmission of Correlated Discrete Sources over a Gaussian Multiple Access Channel

We consider the problem of transmission of correlated discrete alphabet sources over a Gaussian Multiple Access Channel (GMAC). A distributed bit-to-Gaussian mapping is proposed which yields jointly Gaussian codewords. This can guarantee lossless transmission or lossy transmission with given distortions, if possible. The technique can be extended to the system with side information at the encoders and decoder.

Genotype-Phenotype Relationships in Long QT Syndrome: Role of Mental Stress, Adrenergic Activity and a Common KCNH2 Polymorphism

Long QT syndrome is a congenital or acquired arrhythmic disorder which manifests as a prolonged QT-interval on the electrocardiogram and as a tendency to develop ventricular arrhythmias which can lead to sudden death. Arrhythmias often occur during intense exercise and/or emotional stress. The two most common subtypes of LQTS are LQT1, caused by mutations in the KCNQ1 gene and LQT2, caused by mutations in the KCNH2 gene. LQT1 and LQT2 patients exhibit arrhythmias in different types of situations: in LQT1 the trigger is usually vigorous exercise whereas in LQT2 arrhythmia results from the patient being startled from rest. It is not clear why trigger factors and clinical outcome differ from each other in the different LQTS subtypes. It is possible that stress hormones such as catecholamines may show different effects depending on the exact nature of the genetic defect, or sensitivity to catecholamines varies from subject to subject. Furthermore, it is possible that subtle genetic variants of putative modifier genes, including those coding for ion channels and hormone receptors, play a role as determinants of individual sensitivity to life-threatening arrhythmias. The present study was designed to identify some of these risk modifiers. It was found that LQT1 and LQT2 patients show an abnormal QT-adaptation to both mental and physical stress. Furthermore, as studied with epinephrine infusion experiments while the heart was paced and action potentials were measured from the right ventricular septum, LQT1 patients showed repolarization abnormalities which were related to their propensity to develop arrhythmia during intense, prolonged sympathetic tone, such as exercise. In LQT2 patients, this repolarization abnormality was noted already at rest corresponding to their arrhythmic episodes as a result of intense, sudden surges in adrenergic tone, such as fright or rage. A common KCNH2 polymorphism was found to affect KCNH2 channel function as demonstrated by in vitro experiments utilizing mammalian cells transfected with the KCNH2 potassium channel as well as QT-dynamics in vivo. Finally, the present study identified a common β-1-adrenergic receptor genotype that is related a shorter QT-interval in LQT1 patients. Also, it was discovered that compound homozygosity for two common β-adrenergic polymorphisms was related to the occurrence of symptoms in the LQT1 type of long QT syndrome. The studies demonstrate important genotype-phenotype differences between different LQTS subtypes and suggest that common modifier gene polymorphisms may affect cardiac repolarization in LQTS. It will be important in the future to prospectively study whether variant gene polymorphisms will assist in clinical risk profiling of LQTS patients.

Anomalous reaction-diffusion as a model of nonexponential DNA escape kinetics

We show that data from recent experiments carried out on the kinetics of DNA escape from alpha-hemolysin nanopores [M. Wiggin, C. Tropini, C. T. Cossa, N. N. Jetha, and A. Marziali, Biophys. J. 95, 5317 (2008)] may be rationalized by a model of chain dynamics based on the anomalous diffusion of a particle moving in a harmonic well in the presence of a delta function sink. The experiments of Wiggin found, among other things, that the occasional occurrence of unusually long escape times in the distribution of chain trapping events led to nonexponential decays in the survival probability, S(t), of the DNA molecules within the nanopore. Wiggin ascribed this nonexponentiality to the existence of a distribution of trapping potentials, which they suggested was theresult of stochastic interactions between the bases of the DNA and the amino acids located on the surface of the nanopore. Based on this idea, they showed that the experimentally determined S(t) could be well fit in both the short and long time regimes by a function of the form (1+t/tau)(-alpha) (the so called Becquerel function). In our model, S(t) is found to be given by a Mittag-Leffler function at short times and by a generalized Mittag-Leffler function at long times. By suitable choice of certain parameter values, these functions are found to fit the experimental S(t) even better than the Becquerel function. Anomalous diffusion of DNA within the trap prior to escape over a barrier of fixed height may therefore provide a second, plausible explanation of the data, and may offer fresh perspectives on similar trapping and escape problems.

Bismuth sulfide: A high-capacity anode for sodium-ion batteries

Exploring high-performance anode materials is currently one of the most urgent issues towards practical sodium-ion batteries (SIBs). In this work, Bi2S3 is demonstrated to be a high-capacity anode for SIBs for the first time. The specific capacity of Bi2S3 nanorods achieves up to 658 and 264 mAh g-1 at a current density of 100 and 2000 mA g-1, respectively. A full cell with Na3V2(PO4)3-based cathode is also assembled as a proof of concept and delivers 340 mAh g-1 at 100 mA g-1. The sodium storage mechanism of Bi2S3 is investigated by ex-situ XRD coupled with high-resolution TEM (HRTEM), and it is found that sodium storage is achieved by a combined conversion-intercalation mechanism.

Carbon-coated hierarchical SnO2 hollow spheres for lithium ion batteries

Hierarchical SnO2 hollow spheres self-assembled from nanosheets were prepared with and without carbon coating. The combination of nanosized architecture, hollow structure, and a conductive carbon layer endows the SnO2-based anode with improved specific capacity and cycling stability, making it more promising for use in lithium ion batteries.

Atomic layer-by-layer Co3O4/graphene composite for high performance lithium-ion batteries

An "atomic layer-by-layer" structure of Co3O4/graphene is developed as an anode material for lithium-ion batteries. Due to the atomic thickness of both the Co3O4 nanosheets and the graphene, the composite exhibits an ultrahigh specific capacity of 1134.4 mAh g-1 and an ultralong life up to 2000 cycles at 2.25 C, far beyond the performances of previously reported Co3O4/C composites.

3D Fe2(MoO4)3 microspheres with nanosheet constituents as high-capacity anode materials for lithium-ion batteries

Three-dimensional (3D) Fe2(MoO4)3 microspheres with ultrathin nanosheet constituents are first synthesized as anode materials for the lithium-ion battery. It is interesting that the single-crystalline nanosheets allow rapid electron/ion transport on the inside, and the high porosity ensures fast diffusion of liquid electrolyte in energy storage applications. The electrochemical properties of Fe2(MoO4)3 as anode demonstrates that 3D Fe2(MoO4)3 microspheres deliver an initial capacity of 1855 mAh/g at a current density of 100 mA/g. Particularly, when the current density is increased to 800 mA/g, the reversible capacity of Fe2(MoO4)3 anode still arrived at 456 mAh/g over 50 cycles. The large and reversible capacities and stable charge–discharge cycling performance indicate that Fe2(MoO4)3 is a promising anode material for lithium battery applications. Graphical abstract The electrochemical properties of Fe2(MoO4)3 as anode demonstrates that 3D Fe2(MoO4)3 microspheres delivered an initial capacity of 1855 mAh/g at a current density of 100 mA/g. When the current density was increased to 800 mA/g, the Fe2(MoO4)3 still behaved high reversible capacity and good cycle performance.