Structural basis for the redox sensitivity of the Mycobacterium tuberculosis SigK-RskA sigma-anti-sigma complex

The host-pathogen interactions in Mycobacterium tuberculosis infection are significantly influenced by redox stimuli and alterations in the levels of secreted antigens. The extracyto-plasmic function (ECF) sigma factor sigma(K) governs the transcription of the serodominant antigens MPT70 and MPT83. The cellular levels of sigma(K) are regulated by the membrane-associated anti-sigma(K) (RskA) that localizes sigma(K) in an inactive complex. The crystal structure of M. tuberculosis sigma(K) in complex with the cytosolic domain of RskA (RskAcyto) revealed a disulfide bridge in the -35 promoter-interaction region of sigma(K). Biochemical experiments reveal that the redox potential of the disulfide-forming cysteines in sigma(K) is consistent with its role as a sensor. The disulfide bond in sigma(K) influences the stability of the sigma(K)-RskA(cyto) complex but does not interfere with sigma(K)-promoter DNA interactions. It is noted that these disulfide-forming cysteines are conserved across homologues, suggesting that this could be a general mechanism for redox-sensitive transcription regulation.

Molecular Basis for the Differential Quinolone Susceptibility of Mycobacterial DNA Gyrase

DNA gyrase is a type II topoisomerase that catalyzes the introduction of negative supercoils in the genomes of eubacteria. Fluoroquinolones (FQs), successful as drugs clinically, target the enzyme to trap the gyrase-DNA complex, leading to the accumulation of double-strand breaks in the genome. Mycobacteria are less susceptible to commonly used FQs. However, an 8-methoxy-substituted FQ, moxifloxacin (MFX), is a potent antimycobacterial, and a higher susceptibility of mycobacterial gyrase to MFX has been demonstrated. Although several models explain the mechanism of FQ action and gyrase-DNA-FQ interaction, the basis for the differential susceptibility of mycobacterial gyrase to various FQs is not understood. We have addressed the basis of the differential susceptibility of the gyrase and revisited the mode of action of FQs. We demonstrate that FQs bind both Escherichia coli and Mycobacterium tuberculosis gyrases in the absence of DNA and that the addition of DNA enhances the drug binding. The FQs bind primarily to the GyrA subunit of mycobacterial gyrase, while in E. coli holoenzyme is the target. The binding of MFX to GyrA of M. tuberculosis correlates with its effectiveness as a better inhibitor of the enzyme and its efficacy in cell killing.

Structural basis for the catalytic mechanism of homoserine dehydrogenase

Homoserine dehydrogenase (HSD) is an oxidoreductase in the aspartic acid pathway. This enzyme coordinates a critical branch point of the metabolic pathway that leads to the synthesis of bacterial cell-wall components such as L-lysine and m-DAP in addition to other amino acids such as L-threonine, L-methionine and L-isoleucine. Here, a structural rationale for the hydride-transfer step in the reaction mechanism of HSD is reported. The structure of Staphylococcus aureus HSD was determined at different pH conditions to understand the basis for the enhanced enzymatic activity at basic pH. An analysis of the crystal structure revealed that Lys105, which is located at the interface of the catalytic and cofactor-binding sites, could mediate the hydride-transfer step of the reaction mechanism. The role of Lys105 was subsequently confirmed by mutational analysis. Put together, these studies reveal the role of conserved water molecules and a lysine residue in hydride transfer between the substrate and the cofactor.

Prediction of Indian rainfall during the summer monsoon season on the basis of links with equatorial Pacific and Indian Ocean climate indices

Interannual variation of Indian summer monsoon rainfall (ISMR) is linked to El Nino-Southern oscillation (ENSO) as well as the Equatorial Indian Ocean oscillation (EQUINOO) with the link with the seasonal value of the ENSO index being stronger than that with the EQUINOO index. We show that the variation of a composite index determined through bivariate analysis, explains 54% of ISMR variance, suggesting a strong dependence of the skill of monsoon prediction on the skill of prediction of ENSO and EQUINOO. We explored the possibility of prediction of the Indian rainfall during the summer monsoon season on the basis of prior values of the indices. We find that such predictions are possible for July-September rainfall on the basis of June indices and for August-September rainfall based on the July indices. This will be a useful input for second and later stage forecasts made after the commencement of the monsoon season.

An Algorithm to Decide Feasibility of Linear Integer Constraints Occurrng in Decision Tables

Abstract-To detect errors in decision tables one needs to decide whether a given set of constraints is feasible or not. This paper describes an algorithm to do so when the constraints are linear in variables that take only integer values. Decision tables with such constraints occur frequently in business data processing and in nonnumeric applications. The aim of the algorithm is to exploit. the abundance of very simple constraints that occur in typical decision table contexts. Essentially, the algorithm is a backtrack procedure where the the solution space is pruned by using the set of simple constrains. After some simplications, the simple constraints are captured in an acyclic directed graph with weighted edges. Further, only those partial vectors are considered from extension which can be extended to assignments that will at least satisfy the simple constraints. This is how pruning of the solution space is achieved. For every partial assignment considered, the graph representation of the simple constraints provides a lower bound for each variable which is not yet assigned a value. These lower bounds play a vital role in the algorithm and they are obtained in an efficient manner by updating older lower bounds. Our present algorithm also incorporates an idea by which it can be checked whether or not an (m - 2)-ary vector can be extended to a solution vector of m components, thereby backtracking is reduced by one component.

Structural Basis of Drug Resistance by Genetic Variants of HIV Type 1 Clade C Protease from India

Using computer modeling of three-dimensional structures and structural information available on the crystal structures of HIV-1 protease, we investigated the structural effects of mutations, in treatment-naive and treatment-exposed individuals from India and postulated mechanisms of resistance in clade C variants. A large number of models (14) have been generated by computational mutation of the available crystal structures of drug bound proteases. Localized energy minimization was carried out in and around the sites of mutation in order to optimize the geometry of interactions present. Most of the mutations result in structural differences at the flap that favors the semiopen state of the enzyme. Some of the mutations were also found to confer resistance by affecting the geometry of the active site. The E35D mutation affects the flap structure in clade B strains and E35N and E35K mutation, seen in our modeled strains, have a more profound effect. Common polymorphisms at positions 36 and 63 in clade C also affected flap structure. Apart from a few other residues Gln-58, Asn-83, Asn-88, and Gln-92 and their interactions are important for the transition from the closed to the open state. Development of protease inhibitors by structure-based design requires investigation of mechanisms operative for clade C to improve the efficacy of therapy.

The transient response of certain third-ordernon-linear systems

In this paper a method of solving certain third-order non-linear systems by using themethod of ultraspherical polynomial approximation is proposed. By using the method of variation of parameters the third-order equation is reduced to three partial differential equations. Instead of being averaged over a cycle, the non-linear functions are expanded in ultraspherical polynomials and with only the constant term retained, the equations are solved. The results of the procedure are compared with the numerical solutions obtained on a digital computer. A degenerate third-order system is also considered and results obtained for the above system are compared with numerical results obtained on the digital computer. There is good agreement between the results obtained by the proposed method and the numerical solution obtained on digital computer.

Molecular basis of nitrate reductase induction in Candida utilis

Temporal separaton of transcription and translation during nitrate reductase induction oin Candida utilis was achieved by the use of actinomycin D and cycloheximide. The yeast failed to synthesize nitrate reductase when nitrate was not provided during transcription. Nitrate thus appeared to induce during transcription the capacity to synthesize nitrate reductase. Presence of nitrate, on the other hand, was not obligatory during translation except for its essential role in maintaining the stability of nitrate reductase after its formation as well as its mRNA.

Ultraspherical polynomials approach to the study of third-order non-linear systems

In this study, the Krylov-Bogoliubov-Mitropolskii-Popov asymptotic method is used to determine the transient response of third-order non-linear systems. Instead of averaging the non-linear functions over a cycle, they are expanded in ultraspherical polynomials and the constant term is retained. The resulting equations are solved to obtain the approximate solution. A numerical example is considered and the approximate solution is compared with the digital solution. The results show that there is good agreement between the two values.

Equivalent constitutive model-based design of wave-absorbing material system and controller

A design methodology for wave-absorbing active material system is reported. The design enforces equivalence between an assumed material model having wave-absorbing behavior and a set of target feedback controllers for an array of microelectro-mechanical transducers which are integral part of the active material system. The proposed methodology is applicable to problems involving the control of acoustic waves in passive-active material system with complex constitutive behavior at different length-scales. A stress relaxation type one-dimensional constitutive model involving viscous damping mechanism is considered, which shows asymmetric wave dispersion characteristics about the half-line. The acoustic power flow and asymptotic stability of such material system are studied. A single sensor non-collocated linear feedback control system in a one-dimensional finite waveguide, which is a representative volume element in an active material system, is considered. Equivalence between the exact dynamic equilibrium of these two systems is imposed. It results in the solution space of the design variables, namely the equivalent damping coefficient, the wavelength(s) to be controlled and the location of the sensor. The characteristics of the controller transfer functions and their pole-placement problem are studied. (c) 2005 Elsevier Ltd. All rights reserved.

Analysis of Torsional Interactions with Power System Stabilizer

Torsional interactions can occur due to the speed input Power System Stabilizer (PSS) that are primarily used to damp low frequency oscillations. The solution to this problem can be either in the form of providing a torsional filter or developing an alternate signal for the PSS. This paper deals with the formulation of a linearized state space model of the system and study of the interactions using eigenvalue analysis. The effects of the parameters of PSS and control signals on the damping of torsional modes are investigated.

Intra-molecular hydrogen bonding with organic fluorine in the solution state: Deriving evidence by a two dimensional NMR experiment

The direct evidence for the existence of intra-molecular C-F center dot center dot center dot H-N hydrogen bond in organofluorine molecules, in the liquid state, is derived using NMR spectroscopy by the detection of long range interactions among fluorine, nitrogen and hydrogen atoms. The present study reports the determination of the relative signs and magnitudes of through space and through bond couplings to draw unambiguous evidence on the existence of weak molecular interactions involving organic fluorine. It is a simple, easy to implement, N-15 natural abundant two dimensional heteronuclear N-15-H-1 double quantum-single quantum correlation experiment. The existence of intra-molecular hydrogen bond is conclusively established in the investigated molecules. (C) 2011 Elsevier B.V. All rights reserved.

Improvements in Efficiency of Surface Potential Computation for Independent DG MOSFET

A robust numerical solution of the input voltage equations (IVEs) for the independent-double-gate metal-oxide-semiconductor field-effect transistor requires root bracketing methods (RBMs) instead of the commonly used Newton-Raphson (NR) technique due to the presence of nonremovable discontinuity and singularity. In this brief, we do an exhaustive study of the different RBMs available in the literature and propose a single derivative-free RBM that could be applied to both trigonometric and hyperbolic IVEs and offers faster convergence than the earlier proposed hybrid NR-Ridders algorithm. We also propose some adjustments to the solution space for the trigonometric IVE that leads to a further reduction of the computation time. The improvement of computational efficiency is demonstrated to be about 60% for trigonometric IVE and about 15% for hyperbolic IVE, by implementing the proposed algorithm in a commercial circuit simulator through the Verilog-A interface and simulating a variety of circuit blocks such as ring oscillator, ripple adder, and twisted ring counter.

Structural and molecular basis of interaction of HCV non-structural protein 5A with human casein kinase 1 alpha and PKR

Background: Interaction of non-structural protein 5A (NS5A) of Hepatitis C virus (HCV) with human kinases namely, casein kinase 1 alpha (ck1 alpha) and protein kinase R (PKR) have different functional implications such as regulation of viral replication and evasion of interferon induced immune response respectively. Understanding the structural and molecular basis of interactions of the viral protein with two different human kinases can be useful in developing strategies for treatment against HCV. Results: Serine 232 of NS5A is known to be phosphorylated by human ck1 alpha. A structural model of NS5A peptide containing phosphoacceptor residue Serine 232 bound to ck1 alpha has been generated using the known 3-D structures of kinase-peptide complexes. The substrate interacting residues in ck1 alpha has been identified from the model and these are found to be conserved well in the ck1 family. ck1 alpha - substrate peptide complex has also been used to understand the structural basis of association between ck1 alpha and its other viral stress induced substrate, tumour suppressor p53 transactivation domain which has a crystal structure available. Interaction of NS5A with another human kinase PKR is primarily genotype specific. NS5A from genotype 1b has been shown to interact and inhibit PKR whereas NS5A from genotype 2a/3a are unable to bind and inhibit PKR efficiently. This is one of the main reasons for the varied response to interferon therapy in HCV patients across different genotypes. Using PKR crystal structure, sequence alignment and evolutionary trace analysis some of the critical residues responsible for the interaction of NS5A 1b with PKR have been identified. Conclusions: The substrate interacting residues in ck1 alpha have been identified using the structural model of kinase substrate peptide. The PKR interacting NS5A 1b residues have also been predicted using PKR crystal structure, NS5A sequence analysis along with known experimental results. Functional significance and nature of interaction of interferon sensitivity determining region and variable region 3 of NS5A in different genotypes with PKR which was experimentally shown are also supported by the findings of evolutionary trace analysis. Designing inhibitors to prevent this interaction could enable the HCV genotype 1 infected patients respond well to interferon therapy.