bluuebunny/crossref_metadata_2025 huggingface dataset:
Spacecraft Vibration Suppression Using Variable Structure Output Feedback Control and Smart Materials
Authors: Hu Qinglei , Ma Guangfu
Published: March 2006

A hybrid control scheme for vibration reduction of flexible spacecraft during rotational maneuvers is investigated by using variable structure output feedback control (VSOFC) for attitude control and smart materials for active vibration suppression. The proposed control design process is twofold: design of the attitude controller using VSOFC theory acting on the hub and design of an independent flexible vibration controller acting on the flexible part using piezoceramics as sensors and actuators to actively suppress certain flexible modes. The attitude controller, using only the attitude and angular rate measurement, consists of a linear feedback term and a discontinuous feedback term, which are designed so that the sliding surface exists and is globally reachable. With the presence of this attitude controller, an additional independent flexible control system acting on the flexible parts is designed for further vibration suppression. Using the piezoelectric materials as actuator/sensor, both single-mode vibration suppression and multimode vibration suppression are studied and compared for the different active vibration control algorithms, constant-gain negative velocity feedback (CGNVF) control, positive position feedback (PPF) control, and linear-quadratic Gaussian (LQG) control. Numerical simulations demonstrate that the proposed approach can significantly reduce the vibration of the flexible appendages and further greatly improve the precision during and after the maneuver operations.
Parametric Instability of a Rotating Circular Ring With Moving, Time-Varying Springs
Authors: Canchi Sripathi Vangipuram , Parker Robert G.
Published: March 2006

Parametric instabilities of in-plane bending vibrations of a rotating ring coupled to multiple, discrete, rotating, time-varying stiffness spring-sets of general geometric description are investigated in this work. Instability boundaries are identified analytically using perturbation analysis and given as closed-form expressions in the system parameters. Ring rotation and time-varying stiffness significantly affect instability regions. Different configurations with a rotating and nonrotating ring, and rotating spring-sets are examined. Simple relations governing the occurrence and suppression of instabilities are discussed for special cases with symmetric circumferential spacing of spring-sets. These results are applied to identify possible conditions of ring gear instability in example planetary gears.
Steady-State Dynamic Response of Preisach Hysteretic Systems
Authors: Spanos P. D. , Kontsos A. , Cacciola P.
Published: March 2006

The goal of this paper is to study the steady-state dynamic response of an oscillator involving a hysteretic component and exposed to harmonic excitation. This is accomplished by using the Preisach formalism in the description of the contribution of the hysteretic component. Two cases are considered. In the first one, the hysteretic component is modeled using a series of “Jenkin’s elements,” while in the second one the same component is modeled by a zero-memory plus a purely hysteretic term. The steady-state amplitude of the response is determined analytically by using the equivalent linearization technique which involves input-output relationships for the equivalent linear system, the stiffness and damping coefficients of which are response-amplitude dependent. The derived results are compared with pertinent numerical data obtained by integrating the nonlinear equation of motion of the oscillator. The analytical and the numerical results are found in excellent agreement and supplement the findings of certain previous studies.
Optimal Placement of Piezoelectric Sensors and Actuators for Controlled Flexible Linkage Mechanisms
Authors: Zhang Xianmin , Erdman Arthur G.
Published: March 2006

The optimal placement of sensors and actuators in active vibration control of flexible linkage mechanisms is studied. First, the vibration control model of the flexible mechanism is introduced. Second, based on the concept of the controllability and the observability of the controlled subsystem and the residual subsystem, the optimal model is developed aiming at the maximization of the controllability and the observability of the controlled modes and minimization of those of the residual modes. Finally, a numerical example is presented, which shows that the proposed method is feasible. Simulation analysis shows that to achieve the same control effect, the control system is easier to realize if the sensors and actuators are located in the optimal positions.
Adaptive Control of a Base Isolated System for Protection of Building Structures
Authors: Zhou Jing , Wen Changyun , Cai Wenjian
Published: March 2006

In this paper, we present two adaptive backstepping control algorithms for a second-order uncertain hysteretic structural system found in base isolation scheme for seismic active protection of building structures. The hysteretic nonlinear behavior is described by a Bouc–Wen model. The structural parameters and isolation parameters are all uncertain parameters. In the first scheme, there is no apriori information required from these parameters and the residual effect of the hysteresis is treated as a bounded disturbance. An update law is used to estimate the bound involving this partial hysteresis effect and external disturbance. In the second scheme, we further take the structure of the Bouc–Wen model describing the hysteresis into account in the controller design, if apriori knowledge on some parameters of the model is available. It is shown that not only is global stability guaranteed by the proposed controller, but also both transient and asymptotic performances are quantified as explicit functions of the design parameters so that designers can tune the design parameters in an explicit way to obtain the required closed loop behavior.
Multibody Approach for Model-Based Fault Detection of a Reel
Authors: Korkealaakso Pasi , Rouvinen Asko , Mikkola Aki
Published: March 2006

In order to improve the recognition of faulty situations, model-based fault detection can be used together with signal processing methods. In this study, faults and abnormalities of a reel are studied by employing the multibody simulation approach. The reel under consideration consists of a number of subsystems, including hydraulics, electrical drives, and mechanical parts. These subsystems are coupled by joints, friction forces, and contact forces. Using the multibody simulation approach, the complete model of the reel can be obtained by coupling different subsystems together. Three well-known multibody formulations, a method of Lagrange multipliers, an Augmented Lagrangian method, and a method based on projection matrix R, are briefly described and compared in order to find out the most efficient method for simulating the studied reel. Although this study is focused on the simulation of fault scenarios, the introduced multibody simulation approach can be utilized in real-time simulation. This makes it possible to apply the model to an existing reel.
Nonlinear Modeling of Flexible Manipulators Using Nondimensional Variables
Authors: Shaker M. Chandra , Ghosal Ashitava
Published: March 2006

This paper deals with nonlinear modeling of planar one- and two-link, flexible manipulators with rotary joints using finite element method (FEM) based approaches. The equations of motion are derived taking into account the nonlinear strain-displacement relationship and two characteristic velocities, Ua and Ug, representing material and geometric properties (also axial and flexural stiffness) respectively, are used to nondimensionalize the equations of motion. The effect of variation of Ua and Ug on the dynamics of a planar flexible manipulator is brought out using numerical simulations. It is shown that above a certain Ug value (approximately ⩾45m∕s), a linear model (using a linear strain-displacement relationship) and the nonlinear model give approximately the same tip deflection. Likewise, it was found that the effect of Ua is prominent only if Ug is small. The natural frequencies are seen to be varying in a nonlinear manner with Ua and in a linear manner with Ug.
Chaos and Quasi-Periodic Motions on the Homoclinic Surface of Nonlinear Hamiltonian Systems With Two Degrees of Freedom
Authors: Luo Albert C. J.
Published: March 2006

The numerical prediction of chaos and quasi-periodic motion on the homoclinic surface of a two-degree-of-freedom (2-DOF) nonlinear Hamiltonian system is presented through the energy spectrum method. For weak interactions, the analytical conditions for chaotic motion in such a Hamiltonian system are presented through the incremental energy approach. The Poincaré mapping surfaces of chaotic motions for this specific nonlinear Hamiltonian system are illustrated. The chaotic and quasi-periodic motions on the phase planes, displacement subspace (or potential domains), and the velocity subspace (or kinetic energy domains) are illustrated for a better understanding of motion behaviors on the homoclinic surface. Through this investigation, it is observed that the chaotic and quasi-periodic motions almost fill on the homoclinic surface of the 2-DOF nonlinear Hamiltonian system. The resonant-periodic motions for such a system are theoretically countable but numerically inaccessible. Such conclusions are similar to the ones in the KAM theorem even though the KAM theorem is based on the small perturbation.
New Model of a CVT Rocker-Pin Chain With Exact Joint Kinematics
Authors: Neumann Lutz , Ulbrich Heinz , Pfeiffer Friedrich
Published: March 2006

The improvement of continuously variable transmissions (CVTs) is a challenging task. Detailed dynamic models of the system are needed for the optimization process. This paper introduces a new model of a rocker-pin chain, which is the central part of a chain CVT. During the derivation of the equations of motion, special attention is turned on the exact description of the joint kinematics. An interesting method to speed up the numerical simulation of the chain drive is introduced, which takes into account the special structure of the mass matrix. Simulation results show the strong influence of the geometry of the rocker-pin joints on the dynamics of the whole gear. Further reasonable gear efficiencies can be estimated for arbitrary joint geometries. This model is an excellent basis for further work on optimization of the CVT chain.
A Compliant Contact Model Including Interference Geometry for Polyhedral Objects
Authors: Luo Lianzhen , Nahon Meyer
Published: March 2006

Modeling of contact with the environment is an essential capability for the simulation of space robotics system, which includes tasks such as berthing and docking. The effect of interbody contact on the robotic system has to be determined to predict potential problems in the design cycle. A compliant contact dynamics model is proposed here that considers most possible contact situations for a wide diversity of possible object shapes and using interference geometry information. A uniform formula is provided to determine the contact force as a function of geometric parameters and material properties. A corresponding geometric algorithm is provided in order to obtain the necessary geometric parameters. Some simulation results are presented based on the implementation of the geometric algorithm.
Normal Force-Displacement Relationship of Spherical Joints With Clearances
Authors: Liu Cai-shan , Zhang Ke , Yang Lei
Published: March 2006

The spherical joint with clearances can be modeled as an axi-symmetric quasi-static contact of a sphere in a cavity. The Hertz theory based on the assumption of nonconformal contact is often used to represent the normal force-displacement relationship for the spherical joint with clearances. This assumption limits the application of the theory, especially in the case of occurring large deformation in contact area. The Steuermann theory is effective to solve the conformal contact problem in some cases, but it is strictly dependent on the selecting index n of the polynomial, which is used to represent the contact profiles. In this paper, an approximate contact model of the spherical joints with clearances is developed that is based on using the distributed elastic forces to model the compliant of the surfaces in contact. The new formulation is simple and straightforward, and its validity has been tested by comparison to the finite element results.
The Nonlinear Behaviors of a Symmetric Isotropic Laminate in a Mixed Boundary Condition Subject to an Arbitrary Thermal Field Coupled with Mechanical Loading
Authors: He Xiaoling
Published: March 2006

This paper investigates the nonlinear dynamics of a symmetric isotropic rectangular laminated structure by using the method of weighted residuals with total energy conservation. We obtain the equation of motion in a decoupled form Duffing equation for the laminate deflection subject to both thermal and mechanical loading while confined in a mixed boundary condition of simply supported and clamped edges at the boundaries. The Duffing equation incorporates both the steady-state and transient state for the in-plane and transverse thermal fields, which allows for investigation of the thermal field influence on the laminate buckling and vibration behaviors.
Numerical Scheme for the Solution of Fractional Differential Equations of Order Greater Than One
Authors: Kumar Pankaj , Agrawal Om P.
Published: March 2006

This paper presents a numerical scheme for the solutions of Fractional Differential Equations (FDEs) of order α, 1<α<2 which have been expressed in terms of Caputo Fractional Derivative (FD). In this scheme, the properties of the Caputo derivative are used to reduce an FDE into a Volterra-type integral equation. The entire domain is divided into several small domains, and the distribution of the unknown function over the domain is expressed in terms of the function values and its slopes at the node points. These approximations are then substituted into the Volterra-type integral equation to reduce it to algebraic equations. Since the method enforces the continuity of variables at the node points, it provides a solution that is continuous and with a slope that is also continuous over the entire domain. The method is used to solve two problems, linear and nonlinear, using two different types of polynomials, cubic order and fractional order. Results obtained using both types of polynomials agree well with the analytical results for problem 1 and the numerical results obtained using another scheme for problem 2. However, the fractional order polynomials give more accurate results than the cubic order polynomials do. This suggests that for the numerical solutions of FDEs fractional order polynomials may be more suitable than the integer order polynomials. A series of numerical studies suggests that the algorithm is stable.
Suppression of Planet Mode Response in Planetary Gear Dynamics Through Mesh Phasing
Authors: Ambarisha Vijaya Kumar , Parker Robert G.
Published: March 2006

This work analytically derives design rules to suppress certain harmonics of planet mode response in planetary gear dynamics through mesh phasing. Planet modes are one of three categories of planetary gear vibration modes. In these modes, only the plantes deflect while the carrier, ring, and sun gears have no motion (Lin, J., and Parker, R. G., 1999, ASME J. Vib. Acoust., 121, pp. 316–321;J. Sound Vib, 233(5), pp. 921–928). The dynamic mesh forces are not explicitly modeled for this study; instead, the symmetry of planetary gear systems and gear tooth mesh periodicity are sufficient to establish rules to suppress planet modes. Thus, the conclusions are independent of the mesh modeling details. Planetary gear systems with equally spaced planets and with diametrically opposed planet pairs are examined. Suppression of degenerate mode response in purely rotational degree-of-freedom models achieved in the limit of infinite bearing stiffness is also investigated. The mesh phasing conclusions are verified by dynamic simulations of various planetary gears using a lumped-parameter analytical model and by comparisons to others’ research.
Differential Transformation Approach for Free Vibration Analysis of a Centrifugally Stiffened Timoshenko Beam
Authors: Mei C.
Published: March 2006

In this paper, the differential transformation approach is applied to analyze the free vibration of centrifugally stiffened Timoshenko beam structures. Such structures involve variable coefficients in the governing equations, which in general cannot be solved analytically in closed form. Both the natural frequencies and the mode shapes are obtained using the differential transformation technique. Numerical examples are presented and results are compared with available results in the literature.
A Mitotic Lamin B Matrix Induced by RanGTP Required for Spindle Assembly
Authors: Tsai Ming-Ying , Wang Shusheng , Heidinger Jill M. , Shumaker Dale K. , Adam Stephen A. , Goldman Robert D. , Zheng Yixian
Published: March 2006

Mitotic spindle morphogenesis is a series of highly coordinated movements that lead to chromosome segregation and cytokinesis. We report that the intermediate filament protein lamin B, a component of the interphase nuclear lamina, functions in spindle assembly. Lamin B assembled into a matrix-like network in mitosis through a process that depended on the presence of the guanosine triphosphate–bound form of the small guanosine triphosphatase Ran. Depletion of lamin B resulted in defects in spindle assembly. Dominant negative mutant lamin B proteins that disrupt lamin B assembly in interphase nuclei also disrupted spindle assembly in mitosis. Furthermore, lamin B was essential for the formation of the mitotic matrix that tethers a number of spindle assembly factors. We propose that lamin B is a structural component of the long-sought-after spindle matrix that promotes microtubule assembly and organization in mitosis.
Deconvolution of the Factors Contributing to the Increase in Global Hurricane Intensity
Authors: Hoyos C. D. , Agudelo P. A. , Webster P. J. , Curry J. A.
Published: March 2006

To better understand the change in global hurricane intensity since 1970, we examined the joint distribution of hurricane intensity with variables identified in the literature as contributing to the intensification of hurricanes. We used a methodology based on information theory, isolating the trend from the shorter-term natural modes of variability. The results show that the trend of increasing numbers of category 4 and 5 hurricanes for the period 1970–2004 is directly linked to the trend in sea-surface temperature; other aspects of the tropical environment, although they influence shorter-term variations in hurricane intensity, do not contribute substantially to the observed global trend.
SV2 Is the Protein Receptor for Botulinum Neurotoxin A
Authors: Dong Min , Yeh Felix , Tepp William H. , Dean Camin , Johnson Eric A. , Janz Roger , Chapman Edwin R.
Published: March 2006

How the widely used botulinum neurotoxin A (BoNT/A) recognizes and enters neurons is poorly understood. We found that BoNT/A enters neurons by binding to the synaptic vesicle protein SV2 (isoforms A, B, and C). Fragments of SV2 that harbor the toxin interaction domain inhibited BoNT/A from binding to neurons. BoNT/A binding to SV2A and SV2B knockout hippocampal neurons was abolished and was restored by expressing SV2A, SV2B, or SV2C. Reduction of SV2 expression in PC12 and Neuro-2a cells also inhibited entry of BoNT/A, which could be restored by expressing SV2 isoforms. Finally, mice that lacked an SV2 isoform (SV2B) displayed reduced sensitivity to BoNT/A. Thus, SV2 acts as the protein receptor for BoNT/A.
Structure and Receptor Specificity of the Hemagglutinin from an H5N1 Influenza Virus
Authors: Stevens James , Blixt Ola , Tumpey Terrence M. , Taubenberger Jeffery K. , Paulson James C. , Wilson Ian A.
Published: March 2006

The hemagglutinin (HA) structure at 2.9 angstrom resolution, from a highly pathogenic Vietnamese H5N1 influenza virus, is more related to the 1918 and other human H1 HAs than to a 1997 duck H5 HA. Glycan microarray analysis of this Viet04 HA reveals an avian α2-3 sialic acid receptor binding preference. Introduction of mutations that can convert H1 serotype HAs to human α2-6 receptor specificity only enhanced or reduced affinity for avian-type receptors. However, mutations that can convert avian H2 and H3 HAs to human receptor specificity, when inserted onto the Viet04 H5 HA framework, permitted binding to a natural human α2-6 glycan, which suggests a path for this H5N1 virus to gain a foothold in the human population.
Porin-Mediated Antibiotic Resistance in
Neisseria gonorrhoeae
: Ion, Solute, and Antibiotic Permeation through PIB Proteins with
penB
Mutations
Authors: Olesky Melanie , Zhao Shuqing , Rosenberg Robert L. , Nicholas Robert A.
Published: March 2006

Neisseria gonorrhoeae
has two porins, PIA and PIB, whose genes (
porA
and
porB
, respectively) are alleles of a single
por
locus. We recently demonstrated that
penB
mutations at positions 120 and 121 in PIB, which are presumed to reside in loop 3 that forms the pore constriction zone, confer intermediate-level resistance to penicillin and tetracycline (M. Olesky, M. Hobbs, and R. A. Nicholas, Antimicrob. Agents Chemother.
46:
2811-2820, 2002). In the present study, we investigated the electrophysiological properties as well as solute and antibiotic permeation rates of recombinant PIB proteins containing
penB
mutations (G120K, G120D/A121D, G120P/A121P, and G120R/A121H). In planar lipid bilayers, the predominant conducting state of each porin variant was 30 to 40% of the wild type, even though the anion selectivity and maximum channel conductance of each PIB variant was similar to that of the wild type. Liposome-swelling experiments revealed no significant differences in the permeation of sugars or β-lactam antibiotics through the wild type or PIB variants. Although these results are seemingly contradictory with the ability of these variants to increase antibiotic resistance, they are consistent with MIC data showing that these porin mutations confer resistance only in strains containing an
mtrR
mutation, which increases expression of the MtrC-MtrD-MtrE efflux pump. Moreover, both the
mtrR
and
penB
mutations were required to decrease in vivo permeation rates below those observed in the parental strain containing either
mtrR
or porin mutations alone. Thus, these data demonstrate a novel mechanism of porin-mediated resistance in which mutations in PIB have no affect on antibiotic permeation alone but instead act synergistically with the MtrC-MtrD-MtrE efflux pump in the development of antibiotic resistance in gonococci.
Phenotypic Characterization ofStreptococcus pneumoniaeBiofilm Development
Authors: Allegrucci Magee , Hu F. Z. , Shen K. , Hayes J. , Ehrlich Garth D. , Post J. Christopher , Sauer Karin
Published: March 2006

Streptococcus pneumoniaeis among the most common pathogens associated with chronic otitis media with effusion, which has been hypothesized to be a biofilm disease.S. pneumoniaehas been shown to form biofilms, however, little is known about the developmental process, the architecture, and the changes that occur upon biofilm development. In the current study we made use of a continuous-culture biofilm system to characterize biofilm development of 14 differentS. pneumoniaestrains representing at least 10 unique serotypes. The biofilm development process was found to occur in three distinct stages, including initial attachment, cluster formation, and biofilm maturation. While all 14 pneumococcal strains displayed similar developmental stages, the mature biofilm architecture differed significantly among the serotypes tested. Overall, three biofilm architectural groups were detected based on biomass, biofilm thickness, and cluster size. The biofilm viable cell counts and total protein concentration increased steadily over the course of biofilm development, reaching ∼8 × 108cells and ∼15 mg of protein per biofilm after 9 days of biofilm growth. Proteomic analysis confirmed the presence of distinct biofilm developmental stages by the detection of multiple phenotypes over the course of biofilm development. The biofilm development process was found to correlate not only with differential production of proteins but also with a dramatic increase in the number of detectable proteins, indicating that biofilm formation byS. pneumoniaemay be a far more complex process than previously anticipated. Protein identification revealed that proteins involved in virulence, adhesion, and resistance were more abundant under biofilm growth conditions. A possible role of the identified proteins in biofilm formation is discussed.
Single-Strand-Specific Exonucleases Prevent Frameshift Mutagenesis by Suppressing SOS Induction and the Action of DinB/DNA Polymerase IV in Growing Cells
Authors: Hersh Megan N. , Morales Liza D. , Ross Kimberly J. , Rosenberg Susan M.
Published: March 2006

Escherichia coli
strains carrying null alleles of genes encoding single-strand-specific exonucleases ExoI and ExoVII display elevated frameshift mutation rates but not base substitution mutation rates. We characterized increased spontaneous frameshift mutation in ExoI
−
ExoVII
−
cells and report that some of this effect requires RecA, an inducible SOS DNA damage response, and the low-fidelity, SOS-induced DNA polymerase DinB/PolIV, which makes frameshift mutations preferentially. We also find that SOS is induced in ExoI
−
ExoVII
−
cells. The data imply a role for the single-stranded exonucleases in guarding the genome against mutagenesis by removing excess single-stranded DNA that, if left, leads to SOS induction and PolIV-dependent mutagenesis. Previous results implicated PolIV in
E. coli
mutagenesis specifically during starvation or antibiotic stresses. Our data imply that PolIV can also promote mutation in growing cells under genome stress due to excess single-stranded DNA.
Oxygen-Dependent Regulation of the Central Pathway for the Anaerobic Catabolism of Aromatic Compounds inAzoarcussp. Strain CIB
Authors: Durante-Rodríguez Gonzalo , Zamarro María Teresa , García José Luis , Díaz Eduardo , Carmona Manuel
Published: March 2006

The role of oxygen in the transcriptional regulation of thePNpromoter that controls thebzdoperon involved in the anaerobic catabolism of benzoate in the denitrifyingAzoarcussp. strain CIB has been investigated. In vivo experiments usingPN::lacZtranslational fusions, in bothAzoarcussp. strain CIB andEscherichia colicells, have shown an oxygen-dependent repression effect on the transcription of thebzdcatabolic genes.E. coliFnr was required for the anaerobic induction of thePNpromoter, and the oxygen-dependent repression of thebzdgenes could be bypassed by the expression of a constitutively active Fnr* protein. In vitro experiments revealed that Fnr binds to thePNpromoter at a consensus sequence centered at position −41.5 from the transcription start site overlapping the −35 box, suggesting thatPNbelongs to the class II Fnr-dependent promoters. Fnr interacts with RNA polymerase (RNAP) and is strictly required for transcription initiation after formation of the RNAP-PNcomplex. Anfnrortholog, theacpRgene, was identified in the genome ofAzoarcussp. strain CIB. TheAzoarcussp. strain CIBacpRmutant was unable to grow anaerobically on aromatic compounds and it did not drive the expression of thePN::lacZfusion, suggesting that AcpR is the cognate transcriptional activator of thePNpromoter. Since the lack of AcpR inAzoarcussp. strain CIB did not affect growth on nonaromatic carbon sources, AcpR can be considered a transcriptional regulator of the Fnr/Crp superfamily that has evolved to specifically control the central pathway for the anaerobic catabolism of aromatic compounds inAzoarcus.
Polyamines Are Essential for the Formation of Plague Biofilm
Authors: Patel Chandra N. , Wortham Brian W. , Lines J. Louise , Fetherston Jacqueline D. , Perry Robert D. , Oliveira Marcos A.
Published: March 2006

We provide the first evidence for a link between polyamines and biofilm levels inYersinia pestis, the causative agent of plague. Polyamine-deficient mutants ofY. pestiswere generated with a single deletion inspeAorspeCand a double deletion mutant. The genesspeAandspeCcode for the biosynthetic enzymes arginine decarboxylase and ornithine decarboxylase, respectively. The level of the polyamine putrescine compared to the parentalspeA+speC+strain (KIM6+) was depleted progressively, with the highest levels found in theY. pestisΔspeCmutant (55% reduction), followed by the ΔspeAmutant (95% reduction) and the ΔspeAΔspeCmutant (>99% reduction). Spermidine, on the other hand, remained constant in the single mutants but was undetected in the double mutant. The growth rates of mutants with single deletions were not altered, while the ΔspeAΔspeCmutant grew at 65% of the exponential growth rate of thespeA+speC+strain. Biofilm levels were assayed by three independent measures: Congo red binding, crystal violet staining, and confocal laser scanning microscopy. The level of biofilm correlated to the level of putrescine as measured by high-performance liquid chromatography-mass spectrometry and as observed in a chemical complementation curve. Complementation of the ΔspeAΔspeCmutant withspeAshowed nearly full recovery of biofilm to levels observed in thespeA+speC+strain. Chemical complementation of the double mutant and recovery of the biofilm defect were only observed with the polyamine putrescine.
Chromosome Evolution in theThermotogales: Large-Scale Inversions and Strain Diversification of CRISPR Sequences
Authors: DeBoy Robert T. , Mongodin Emmanuel F. , Emerson Joanne B. , Nelson Karen E.
Published: March 2006

In the present study, the chromosomes of two members of theThermotogaleswere compared. A whole-genome alignment ofThermotoga maritimaMSB8 andThermotoga neapolitanaNS-E has revealed numerous large-scale DNA rearrangements, most of which are associated with CRISPR DNA repeats and/or tRNA genes. These DNA rearrangements do not include the putative origin of DNA replication but move within the same replichore, i.e., the same replicating half of the chromosome (delimited by the replication origin and terminus). Based on cumulative GC skew analysis, both theT. maritimaandT. neapolitanalineages contain one or two major inverted DNA segments. Also, based on PCR amplification and sequence analysis of the DNA joints that are associated with the major rearrangements, the overall chromosome architecture was found to be conserved at most DNA joints for other strains ofT. neapolitana. Taken together, the results from this analysis suggest that the observed chromosomal rearrangements in theThermotogaleslikely occurred by successive inversions after their divergence from a common ancestor and before strain diversification. Finally, sequence analysis shows that size polymorphisms in the DNA joints associated with CRISPRs can be explained by expansion and possibly contraction of the DNA repeat and spacer unit, providing a tool for discerning the relatedness of strains from different geographic locations.
Significant Gene Order and Expression Differences inBordetella pertussisDespite Limited Gene Content Variation
Authors: Brinig Mary M. , Cummings Craig A. , Sanden Gary N. , Stefanelli Paola , Lawrence Andrew , Relman David A.
Published: March 2006

Bordetella pertussis, an obligate human pathogen and the agent of whooping cough, is a clonal species, despite the dynamic selection pressures imposed by host immunity and vaccine usage. Because the generation of variation is critical for species evolution, we employed a variety of approaches to examine features ofB. pertussisgenetic variation. We found a high level of conservation of gene content among 137B. pertussisstrains with different geographical, temporal, and epidemiological associations, using comparative genomic hybridization. The limited number of regions of difference were frequently located adjacent to copies of the insertion element IS481, which is present in high numbers in theB. pertussischromosome. This repeated sequence appears to provide targets for homologous recombination, resulting in deletion of intervening sequences. Using subtractive hybridization, we searched for previously undetected genes in diverse clinical isolates but did not detect any new genes, indicating that gene acquisition is rare inB. pertussis. In contrast, we found evidence of altered gene order in the several strains that were examined and again found an association of IS481with sites of rearrangement. Finally, we compared whole-genome expression profiles of different strains and found significant changes in transcript abundance, even in the same strain after as few as 12 laboratory passages. This combination of approaches provides a detailed picture of a pathogenic species with little gene loss or gain but with the capacity to generate variation by rearranging its chromosome and altering gene expression. These findings have broad implications for host adaptation by microbial pathogens.
Nucleotide-Dependent Dimerization of the C-Terminal Domain of the ABC Transporter CvaB in Colicin V Secretion
Authors: Guo Xiangxue , Harrison Robert W. , Tai Phang C.
Published: March 2006

The cytoplasmic membrane proteins CvaB and CvaA and the outer membrane protein TolC constitute the bacteriocin colicin V secretion system in
Escherichia coli
. CvaB functions as an ATP-binding cassette transporter, and its C-terminal domain (CTD) contains typical motifs for the nucleotide-binding and Walker A and B sites and the ABC signature motif. To study the role of the CvaB CTD in the secretion of colicin V, a truncated construct of this domain was made and overexpressed. Different forms of the CvaB CTD were found during purification and identified as monomer, dimer, and oligomer forms by gel filtration and protein cross-linking. Nucleotide binding was shown to be critical for CvaB CTD dimerization. Oligomers could be converted to dimers by nucleotide triphosphate-Mg, and nucleotide release from dimers resulted in transient formation of monomers, followed by oligomerization and aggregation. Site-directed mutagenesis showed that the ABC signature motif was involved in the nucleotide-dependent dimerization. The spatial proximity of the Walker A site and the signature motif was shown by disulfide cross-linking a mixture of the A530C and L630C mutant proteins, while the A530C or L630C mutant protein did not dimerize on its own. Taken together, these results indicate that the CvaB CTD formed a nucleotide-dependent head-to-tail dimer.
Identification of a Novel α-Galactosidase from the Hyperthermophilic ArchaeonSulfolobus solfataricus
Authors: Brouns Stan J. J. , Smits Nicole , Wu Hao , Snijders Ambrosius P. L. , Wright Phillip C. , de Vos Willem M. , van der Oost John
Published: March 2006

Sulfolobus solfataricusis an aerobic crenarchaeon that thrives in acidic volcanic pools. In this study, we have purified and characterized a thermostable α-galactosidase from cell extracts ofS. solfataricusP2 grown on the trisaccharide raffinose. The enzyme, designated GalS, is highly specific for α-linked galactosides, which are optimally hydrolyzed at pH 5 and 90°C. The protein consists of 74.7-kDa subunits and has been identified as the gene product of open reading frame Sso3127. Its primary sequence is most related to plant enzymes of glycoside hydrolase family 36, which are involved in the synthesis and degradation of raffinose and stachyose. Both thegalSgene fromS. solfataricusP2 and an orthologous gene fromSulfolobus tokodaiihave been cloned and functionally expressed inEscherichia coli, and their activity was confirmed. At present, theseSulfolobusenzymes not only constitute a distinct type of thermostable α-galactosidases within glycoside hydrolase clan D but also represent the first members from theArchaea.
Receptor-Binding Protein of
Lactococcus lactis
Phages: Identification and Characterization of the Saccharide Receptor-Binding Site
Authors: Tremblay Denise M. , Tegoni Mariella , Spinelli Silvia , Campanacci Valérie , Blangy Stéphanie , Huyghe Céline , Desmyter Aline , Labrie Steve , Moineau Sylvain , Cambillau Christian
Published: March 2006

Phage p2, a member of the lactococcal 936 phage species, infects
Lactococcus lactis
strains by binding initially to specific carbohydrate receptors using its receptor-binding protein (RBP). The structures of p2 RBP, a homotrimeric protein composed of three domains, and of its complex with a neutralizing llama VH domain (VHH5) have been determined (S. Spinelli, A. Desmyter, C. T. Verrips, H. J. de Haard, S. Moineau, and C. Cambillau, Nat. Struct. Mol. Biol. 13:85-89, 2006). Here, we show that VHH5 was able to neutralize 12 of 50 lactococcal phages belonging to the 936 species. Moreover, escape phage mutants no longer neutralized by VHH5 were isolated from 11 of these phages. All of the mutations (but one) cluster in the RBP/VHH5 interaction surface that delineates the receptor-binding area. A glycerol molecule, observed in the 1.7-Å resolution structure of RBP, was found to bind tightly (

K
d

= 0.26 μM) in a crevice located in this area. Other saccharides bind RBP with comparable high affinity. These data prove the saccharidic nature of the bacterial receptor recognized by phage p2 and identify the position of its binding site in the RBP head domain.
The First 45 Amino Acids of SopA Are Necessary for InvB Binding and SPI-1 Secretion
Authors: Higashide Wendy , Zhou Daoguo
Published: March 2006

Salmonella enterica
serovar Typhimurium encodes two type III secretion systems (TTSSs) within pathogenicity island 1 (SPI-1) and island 2 (SPI-2). These type III protein secretion and translocation systems transport a panel of bacterial effector proteins across both the bacterial and the host cell membranes to promote bacterial entry and subsequent survival inside host cells. Effector proteins contain secretion and translocation signals that are often located at their N termini. We have developed a ruffling-based translocation reporter system that uses the secretion- and translocation-deficient catalytic domain of SopE, SopE
78-240
, as a reporter. Using this assay, we determined that the N-terminal 45 amino acid residues of
Salmonella
SopA are necessary and sufficient for directing its secretion and translocation through the SPI-1 TTSS. SopA
1-45
, but not SopA
1-44
, is also able to bind to its chaperone, InvB, indicating that SPI-1 type III secretion and translocation of SopA require its chaperone.
Influence of Culture Heterogeneity in Cell Surface Charge on Adhesion and Biofilm Formation by
Enterococcus faecalis
Authors: van Merode Annet E. J. , van der Mei Henny C. , Busscher Henk J. , Krom Bastiaan P.
Published: March 2006

Biofilm formation is an increasing problem in medicine, due to the intrinsic resistance of microorganisms in the biofilm mode of growth against the host immune system and antimicrobial therapy. Adhesion is an important step in biofilm formation, influenced, among other factors, by the surface hydrophobicities and charges of both the substratum and the adhering microorganisms.
Enterococcus faecalis
strains generally display subpopulations with different surface charges, expressed as bimodal zeta potential distributions. Two-thirds of
E. faecalis
strains isolated from clogged biliary stents displayed such heterogeneity of surface charges in culture. In this study, the influence of this culture heterogeneity on initial adhesion and subsequent biofilm formation was investigated. Heterogeneous strains were retained in higher numbers on polystyrene than homogeneous strains. Also, biofilm formation was much more pronounced for heterogeneous strains than for homogeneous strains. In a population enriched to display only one subpopulation, fewer bacteria were retained than in its original heterogeneous culture. Also, the enriched subpopulation formed less biofilm than its original heterogeneous culture. The presence of ox bile during adhesion resulted in fewer retained bacteria, although heterogeneous strains were still retained in significantly higher numbers than were homogeneous strains, and, in general, the presence of ox bile reduced biofilm formation. The initial adhesion and biofilm formation were independent of the presence of the gene encoding the enterococcal surface protein (
esp
) or the expression of gelatinase (GelE). It is concluded that heterogeneity in cell surface charge represents an advantage for bacteria in the colonization of surfaces.
Biosynthesis of the N-Linked Glycan inCampylobacter jejuniand Addition onto Protein through Block Transfer
Authors: Kelly John , Jarrell Harold , Millar Lorna , Tessier Luc , Fiori Laura M. , Lau Peter C. , Allan Brenda , Szymanski Christine M.
Published: March 2006

In eukaryotes, N-linked protein glycosylation is a universal modification involving addition of preformed oligosaccharides to select Asn-Xaa-Ser/Thr motifs and influencing multiple biological events. We recently demonstrated thatCampylobacter jejuniis the first member of theBacteriato possess an N-linked glycan pathway. In this study, high-resolution magic angle spinning nuclear magnetic resonance (HR-MAS NMR) was applied to probe and quantitateC. jejuni N-glycan biosynthesis in vivo. To confirm HR-MAS NMR findings, glycosylation mutants were screened for chicken colonization potential, and glycoproteins were examined by mass spectrometry and lectin blotting. Consistent with the mechanism in eukaryotes, the combined data indicate that bacterial glycans are assembled en bloc, emphasizing the evolutionary conservation of protein N glycosylation. We also show that under the conditions examined, PglG plays no role in glycan biosynthesis, PglI is the glucosyltransferase and the putative ABC transporter, and WlaB (renamed PglK) is required for glycan assembly. These studies underpin the mechanism of N-linked protein glycosylation inBacteriaand provide a simple model system for investigating protein glycosylation and for exploitation in glycoengineering.
Dual Control of Quorum Sensing by Two TraM-Type Antiactivators in
Agrobacterium tumefaciens
Octopine Strain A6
Authors: Wang Chao , Zhang Hai-Bao , Chen Guozhou , Chen Lingling , Zhang Lian-Hui
Published: March 2006

Agrobacterium tumefaciens
wild-type strains have a unique quorum-sensing (QS)-dependent Ti plasmid conjugative transfer phenotype in which QS signaling is activated by corresponding conjugative opine inducers. Strain K588, with a nopaline-type chromosomal background harboring an octopine-type Ti plasmid, however, is a spontaneous mutant displaying a constitutive phenotype in QS. In this study, we show that a single amino acid mutation (L54P) in the QS antiactivator TraM encoded by the
traM
gene of Ti plasmid is responsible for the constitutive phenotype of strain K588. Introduction of the L54P point mutation to the TraM of wild-type strain A6 by allelic replacement, however, failed to generate the expected constitutive phenotype in this octopine-type strain. Intriguingly, the QS-constitutive phenotype appeared when the pTiA6 carrying the mutated
traM
was placed in the chromosomal background of the nopaline-type strain C58C1RS, suggesting an unknown inhibitory factor(s) encoded by the chromosomal background of strain A6 but not by C58C1RS. Low-stringency Southern blotting analysis showed that strain A6, but not strain C58 and its derivatives, contains a second
traM
homologue. The homologue, designated
traM2
, has 64% and 65% identities with
traM
at the DNA and peptide levels, respectively. Similar to TraM, TraM2 is a potent antiactivator that functions by blocking TraR, the QS activator, from specific binding to the
tra
gene promoters. Deletion of
traM2
in strain A6 harboring the mutated
traM
confers a constitutive QS phenotype. The results demonstrate that the QS system in strain A6 is subjected to the dual control of TraM and TraM2.
The Transcriptional Regulator VqmA Increases Expression of the Quorum-Sensing Activator HapR in
Vibrio cholerae
Authors: Liu Zhi , Hsiao Ansel , Joelsson Adam , Zhu Jun
Published: March 2006

Vibrio cholerae
is the causative agent of the severe diarrheal disease cholera. A number of environmental stimuli regulate virulence gene expression in
V. cholerae
, including quorum-sensing signals. At high cell densities, quorum sensing in
V. cholerae
invokes a series of signal transduction pathways in order to activate the expression of the master regulator HapR, which then represses the virulence regulon and biofilm-related genes and activates protease production. In this study, we identified a transcriptional regulator, VqmA (VCA1078), that activates
hapR
expression at low cell densities. Under in vitro inducing conditions, constitutive expression of VqmA represses the virulence regulon in a HapR-dependent manner. VqmA increases
hapR
transcription as measured by the activity of the
hapR-lacZ
reporter, and it increases HapR production as measured by Western blotting. Using a heterogenous
luxCDABE
cosmid, we found that VqmA stimulates quorum-sensing regulation at lower cell densities and that this stimulation bypasses the known LuxO-small-RNA regulatory circuits. Furthermore, we showed that VqmA regulates
hapR
transcription directly by binding to its promoter region and that expression of
vqmA
is cell density dependent and autoregulated. The physiological role of VqmA is also discussed.
Role of
oxyR
in the Oral Anaerobe
Porphyromonas gingivalis
Authors: Diaz Patricia I. , Slakeski Nada , Reynolds Eric C. , Morona Renato , Rogers Anthony H. , Kolenbrander Paul E.
Published: March 2006

Porphyromonas gingivalis
is an anaerobic microorganism that inhabits the oral cavity, where oxidative stress represents a constant challenge. A putative transcriptional regulator associated with oxidative stress, an
oxyR
homologue, is known from the
P. gingivalis
W83 genome sequence. We used microarrays to characterize the response of
P. gingivalis
to H
2
O
2
and examine the role of
oxyR
in the regulation of this response. Most organisms in which
oxyR
has been investigated are facultative anaerobes or aerobes. In contrast to the OxyR-regulated response of these microorganisms to H
2
O
2
, the main feature of the response in
P. gingivalis
was a concerted up-regulation of insertion sequence elements related to IS
1
transposases. Common OxyR-regulated genes such as
dps
and
ahpFC
were not positively regulated in
P. gingivalis
in response to H
2
O
2
. However, their expression was dependent on the presence of a functional OxyR, as revealed by microarray comparison of an
oxyR
mutant to the wild type. Phenotypic characterization of the
oxyR
mutant showed that OxyR plays a role in both the resistance to H
2
O
2
and the aerotolerance of
P. gingivalis. Escherichia coli
and other bacteria with more complex respiratory requirements use OxyR for regulating resistance to H
2
O
2
and use a separate regulator for aerotolerance. In
P. gingivalis
, the presence of a single protein combining the two functions might be related to the comparatively smaller genome size of this anaerobic microorganism. In conclusion, these results suggest that OxyR does not act as a sensor of H
2
O
2
in
P. gingivalis
but constitutively activates transcription of oxidative-stress-related genes under anaerobic growth.