A system for the heterologous expression of complex redox proteins in Rhodobacter capsulatus: characterisation of recombinant sulphite : cytochrome c oxidoreductase from Starkeya novella

The phototrophic purple non-sulfur bacterium Rhodobacter capsulatus expresses a wide variety of complex redox proteins in response to changing environmental conditions. Here we report the construction and evaluation of an expression system for recombinant proteins in that organism which makes use of the dor promoter from the same organism. A generic expression vector, pDorEX, was constructed and used to express sulphite:cytochrome c oxidoreductase from Starkeya novella, a heterodimeric protein containing both molybdenum and haem c. The recombinant protein was secreted to the periplasm and its biochemical properties were very similar to those of the native enzyme. The pDorEX system therefore seems to be potentially useful for heterologous expression of multi-subunit proteins containing complex redox cofactors. (C) 2002 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.

Phase variable restriction-modification systems in Moraxella catarrhalis

A repetitive DNA motif was used as a marker to identify novel genes in the mucosal pathogen Moraxella catarrhalis. There is a high prevalence of such repetitive motifs in virulence genes that display phase variable expression. Two repeat containing loci were identified using a digoxigenin-labelled 5'-(CAAC)(6)-3' oligonucleotide probe. The repeats are located in the methylase components of two distinct type III restriction-modification (R-M) systems. We suggest that the phase variable nature of these R-M systems indicates that they have an important role in the biology of M. catarrhalis. (C) 2002 Published by Elsevier Science B.V. on behalf of the Federation of European Microbiological Societies.

A variable stepsize implementation for stochastic differential equations

Stochastic differential equations (SDEs) arise from physical systems where the parameters describing the system can only be estimated or are subject to noise. Much work has been done recently on developing higher order Runge-Kutta methods for solving SDEs numerically. Fixed stepsize implementations of numerical methods have limitations when, for example, the SDE being solved is stiff as this forces the stepsize to be very small. This paper presents a completely general variable stepsize implementation of an embedded Runge Kutta pair for solving SDEs numerically; in this implementation, there is no restriction on the value used for the stepsize, and it is demonstrated that the integration remains on the correct Brownian path.

Role of peripheral afference during acquisition of a complex coordination task

It has long been supposed that the interference observed in certain patterns of coordination is mediated, at least in part, by peripheral afference from the moving limbs. We manipulated the level of afferent input, arising from movement of the opposite limb, during the acquisition of a complex coordination task. Participants learned to generate flexion and extension movements of the right wrist, of 75degrees amplitude, that were a quarter cycle out of phase with a 1-Hz sinusoidal visual reference signal. On separate trials, the left wrist either was at rest, or was moved passively by a torque motor through 50degrees, 75degrees or 100degrees, in synchrony with the reference signal. Five acquisition sessions were conducted on successive days. A retention session was conducted I week later. Performance was initially superior when the opposite limb was moved passively than when it was static. The amplitude and frequency of active movement were lower in the static condition than in the driven conditions and the variation in the relative phase relation across trials was greater than in the driven conditions. In addition, the variability of amplitude, frequency and the relative phase relation during each trial was greater when the opposite limb was static than when driven. Similar effects were expressed in electromyograms. The most marked and consistent differences in the accuracy and consistency of performance (defined in terms of relative phase) were between the static condition and the condition in which the left wrist was moved through 50degrees. These outcomes were exhibited most prominently during initial exposure to the task. Increases in task performance during the acquisition period, as assessed by a number of kinematic variables, were generally well described by power functions. In addition, the recruitment of extensor carpi radialis (ECR), and the degree of co-contraction of flexor carpi radialis and ECR, decreased during acquisition. Our results indicate that, in an appropriate task context, afferent feedback from the opposite limb, even when out of phase with the focal movement, may have a positive influence upon the stability of coordination.

On the overabundance of light rare earth elements in terrestrial zircons and its implication for Earth's earliest magmatic differentiation

We present whole-rock and zircon rare earth element (REE) data from two early Archaean gneisses (3.81 Ga and 3.64 Ga) from the Itsaq gneiss complex, south-west Greenland. Both gneisses represent extremely rare examples of unaltered, fresh and relatively undeformed igneous rocks of such antiquity. Cathodoluminescence imaging of their zircons indicates a single crystallisation episode with no evidence for either later metamorphic and/or anatectic reworking or inheritance of earlier grains. Uniform, single-population U/Pb age data confirm the structural simplicity of these zircons. One sample, a 3.64 Ga granodioritic gneiss from the Gothabsfjord, yields a chondrite-normalised REE pattern with a positive slope from La to Lu as well as substantial positive Ce and slight negative Eu anomalies, features generally considered to be typical of igneous zircon. In contrast, the second sample, a 3.81 Ga tonalite from south of the Isua Greenstone Belt, has variable but generally much higher light REE abundances, with similar middle to heavy REE. Calculation of zircon/melt distribution coefficients (D-REE(zircon/melt)) from each sample yields markedly different values for the trivalent REE (i.e. Ce and Eu omitted) and simple application of one set of D-REE(zircon/melt) to model the melt composition for the other sample yields concentrations that are in error by up to two orders of magnitude for the light REE (La-Nd). The observed light REE overabundance in the 3.81 Ga tonalite is a commonly observed feature in terrestrial zircons for which a number of explanations ranging from lattice strain to disequilibrium crystallisation have been proposed and are further investigated herein. Regardless of the cause of light REE overabundance, our study shows that simple application of zircon/melt distribution coefficients is not an unambiguous method for ascertaining original melt composition. In this context, recent studies that use REE data to claim that > 4.3 Ga Hadean detrital zircons originally crystallised from an evolved magma, in turn suggesting the operation of geological processes in the early Earth analogous to those of the present day (e.g. subduction and melting of hydrated oceanic crust), must be regarded with caution. Indeed, comparison of terrestrial Hadean and > 3.9 Ga lunar highland zircons shows remarkable similarities in the light REE, even though subduction processes that have been used to explain the terrestrial zircons have never operated on the Moon. (C) 2002 Elsevier Science B.V. All rights reserved.

Robust model-order reduction of complex biological processes

This paper addresses robust model-order reduction of a high dimensional nonlinear partial differential equation (PDE) model of a complex biological process. Based on a nonlinear, distributed parameter model of the same process which was validated against experimental data of an existing, pilot-scale BNR activated sludge plant, we developed a state-space model with 154 state variables in this work. A general algorithm for robustly reducing the nonlinear PDE model is presented and based on an investigation of five state-of-the-art model-order reduction techniques, we are able to reduce the original model to a model with only 30 states without incurring pronounced modelling errors. The Singular perturbation approximation balanced truncating technique is found to give the lowest modelling errors in low frequency ranges and hence is deemed most suitable for controller design and other real-time applications. (C) 2002 Elsevier Science Ltd. All rights reserved.

Genotyping of methicillin-resistant Staphylococcus aureus by assaying for the presence of variable elements associated with mecA

The region surrounding mecA in methicillin-resistant Staphylococcus aureus (MRSA) is highly variable. We describe an approach for the rapid genotyping of MRSA by assaying for the presence or absence of variable or mobile elements previously shown to be associated with the mecA region.

Simulation of the influence of rate- and state-dependent friction on the macroscopic behavior of complex fault zones with the lattice solid model

In order to understand the earthquake nucleation process, we need to understand the effective frictional behavior of faults with complex geometry and fault gouge zones. One important aspect of this is the interaction between the friction law governing the behavior of the fault on the microscopic level and the resulting macroscopic behavior of the fault zone. Numerical simulations offer a possibility to investigate the behavior of faults on many different scales and thus provide a means to gain insight into fault zone dynamics on scales which are not accessible to laboratory experiments. Numerical experiments have been performed to investigate the influence of the geometric configuration of faults with a rate- and state-dependent friction at the particle contacts on the effective frictional behavior of these faults. The numerical experiments are designed to be similar to laboratory experiments by DIETERICH and KILGORE (1994) in which a slide-hold-slide cycle was performed between two blocks of material and the resulting peak friction was plotted vs. holding time. Simulations with a flat fault without a fault gouge have been performed to verify the implementation. These have shown close agreement with comparable laboratory experiments. The simulations performed with a fault containing fault gouge have demonstrated a strong dependence of the critical slip distance D-c on the roughness of the fault surfaces and are in qualitative agreement with laboratory experiments.

Cadherin-directed actin assembly: E-cadherin physically associates with the Arp2/3 complex to direct actin assembly in nascent adhesive contacts

Cadherin cell adhesion molecules are major determinants of tissue patterning which function in cooperation with the actin cytoskeleton [1-4]. In the context of stable adhesion [1], cadherin/catenin complexes are often envisaged to passively scaffold onto cortical actin filaments. However, cadherins also form dynamic adhesive contacts during wound healing and morphogenesis [2]. Here actin polymerization has been proposed to drive cell surfaces together [5], although F-actin reorganization also occurs as cell contacts mature [6]. The interaction between cadherins and actin is therefore likely to depend on the functional state of adhesion. We sought to analyze the relationship between cadherin homophilic binding and cytoskeletal activity during early cadherin adhesive contacts. Dissecting the specific effect of cadherin ligation alone on actin regulation is difficult in native cell-cell contacts, due to the range of juxtacrine signals that can arise when two cell surfaces adhere [7]. We therefore activated homophilic ligation using a specific functional recombinant protein. We report the first evidence that E-cadherin associates with the Arp2/3 complex actin nucleator and demonstrate that cadherin binding can exert an active, instructive influence on cells to mark sites for actin assembly at the cell surface.

Exploring complex fitness surfaces: Multiple ornamentation and polymorphism in male guppies

The sexual ornamentation used by male guppies to attract females comprises many components, each of which varies considerably among males. Although natural and sexual selection have been shown to contribute to divergence among populations in male sexual ornaments, the role of sexual selection in maintaining polymorphism within populations is less clear. We used both parametric quadratic regression and nonparametric projection pursuit regression techniques to reveal the major axes of non-linear sexual selection on male ornaments. We visualized the fitness surfaces defined by these axes using thin-plate splines to allow a direct comparison of the two methodologies. Identification of the major axes of selection and their visualization was critical in determining the form and strength of nonlinear selection. Both types of analysis revealed fitness surfaces comprising three peaks, suggesting that there is more than one way to make an attractive guppy. Disruptive selection may be an important process underlying the presence of multiple sexual ornaments and may contribute to the maintenance of the high levels of polymorphism in male sexual ornaments found in guppy populations.