Lessons from tomographic studies of the mammalian Golgi

Basic structure studies of the biosynthetic machinery of the cell by electron microscopy (EM) have underpinned much of our fundamental knowledge in the areas of molecular cell biology and membrane traffic. Driven by our collective desire to understand how changes in the complex and dynamic structure of this enigmatic organelle relate to its pivotal roles in the cell, the comparatively high-resolution glimpses of the Golgi and other compartments of the secretory pathway offered to us through EM have helped to inspire the development and application of some of our most informative, complimentary (molecular, biochemical and genetic) approaches. Even so, no one has yet even come close to relating the basic molecular mechanisms of transport, through and from the Golgi, to its ultrastructure, to everybody's satisfaction. Over the past decade, EM tomography has afforded new insights into structure -function relationships of the Golgi and provoked a re-evaluation of older paradigms. By providing a set of tools for structurally dissecting cells at high-resolution in three-dimensions (3D), EM tomography has emerged as a method for studying molecular cell biology in situ. As we move rapidly toward the establishment of molecular atlases of organelles through advances in proteomics and genomics, tomographic studies of the Golgi offer the tantalizing possibility that one day, we will be able to map the spatio-temporal coordinates of Golgi-related proteins and lipids accurately in the context of 4D cellular space. (c) 2005 Elsevier B.V. All rights reserved.

Structure and folding of potato type II proteinase inhibitors: Circular permutation and intramolecular domain swapping

Potato type II serine proteinase inhibitors are proteins that consist of multiple sequence repeats, and exhibit a multidomain structure. The structural domains are circular permutations of the repeat sequence.. as a result or intramolecular domain swapping. Structural studies give indications for the origins of this folding behaviour, and the evolution of the inhibitor family.

Conotoxins as research tools and drug leads

The complex mixture of biologically active peptides that constitute the venom of Conus species provides a rich source of ion channel neurotoxins. These peptides, commonly known as conotoxins, exhibit a high degree of selectivity and potency for different ion channels and their subtypes making them invaluable tools for unravelling the secrets of the nervous system. Furthermore, several conotoxin molecules have profound applications in drug discovery, with some examples currently undergoing clinical trials. Despite their relatively easy access by chemical synthesis, rapid access to libraries of conotoxin analogues for use in structure-activity relationship studies still poses a significant limitation. This is exacerbated in conotoxins containing multiple disulfide bonds, which often require synthetic strategies utilising several steps. This review will examine the structure and activity of some of the known classes of conotoxins and will highlight their potential as neuropharmacological tools and as drug leads. Some of the classical and more recent approaches to the chemical synthesis of conotoxins, particularly with respect to the controlled formation of disulfide bonds will be discussed in detail. Finally, some examples of structure-activity relationship studies will be discussed, as well as some novel approaches for designing conotoxin analogues.

CFD simulations of hydrocyclones with an air core - Comparison between large eddy simulations and a second moment closure

CFD simulations of the 75 mm, hydrocyclone of Hsieh (1988) have been conducted using Fluent TM. The simulations used 3-dimensional body fitted grids. The simulations were two phase simulations where the air core was resolved using the mixture (Manninen et al., 1996) and VOF (Hirt and Nichols, 1981) models. Velocity predictions from large eddy simulations (LES), using the Smagorinsky-Lilly sub grid scale model (Smagorinsky, 1963; Lilly, 1966) and RANS simulations using the differential Reynolds stress turbulence model (Launder et al., 1975) were compared with Hsieh's experimental velocity data. The LES simulations gave very good agreement with Hsieh's data but required very fine grids to predict the velocities correctly in the bottom of the apex. The DRSM/RANS simulations under predicted tangential velocities, and there was little difference between the velocity predictions using the linear (Launder, 1989) and quadratic (Speziale et al., 1991) pressure strain models. Velocity predictions using the DRSM turbulence model and the linear pressure strain model could be improved by adjusting the pressure strain model constants.

Engineered T cell receptors and their potential in molecular medicine

T cell receptors are among the most specific biological structures found in nature and are therefore excellent candidates for the molecular targeting of antigen. It is becoming increasingly apparent that common sets of T cell receptors are frequently used in humans to combat pathogen and cancer derived threats. Given that many of these conserved T cell receptors have high affinity for their target ligands, there is potential to amass virtual banks of “off-the-shelf” receptors for use in a wide range of immunotherapeutic strategies. Additionally, such T cell receptors could become basic blueprints for artificial enhancement through mutagenesis, thereby creating an even better 3-dimensional fit for their cognate targets. Indeed, preliminary approaches using both “natural” and “supernatural” T cell receptors have shown promise in treating autoimmunity and malignancy. This review will discuss these studies and other approaches through which T cell receptors can be exploited in immunodiagnostics, pathogen control and gene therapy.

Factors influencing the stability of cyclotides: Proteins with a circular backbone and cystine knot motif

Cyclotides are a large family of mini-proteins that have the distinguishing features of a head-to-tail cyclised backbone and a cystine knot formed by six conserved cysteine residues. They are present in plants from the Rubiaceae, Violaceae and Cucurbitaceae families. The unique structural features of the cyclotides make them extremely resistant to chemical, thermal and proteolytic degradation. In this article we review recent Studies from our laboratory that dissect the role of the individual structural elements in defining the stability of cyclotides. The resistance of cyclotides to chemical and proteolytic degradation is in large part due to the cystine knot, whereas the thermal stability is I composite of several features including the cystine knot, the cyclic backbone and the hydrogen bonding network. A range of biological activities of cyclotides is critically dependent oil the presence of the cyclic backbone.

NMR of conotoxins: structural features and an analysis of chemical shifts of post-translationally modified amino acids

Conotoxins are small conformationally constrained peptides found in the venom of marine snails of the genus Conus. They are usually cysteine rich and frequently contain a high degree of post-translational modifications such as C-terminal amidation, hydroxylation, carboxylation, bromination, epimerisation and glycosylation. Here we review the role of NMR in determining the three-dimensional structures of conotoxins and also provide a compilation and analysis of H-1 and C-13 chemical shifts of post-translationally modified amino acids and compare them with data from common amino acids. This analysis provides a reference source for chemical shifts of post-translationally modified amino acids. Copyright (C) 2006 John Wiley & Sons, Ltd.

Omega-conotoxins GVIA, MVIIA and CVID: SAR and clinical potential

Highly selective N-type voltage-gated calcium (Ca-V) channel inhibitors from cone snail venom (the omega-conotoxins) have emerged as a new class of therapeutics for the treatment of chronic and neuropathic pain. Earlier in 2005, Prialt ( Elan) or synthetic omega-conotoxin MVIIA, was the first omega-conotoxin to be approved by Food and Drug Administration for human use. This review compares the action of three omega-conotoxins, GVIA, MVIIA and CVID, describing their structure-activity relationships and potential as leads for the design of improved N-type therapeutics that are more useful in the treatment of chronic pain.

Edge-coloured cube decompositions

An edge-colored graph is a graph H together with a function f:E(H) → C where C is a set of colors. Given an edge-colored graph H, the graph induced by the edges of color c C is denoted by H(c). Let G, H, and J be graphs and let μ be a positive integer. A (J, H, G, μ) edge-colored graph decomposition is a set S = {H 1,H 2,...,H t} of edge-colored graphs with color set C = {c 1, c 2,..., c k} such that Hi ≅ H for 1 ≤ i ≤ t; Hi (cj) ≅ G for 1 ≤ i ≤ t and ≤ j ≤ k; and for j = 1, 2,..., k, each edge of J occurs in exactly μ of the graphs H 1(c j ), H 2(c j ),..., H t (c j ). Let Q 3 denote the 3-dimensional cube. In this paper, we find necessary and sufficient conditions on n, μ and G for the existence of a (K n ,Q 3,G, μ) edge-colored graph decomposition. © Birkhäuser Verlag, Basel 2007.

Anisotropic viscous models of large-deformation Mohr-Coulomb failure

We have developed a way to represent Mohr-Coulomb failure within a mantle-convection fluid dynamics code. We use a viscous model of deformation with an orthotropic viscoplasticity (a different viscosity is used for pure shear to that used for simple shear) to define a prefered plane for slip to occur given the local stress field. The simple-shear viscosity and the deformation can then be iterated to ensure that the yield criterion is always satisfied. We again assume the Boussinesq approximation, neglecting any effect of dilatancy on the stress field. An additional criterion is required to ensure that deformation occurs along the plane aligned with maximum shear strain-rate rather than the perpendicular plane, which is formally equivalent in any symmetric formulation. We also allow for strain-weakening of the material. The material can remember both the accumulated failure history and the direction of failure. We have included this capacity in a Lagrangian-integration-point finite element code and show a number of examples of extension and compression of a crustal block with a Mohr-Coulomb failure criterion. The formulation itself is general and applies to 2- and 3-dimensional problems.

Structured engagement in the extended enterprise

Discusses the necessity for the conscious recognition of the phenomenon known as the extended enterprise; this demands that product, process and supply chain design are all considered simultaneously. Structure must be given to the extended enterprise in order to understand and manage it efficaciously. The authors discuss multiple perspectives for doing this, and employ the notions of “3-dimensional concurrent engineering” and “holonic thinking” for conceiving what the structure may look like. Describes a current “action research” project that is investigating potential lead-time reductions within an extended enterprise’s product introduction process. This aims to produce process visualisations, a framework for structuring and sychronising phases and stage-gates within the extended enterprise, and a new simulation tool which will provide a synthetic distributed hypermedia network. These deliverables will be used to play strategic “games” to explore problem issues within the product introduction process that belongs to the extended enterprise, develop teamwork across autonomous companies, and ultimately, contribute to the design of future extended enterprise supply chains.

Approximate Bayesian techniques for inference in stochastic dynamical systems

This thesis is concerned with approximate inference in dynamical systems, from a variational Bayesian perspective. When modelling real world dynamical systems, stochastic differential equations appear as a natural choice, mainly because of their ability to model the noise of the system by adding a variant of some stochastic process to the deterministic dynamics. Hence, inference in such processes has drawn much attention. Here two new extended frameworks are derived and presented that are based on basis function expansions and local polynomial approximations of a recently proposed variational Bayesian algorithm. It is shown that the new extensions converge to the original variational algorithm and can be used for state estimation (smoothing). However, the main focus is on estimating the (hyper-) parameters of these systems (i.e. drift parameters and diffusion coefficients). The new methods are numerically validated on a range of different systems which vary in dimensionality and non-linearity. These are the Ornstein-Uhlenbeck process, for which the exact likelihood can be computed analytically, the univariate and highly non-linear, stochastic double well and the multivariate chaotic stochastic Lorenz '63 (3-dimensional model). The algorithms are also applied to the 40 dimensional stochastic Lorenz '96 system. In this investigation these new approaches are compared with a variety of other well known methods such as the ensemble Kalman filter / smoother, a hybrid Monte Carlo sampler, the dual unscented Kalman filter (for jointly estimating the systems states and model parameters) and full weak-constraint 4D-Var. Empirical analysis of their asymptotic behaviour as a function of observation density or length of time window increases is provided.

Simulation based knowledge elicitation:effect of visual representation and model parameters

Since much knowledge is tacit, eliciting knowledge is a common bottleneck during the development of knowledge-based systems. Visual interactive simulation (VIS) has been proposed as a means for eliciting experts’ decision-making by getting them to interact with a visual simulation of the real system in which they work. In order to explore the effectiveness and efficiency of VIS based knowledge elicitation, an experiment has been carried out with decision-makers in a Ford Motor Company engine assembly plant. The model properties under investigation were the level of visual representation (2-dimensional, 2½-dimensional and 3-dimensional) and the model parameter settings (unadjusted and adjusted to represent more uncommon and extreme situations). The conclusion from the experiment is that using a 2-dimensional representation with adjusted parameter settings provides the better simulation-based means for eliciting knowledge, at least for the case modelled.

A CFD approach on the effect of particle size on char entrainment in bubbling fluidised bed reactors

The fluid – particle interaction inside a 41.7 mg s-1 fluidised bed reactor is modelled. Three char particles of sizes 500 µm, 250 µm, and 100 µm are injected into the fluidised bed and the momentum transport from the fluidising gas and fluidised sand is modelled. Due to the fluidising conditions and reactor design the char particles will either be entrained from the reactor or remain inside the bubbling bed. The particle size is the factor that differentiates the particle motion inside the reactor and their efficient entrainment out of it. A 3-Dimensional simulation has been performed with a completele revised momentum transport model for bubble three-phase flow according to the literature as an extension to the commercial finite volume code FLUENT 6.2.