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PubMed2003

Definition of domain boundaries and crystallization of the SMN Tudor domain.

Spinal muscular atropy (SMA) is the major genetic disease leading to childhood mortality and is caused by mutations in or deletions of the smn1 gene. The human survival of motor neurons (SMN) protein encoded by this gene plays an important role in the assembly of snRNPs (small nuclear ribonucleoprotein complexes) via binding to the spliceosomal Sm proteins. The tails of these Sm proteins contain symmetrically dimethylated arginines that are recognized by the central SMN Tudor domain. To gain insight in the molecular basis of this specific interaction, the SMN Tudor domain has been crystallized. The rapid crystallization of the protein and the high stability of the crystals is facilitated by redefinition of domain boundaries based on NMR relaxation experiments and the previously determined solution structure. The crystals diffract to high resolution (1.8 A) and a complete data set has been collected from a hexagonal crystal form (P6(1)/P6(5)), with unit-cell parameters a = b = 27.65, c = 110.30 A, alpha = beta = 90, gamma = 120 degrees. Crystal soaks and co-crystallization with peptides derived from the Sm protein tails have been initiated. Molecular replacement with the NMR coordinates is under way.

Crystallization↗
PubMed1993

Thresholds and stability analysis of models for the spatial spread of a fatal disease.

A simple two-class (susceptibles and infectives) model describing the dynamics of a fatal disease in a variable-size population is presented and analysed. Spatial dependence is introduced into the model by considering two different mechanisms for the geographic spread of the disease: nonlocal interaction between susceptibles and infectives, and migratory spread of the animals. The steady states and their stability for these spatially dependent models are deduced; no spatially heterogeneous steady states were possible. For nonlocal interaction, there were two spatially uniform steady states: the trivial state (no infectives or susceptibles), which was unstable, and the endemic state (constant proportion of the population infected), which was locally asymptotically stable. With migratory spread, the number of spatially uniform steady states was dependent on the boundary conditions imposed. With hostile (Dirichlet) boundary conditions, only the trivial steady state was possible and its local stability found to depend on the rate of diffusion of the total population. With no-flux (Neumann) boundary conditions, the steady states are the trivial and endemic states; these were unstable and locally asymptotically stable, respectively.

Animals↗
PubMed2000

Influence of gelatin and bovine serum lubricants on ultra-high molecular weight polyethylene wear debris generated in in vitro simulations.

Ultra-high molecular weight polyethylene (UHMWPE) wear debris induced osteolysis has a major role in the late aseptic loosening and ultimate failure of total hip replacements (THR). Clinically relevant in vitro simulations of wear are essential to predict the osteolytic potential of bearing surfaces in artificial hip joints. Newborn calf or bovine serum has been accepted as a boundary lubricant for such in vitro tests, but its biological stability has been questioned. This study compared the wear factors, number of wear particles and levels of microbial contamination produced in bovine serum and a gelatin-based lubricant. The wear factors produced by the two lubricants were not significantly different, however the wear debris morphology produced was substantially different. The bovine serum became contaminated with micro-organisms within 28 h, whereas the protein-based lubricant remained uncontaminated. The results showed that bovine serum was not a stable boundary lubricant. They also showed that although the wear factors for the two solutions were not significantly different, the protein-based lubricant was not a suitable alternative to bovine serum because the wear debris produced was not clinically relevant.

Animals↗
PubMed2000

Spatiotemporal chaos and nonequilibrium transitions in a model excitable medium

We present a detailed study of the statistical steady states of a model for CO oxidation on Pt(110) proposed by Bar and co-workers. We show that the stability diagram of this model depends sensitively on the boundary conditions. We elucidate several novel properties of a state with meandering spirals (M) briefly mentioned by Bar and co-workers. (1) We show that, with periodic boundary conditions, M is the state MP, a binary mixture displaying a coexistence of quasiperiodically rotating spirals and chaotically moving pointlike defects. We show that the transition from MP to the turbulent state T1 is continuous; the transition line marks the locus where the two phases cease to be distinct. (2) With Neumann boundary conditions M is the state MN, a single quasiperiodically rotating spiral. We show that the MN-T1 transition is discontinuous or first order. We also characterize the transitions from MP and MN to the state S, which has quasiperiodically rotating spirals. We also propose qualitative mechanisms for these transitions.

Journal Article↗
PubMed2002

Fingering instability of thin evaporating liquid films.

The fingering instability of growing dry patches in an evaporating film of a polar liquid placed on a solid substrate is investigated. The instability manifests itself as fingering of mobile fronts between growing "dry" (thin) and shrinking "wet" (thick) regions of the film corresponding to two stable states of the evaporating film in contact with its vapor. The boundaries of the fingering instability are found through linear stability analysis of numerical solutions of the nonlinear evolution equation defining the film profile, and the influence of the evaporation rate, polar intermolecular forces, and chemical heterogeneity of the substrate is investigated.

Journal Article↗
PubMed2005

Development of magnetic resonance imaging contrast material for in vivo mapping of tissue transglutaminase activity.

Transglutaminases are a family of enzymes that play an important role in tissue remodeling by catalyzing covalent cross-links between proteins of the extracellular matrix. Elevated activity of transglutaminase was shown at the boundaries of invading tumors, in association with angiogenesis, in stabilization of atherosclerotic plaques, and in generation of blood clots. The aim of this work was to develop a low molecular weight substrate of transglutaminase that could serve for noninvasive magnetic resonance and optical mapping of transglutaminase-mediated cross-linking activity. A 2 kDa contrast material was generated which showed cross-linking by either tissue transglutaminase or factor XIII in the context of multicellular tumor spheroids or fibrin clots, respectively. Successful detection by nuclear magnetic resonance microscopy of transglutaminase-mediated cross-linking of the contrast material to MCF7 multicellular spheroids provides hope that this approach could potentially be developed for clinical demarcation of sites of transglutaminase activity.

Breast Neoplasms↗
PubMed1994

Bone grafts and bone induction substitutes.

The use of bone grafts is the basis of all craniofacial surgery. Bone grafts are used to stabilize the open segments, expand the patient's structural boundaries, and fill defects created by trauma or congenital malformations. Bone grafts are harvested from local or distant sites as indicated. Biomaterial implants are used as bone substitutes when it is not possible to use bone grafts.

Adult↗
PubMed2004

Long-range attraction between particles in dusty plasma and partial surface tension of a dusty phase boundary.

Effective potential of a charged dusty particle moving in homogeneous plasma has a negative part that provides attraction between similarly charged dusty particles. A depth of this potential well is great enough to ensure both stability of crystal structure of dusty plasma and sizable value of surface tension of a boundary surface of dusty region. The latter depends on the orientation of the surface relative to the ion flow, namely, it is maximal and positive for the surface normal to the flow and minimal and negative for the surface along the flow. For the most cases of dusty plasma in a gas discharge, a value of the first of them is more than sufficient to ensure stability of lenticular dusty phase void oriented across the counter-ion flow.

Journal Article↗
PubMed1997

Human pancreatic ribonuclease--deletion of the carboxyl-terminal EDST extension enhances ribonuclease activity and thermostability.

Mammalian ribonucleases constitute one of the fastest evolving protein families in nature. The addition of a four-residue carboxyl-terminal tail: Glu-Asp-Ser-Thr (EDST) in human pancreatic ribonuclease (HPR) in comparison with bovine pancreatic RNase (RNase A) could have adaptive significance in humans. We have cloned and expressed human pancreatic ribonuclease in Escherichia coli to probe the influence of the four-residue extension and neighboring C-terminal residues on the biochemical properties of the enzyme. Removal of the C-terminal extension from HPR yielded an enzyme, HPR-(1-124)-peptide, with enhanced ability to cleave poly(C). HPR-(1-124)-peptide also exhibited a steep increase in thermal stability mimicking that known for RNase A. Wild-type HPR had significantly low thermal stability compared to RNase A. The study identifies the C-terminal boundary in the human pancreatic ribonuclease required for efficient catalysis.

Amino Acid Sequence↗
PubMed1992

Unfolding domains of recombinant fusion alpha alpha-tropomyosin.

The thermal unfolding of the coiled-coil alpha-helix of recombinant alpha alpha-tropomyosin from rat striated muscle containing an additional 80-residue peptide of influenza virus NS1 protein at the N-terminus (fusion-tropomyosin) was studied with circular dichroism and fluorescence techniques. Fusion-tropomyosin unfolded in four cooperative transitions: (1) a pretransition starting at 35 degrees C involving the middle of the molecule; (2) a major transition at 46 degrees C involving no more than 36% of the helix from the C-terminus; (3) a major transition at 56 degrees C involving about 46% of the helix from the N-terminus; and (4) a transition from the nonhelical fusion domain at about 70 degrees C. Rabbit skeletal muscle tropomyosin, which lacks the fusion peptide but has the same tropomyosin sequence, does not exhibit the 56 degrees C or 70 degrees C transition. The very stable fusion unfolding domain of fusion-tropomyosin, which appears in electron micrographs as a globular structural domain at one end of the tropomyosin rod, acts as a cross-link to stabilize the adjacent N-terminal domain. The least stable middle of the molecule, when unfolded, acts as a boundary to allow the independent unfolding of the C-terminal domain at 46 degrees C from the stabilized N-terminal unfolding domain at 56 degrees C. Thus, strong localized interchain interactions in coiled-coil molecules can increase the stability of neighboring domains.

Animals↗
PubMed2003

Molecular characterization of the principal substrate binding site of the ubiquitous folding catalyst protein disulfide isomerase.

Disulfide bond formation in the endoplasmic reticulum of eukaryotes is catalyzed by the ubiquitously expressed enzyme protein disulfide isomerase (PDI). The effectiveness of PDI as a catalyst of native disulfide bond formation in folding polypeptides depends on the ability to catalyze disulfide-dithiol exchange, to bind non-native proteins, and to trigger conformational changes in the bound substrate, allowing access to buried cysteine residues. It is known that the b' domain of PDI provides the principal peptide binding site of PDI and that this domain is critical for catalysis of isomerization but not oxidation reactions in protein substrates. Here we use homology modeling to define more precisely the boundaries of the b' domain and show the existence of an intradomain linker between the b' and a' domains. We have expressed the recombinant b' domain thus defined; the stability and conformational properties of the recombinant product confirm the validity of the domain boundaries. We have modeled the tertiary structure of the b' domain and identified the primary substrate binding site within it. Mutations within this site, expressed both in the isolated domain and in full-length PDI, greatly reduce the binding affinity for small peptide substrates, with the greatest effect being I272W, a mutation that appears to have no structural effect.

Binding Sites↗
PubMed2005

Energy dissipation and stability of propagating surfaces.

Thermodynamic equilibrium states are given by the minimum of a convex free energy function with suitable boundary conditions. Nonconvexity may lead to the coexistence of several phases and the classical Gibbs phase rule allows constructing their equilibrium properties (e.g., density or pressure). Within the framework of nonequilibrium thermodynamics, the maximization of energy dissipation (under suitable boundary conditions) can be used as an extremal principle to find stationary states. We show that stationary states generally exist for convex energy dissipation functions and that nonconvexity leads to metastable and unstable states. A geometric argument, similar in spirit to Gibbs' double-tangent construction, yields the stability limits of stationary states. This argument is applied to study a classical problem of materials science, namely the motion of a grain boundary under the influence of solute drag.

Journal Article↗
PubMed1983

Correlation between thermal stability maps and genetic maps of double-stranded DNAs.

The positional correlation of boundaries of cooperatively melting regions with boundaries of protein coding regions and mere open reading frames, and with cleavage sites of restriction enzymes and S1 nuclease are statistically examined for phi X174, G4, fd, SV40, BKV, and polyoma DNAs. A statistically significant correlation does exist in the case of boundaries of protein coding regions, but none is detected for boundaries of mere open reading frames or cleavage sites of restriction enzymes and S1 nuclease. The significant correlation disappears when the cooperativity of melting of the DNA double strand decreases to a nonphysiological condition. The result presents the first evidence showing that a physical property of DNAs correlates with their biological function.

Base Composition↗
PubMed1990

The generalized multiplicative model for viability selection at multiple loci.

Selection due to differential viability is studied in an n-locus two-allele model using a set indexation that allows the simplicity of the one-locus two-allele model to be carried to multi-locus models. The existence condition is analyzed for polymorphic equilibria with linkage equilibrium: Robbins' equilibria. The local stability condition is given for the Robbins' equilibria on the boundaries in the generalized non-epistatic selection regimes of Karlin and Liberman (1979). These generalized non-epistatic regimes include the additive selection model, the multiplicative selection model and the multiplicative interaction model, and their symmetric versions cover all the symmetric viability models.

Alleles↗
PubMed1989

A continuous analog for RNA folding.

A linear segment in which a number of pairs of intervals of equal length are identified as potential stems is the subject of a folding problem analogous to inference of RNA secondary structure. A quantity of free energy (or equivalently, energy per unit length) is associated with each stem, and the various types of loops are assigned energy costs as a function of their lengths. Inference of stable structures can then be carried out in the same way as in RNA folding. More important, perturbation of stem lengths and energy densities (modelling various mutational processes affecting nucleotide sequences) allows the delineation of domains of stability of various foldings, through the explicit calculation of their boundaries, in a low-dimensional parameter space.

Mathematics↗
PubMed2004

Transient apical polarization of Gliotactin and Coracle is required for parallel alignment of wing hairs in Drosophila.

In Drosophila, wing hairs are aligned in a distally oriented, parallel array. The frizzled pathway determines proximal-distal cell polarity in the wing; however, in frizzled pathway mutants, wing hairs remain parallel. How wing hairs align has not been determined. We have demonstrated a novel role for the septate junction proteins Gliotactin (Gli) and Coracle (Cora) in this process. Prior to prehair extension, Gli and Cora were restricted to basolateral membranes. During pupal prehair development, Gli and Cora transiently formed apical ribbons oriented from the distal wing tip to the proximal hinge. These ribbons were aligned beneath prehair bases and persisted for several hours. During this time, Gli was lost entirely from the basolateral domain. A Gliotactin mutation altered the apical polarization Gli and Cora and induced defects in hair alignment in pupal and adult stages. Genetic and cell biological assays demonstrated that Gli and Cora function to align hairs independently of frizzled. Taken together, our results indicate that Gli and Cora function as the first-identified members of a long-predicted, frizzled-independent parallel alignment mechanism. We propose a model whereby the apical polarization of Gli and Cora functions to stabilize and align prehairs relative to anterior-posterior cell boundaries during pupal wing development.

Animals↗
PubMed2004

Dynamic field sampling of airborne organophosphate triesters using solid-phase microextraction under equilibrium and non-equilibrium conditions.

A simple setup for dynamic air sampling using a solid-phase microextraction (SPME) device designed for use in the field was evaluated for organophosphate triester vapour under both equilibrium and non-equilibrium conditions. The effects of varying the applied airflows in the sampling device were evaluated in order to optimise the system with respect to the Reynolds number and magnitude of the boundary layer that developed near the surface. Further, the storage stability of the analytes was studied for both capped and uncapped 100-microm PDMS fibres. Organophosphate triesters are utilized on large scales as flame-retardants and/or plasticizers, for instance in upholstered furniture. In indoor working environments these compounds have become common components in the surrounding air. Measurements were performed in a recently furnished working environment and the concentration of tris(2-choropropyl) phosphate was found to be 7 microg m(-3).

Air Pollutants, Occupational↗
PubMed2001

Electron tomography of mitochondria after the arrest of protein import associated with Tom19 depletion.

In a mutant form of Neurospora crassa, in which sheltered RIP (repeat induced point mutation) was used to deplete Tom19, protein transport through the TOM/TIM pathway is arrested by the addition of p-fluorophenylalanine (FPA). Using intermediate-voltage electron tomography, we have generated three-dimensional reconstructions of 28 FPA-treated mitochondria at four time points (0-32 h) after the addition of FPA. We determined that the cristae surface area and volume were lost in a roughly linear manner. A decrease in mitochondrial volume was not observed until after 16 h of FPA treatment. The inner boundary membrane did not appear to shrink or contract away from the outer membrane. Interestingly, the close apposition of these membranes remained over the entire periphery, even after all of the cristae had disappeared. The different dynamics of the shrinkage of cristae membrane and inner boundary membrane has implications for compartmentalization of electron transport proteins. Two structurally distinct types of contact sites were observed, consistent with recently published work. We determined that the cristae in the untreated (control) mitochondria are all lamellar. The cristae of FPA-treated mitochondria retain the lamellar morphology as they reduce in size and do not adopt tubular shapes. Importantly, the crista junctions exhibit tubular as well as slot-like connections to the inner boundary membrane, persisting until the cristae disappear, indicating that their stability is not dependent on continuous protein import through the complex containing Tom19.

Fungal Proteins↗