A relative gradient theory for layered materials

It is possible to remedy certain difficulties with the description of short wave length phenomena and interfacial slip in standard models of a laminated material by considering the bending stiffness of the layers. If the couple or moment stresses are assumed to be proportional to the relative deformation gradient, then the bending effect disappears for vanishing interface slip, and the model correctly reduces to an isotropic standard continuum. In earlier Cosserat-type models this was not the case. Laminated materials of the kind considered here occur naturally as layered rock, or at a different scale, in synthetic layered materials and composites. Similarities to the situation in regular dislocation structures with couple stresses, also make these ideas relevant to single slip in crystalline materials. Application of the theory to a one-dimensional model for layered beams demonstrates agreement with exact results at the extremes of zero and infinite interface stiffness. Moreover, comparison with finite element calculations confirm the accuracy of the prediction for intermediate interfacial stiffness.

NMR structure of C-terminally tagged gramicidin channels

A biotin group was covalently attached to the C terminus of gramicidin A (gA) through a linker arm comprising a glycine residue with either one (gAXB) or two caproyl groups (gAXXB). High-resolution two-dimensional NMR spectroscopy was used to determine the structure of these modified gA analogues and [Lys(16)]gramicidin A (gA-Lys) in sodium dodecyl-d(25) sulphate micelles. Gated gA ion channels based on linking a receptor group to these gA analogues have been used recently as a component in a sensing device. The conformations of the gA backbones and amino acid side chains of lysinated gA and biotinylated gA in detergent micelles were found to be almost identical to that of native gA, i.e. that of an N-terminal to N-terminal (head to head) dimer formed by two right-handed, single-stranded beta(6.3) helices. The biotin tail of the gAXB and gAXXB and the lysine extremity of gA-Lys appeared to lie outside the micelle. Thus it appears that the covalent attachment of functional groups to the C terminus of gA does not disrupt the peptide's helical configuration. Further, single channel measurements of all three gA analogues showed that functioning ion channels were preserved within a membrane environment. (C) 1999 Elsevier Science B.V. All rights reserved.

Modelling the large deformations in stratified media - the Cosserat continuum approach

Methods employing continuum approximation in describing the deformation of layered materials possess a clear advantage over explicit models, However, the conventional implicit models based on the theory of anisotropic continua suffers from certain difficulties associated with interface slip and internal instabilities. These difficulties can be remedied by considering the bending stiffness of the layers. This implies the introduction of moment (couple) stresses and internal rotations, which leads to a Cosserat-type theory. In the present model, the behaviour of the layered material is assumed to be linearly elastic; the interfaces are assumed to be elastic perfectly plastic. Conditions of slip or no slip at the interfaces are detected by a Coulomb criterion with tension cut off at zero normal stress. The theory is valid for large deformation analysis. The model is incorporated into the finite element program AFENA and validated against analytical solutions of elementary buckling problems in layered medium. A problem associated with buckling of the roof and the floor of a rectangular excavation in jointed rock mass under high horizontal in situ stresses is considered as the main application of the theory. Copyright (C) 1999 John Wiley & Sons, Ltd.

Structure of a putative ancestral protein encoded by a single sequence repeat from a multidomain proteinase inhibitor gene from Nicotiana alata

Background: The ornamental tobacco Nicotiana alata produces a series of proteinase inhibitors (Pls) that are derived from a 43 kDa precursor protein, NaProPl. NaProPl contains six highly homologous repeats that fold to generate six separate structural domains, each corresponding to one of the native Pls. An unusual feature of NaProPl is that the structural domains lie across adjacent repeats and that the sixth Pl domain is generated from fragments of the first and sixth repeats. Although the homology of the repeats suggests that they may have arisen from gene duplication, the observed folding does not appear to support this. This study of the solution structure of a single NaProPl repeat (aPl1) forms a basis for unravelling the mechanism by which this protein may have evolved, Results: The three-dimensional structure of aPl1 closely resembles the triple-stranded antiparallel beta sheet observed in each of the native Pls. The five-residue sequence Glu-Glu-Lys-Lys-Asn, which forms the linker between the six structural domains in NaProPl, exists as a disordered loop in aPl1. The presence of this loop in aPl1 results in a loss of the characteristically flat and disc-like topography of the native inhibitors. Conclusions: A single repeat from NaProPl is capable of folding into a compact globular domain that displays native-like Pl activity. Consequently, it is possible that a similar single-domain inhibitor represents the ancestral protein from which NaProPl evolved.

A review of the Apocreadiidae Skrjabin, 1942 (Trematoda : Digenea) and description of Australian species

The Apocreadiidae is reviewed and is considered to include genera recognised previously within the families Apocreadiidae, Homalometridae, Schistorchiidae, Sphincterostomatidae and Trematobrienidae. Key features of the family are extensive vitelline follicles, eye-spot pigment dispersed in forebody, I-shaped excretory vesicle, no cirrus-sac and genital pore opening immediately anterior to the ventral sucker (usually) or immediately posterior to it (Postporus Manter, 1949). Three subfamilies and 18 genera are recognised within the Apocreadiidae. The Apocreadiinae comprises Homalometron Stafford, 1904 (new syn. Barbulostomum Ramsey, 1965), Callohelmis n. g., Choanodera Manter, 1940, Crassicutis Manter, 1936, Dactylotrema Bravo-Hollis & Manter, 1957, Marsupioacetabulum Yamaguti, 1952, Microcreadium Simer, 1929, Myzotus Manter, 1940, Neoapocreadium Siddiqi & Cable, 1960, Neomegasolena Siddiqi & Cable, 1960, Pancreadium Manter, 1954, Procaudotestis Szidat, 1954 and Trematobrien Dollfus, 1950. The Schistorchiinae comprises Schistorchis Luhe, 1906, Sphincterostoma Yamaguti, 1937, Sphincteristomum Oshmarin, Mamaev & Parukhin, 1961 and Megacreadium Nagaty, 1956. The Postporinae comprises only Postporus. A key to subfamilies and genera of the Apocreadiidae is provided. It is argued that there is no convincing basis for the recognition of the genus Apocreadium Manter, 1937 and all its constituent species are combined with Homalometron. The following new combinations are proposed for species previously recognised within Apocreadium: Homalometron balistis (Manter, 1947), H. caballeroi (Bravo-Hollis, 1953), H. cryptum (Overstreet, 1969), H. longisinosum (Manter, 1937), H. manteri (Overstreet, 1970), H. mexicanum (Manter, 1937) and H. vinodae (Ahmad, 1985). Apocreadium uroproctoferum Sogandares-Bernal, 1959 is found to lack a uroproct and is made a synonym of H. mexicanum. Homalometron verrunculi nom. nov. is proposed to replace the secondarily pre-occupied H. caballeroi Lamothe-Argumedo, 1965. Barbulostomum is made a synonym of Homalometron and H. cupuloris (Ramsey, 1965) n. comb. is proposed. Neochoanodera is made a synonym of Choanodera and Choanodera ghanensis (Fischthal & Thomas, 1970) n. comb. is proposed. Species within the Apocreadiinae and Postporinae are reviewed and the following are recorded or described from Australian fishes: Homalometron wrightae n. sp. from Achlyopa nigra (Macleay), H. synagris (Yamaguti, 1953) n. comb. from Scolopsis monogramma (Cuvier), H. stradbrokensis n. sp. from Gerres subfasciatus Cuvier, Marsupioacetabulum opallioderma n. sp. from G. subfasciatus, Neoapocreadium karwarensis (Hafeezullah, 1970) n. comb. from G. subfasciatus, N. splendens n. sp. from S. monogramma and Callohelmis pichelinae n. g., n. sp. from Hemigymnus melapterus (Bloch), H. fasciatus (Bloch), Stethojulis bandanensis (Bleeker) andChoerodon venustus (De Vis). Callohelmis is recognised by the combination of absence of tegumental spines, caeca terminating midway between the testes and posterior end of body, ventral sucker enclosed in a tegumental pouch, prominent muscles radiating through the body from the ventral sucker, vitelline follicles not extending into the forebody, and a very short excretory vesicle that opens ventrally. New combinations for species previously recognised within Crassicutis are proposed as follows: Neoapocreadium caranxi (Bilqees, 1976) n. comb., N. gerridis (Nahhas & Cable, 1964) n. comb., N. imtiazi (Ahmad, 1984) n. comb. and N. marina (Manter, 1947) n. comb. The host-specificity and zoogeography of the Apocreadiinae are considered.

Weathering of hydrocarbons in mangrove sediments: testing the effects of using dispersants to treat oil spills

This field study was a combined chemical and biological investigation of the relative effects of using dispersants to treat oil spills impacting mangrove habitats. The aim of the chemistry was to determine whether dispersant affected the short- or long-term composition of a medium range crude oil (Gippsland) stranded in a tropical mangrove environment in Queensland, Australia. Sediment cores from three replicate plots of each treatment (oil only and oil plus dispersant) were analyzed for total hydrocarbons and for individual molecular markers (alkanes, aromatics, triterpanes, and steranes). Sediments were collected at 2 days, then 1, 7, 13 and 22 months post-spill. Over this time, oil in the six treated plots decreased exponentially from 36.6 +/- 16.5 to 1.2 +/- 0.8 mg/g dry wt. There was no statistical difference in initial oil concentrations, penetration of oil to depth, or in the rates of oil dissipation between oiled or dispersed oil plots. At 13 months, alkanes were >50% degraded, aromatics were similar to 30% degraded based upon ratios of labile to resistant markers. However, there was no change in the triterpane or sterane biomarker signatures of the retained oil. This is of general forensic interest for pollution events. The predominant removal processes were evaporation (less than or equal to 27%) and dissolution (greater than or equal to 56%), with a lag-phase of 1 month before the start of significant microbial degradation (less than or equal to 7%). The most resistant fraction of the oil that remained after 7 months (the higher molecular weight hydrocarbons) correlated with the initial total organic carbon content of the soil. Removal rate in the Queensland mangroves was significantly faster than that observed in the Caribbean and was related to tidal flushing. (C) 1999 Elsevier Science Ltd. All rights reserved.

Plant cyclotides: A unique family of cyclic and knotted proteins that defines the cyclic cystine knot structural motif

Several macrocyclic peptides (similar to 30 amino acids), with diverse biological activities, have been isolated from the Rubiaceae and Violaceae plant families over recent years. We have significantly expanded the range of known macrocyclic peptides with the discovery of 16 novel peptides from extracts of Viola hederaceae, Viola odorata and Oldenlandia affinis. The Viola plants had not previously been examined for these peptides and thus represent novel species in which these unusual macrocyclic peptides are produced. Further, we have determined the three-dimensional struc ture of one of these novel peptides, cycloviolacin O1, using H-1 NMR spectroscopy. The structure consists of a distorted triple-stranded beta-sheet and a cystine-knot arrangement of the disulfide bonds. This structure is similar to kalata B1 and circulin A, the only two macrocyclic peptides for which a structure was available, suggesting that despite the sequence variation throughout the peptides they form a family in which the overall fold is conserved. We refer to these peptides as the cyclotide family and their embedded topology as the cyclic cystine knot (CCK) motif. The unique cyclic and knotted nature of these molecules makes them a fascinating example of topologically complex proteins. Examination of the sequences reveals they can be separated into two subfamilies, one of which tends to contain a larger number of positively charged residues and has a bracelet-like circularization of the backbone. The second subfamily contains a backbone twist due to a cis-Pro peptide bond and may conceptually be regarded as a molecular Moebius strip. Here we define the structural features of the two apparent subfamilies of the CCK peptides which may be significant for the likely defense related role of these peptides within plants. (C) 1999 Academic Press.

A review of the Arhythmacanthidae (Acanthocephala) with a description of Heterosentis hirsutus n. sp. from Cnidoglanis macrocephala (Plotosidae) in Australia

Heterosentis hirsutus n. sp. is described from Cnidoglanis macrocephalo (Siluriformes: Plotosidae) from the Swan Estuary, Western Australia. It is distinguished by having 14 longitudinal rows of 6-7 hooks per row on the proboscis, a trunk armed anteriorly and posteriorly (=genital spines) with minute spines and lemnisci that may extend to the poster;or margin of the proboscis receptacle The new species also has prominent fragmented nuclei in its trunk well. New information is given for Heterosentis plotosi Yamoguti, 1935 from Plotosus lineatus (Siluriformes: Plotosidae) and H. poraplagusiarum (Nickol, 1972) Amin, 1985 from Paraplogusia guttata (Pleuronectiformes: Cynoglossidoe), both from Queensland. A key to the species of Heterosentis Van Cleave, 1931 is provided. The Arhythmacanthidae subfamilies are reviewed: there is little utility in the recognition of these taxa because of the small number of genera involved and the validity/ of the characters on which they ore based is in doubt, particularly whether trunk spines are present or absent. Only Acanthocephaloides Meyer, 1932, Breizocanthus Golvon, 1969, Euzetocanthus Golvan & Houin, 1964, Heterosentis, Hypoechinorhynchus Yamaguti, 1939 and Paracanthocepholoides Golvan, 1969 of the Arhythmacanthidae are considered valid. A key to these genera is provided. The monotypic genus Neocanthocepholoides Cable & Quick, 1954 is considered a new synonym of Acanthocephaloides thus creating Acanthocephaloides spinicaudatus (Cable & Quick, 1954) n. comb. Arhythmocanthus Yamaguti, 1935 is maintained as a synonym of Heterosentis because the distinction between two and three hook types is made equivocal when the transition between the opical and subapical hooks is gradual.

Reverse transcription of a naturally occurring nonretroviral RNA produces a precise deletion in the majority of its cDNA products

A precise, reproducible deletion made during in vitro reverse transcription of RNA2 from the icosahedral positive-stranded Helicoverpa armigera stunt virus (Tetraviridae) is described. The deletion, located between two hexamer repeats, is a 50-base sequence that includes one copy of the hexamer repeat. Only the Moloney murine leukemia virus reverse transcriptase and its derivative Superscript I, carrying a deletion of the carboxy-terminal RNase H region, showed this response, indicating a template-switching mechanism different from one proposed that involves a RNase H-dependent strand transfer, Superscript II, however, which carries point mutations to reduce RNase H activity, does not cause a deletion. A possible mechanism involves the enzyme pausing at the 3' side of a stem-loop structure and the 3' end of the nascent DNA strand separating from the template and reannealing to the upstream hexamer repeat.

Acyclic permutants of naturally occurring cyclic proteins - Characterization of cystine knot and beta-sheet formation in the macrocyclic polypeptide kalata B1

Kalata B1 is a prototypic member of the unique cyclotide family of macrocyclic polypeptides in which the major structural features are a circular peptide backbone, a triple stranded beta-sheet, and a cystine knot arrangement of three disulfide bonds. The cyclotides are the only naturally occurring family of circular proteins and have prompted us to explore the concept of acyclic permutation, i.e. opening the backbone of a cross-linked circular protein in topologically permuted ways. We have synthesized the complete suite of acyclic permutants of kalata B1 and examined the effect of acyclic permutation on structure and activity. Only two of six topologically distinct backbone loops are critical for folding into the native conformation, and these involve disruption of the embedded ring in the cystine knot. Surprisingly, it is possible to disrupt regions of the p-sheet and still allow folding into native-like structure, provided the cystine knot is intact. Kalata B1 has mild hemolytic activity, but despite the overall structure of the native peptide being retained in all but two cases, none of the acyclic permutants displayed hemolytic activity. This loss of activity is not localized to one particular region and suggests that cyclization is critical for hemolytic activity.

Preliminary experiences with a single-patient trials service in general practice

Objective: To pilot a single-patient trials (SPTs) service in general practice, designed to improve decision-making about long-term medications for chronic conditions. Design: 12-week within-patient, randomised, double-blind, placebo-controlled, crossover comparison of ibuprofen with paracetamol for osteoarthritis, involving three pairs of two-week treatment periods for each participating patient. Setting and patients: Patients attending an academic general practice with a clinical diagnosis of osteoarthritis, with pain of at least a month's duration severe enough to warrant consideration of long-term non-steroidal anti-inflammatory drug (NSAID) use. Main outcome measures: Pain and stiffness; measures of overall arthritis compared with previous fortnight; preference for NSAID at the end of each two-week treatment period; use of escape analgesia; side effects; and management changes as a result of the SPTs. Results: Eight of 14 patients completed SPTs. One was a clear responder to NSAIDs, five were non-responders, and two were indefinite. Of the five who were using NSAIDs before the SPT, two continued and three ceased using them. Clinically useful information assisted decision-making for all eight participants. Medication management changed for six. Conclusions: Single-patient trials can be successfully implemented in general practice and might be a valuable method for GPs to identify patients who respond to medication for chronic stable conditions such as osteoarthritis, in which individual response to medication is variable.

Conotoxin TVIIA, a novel peptide from the venom of Conus tulipa - 2. Three-dimensional solution structure

The three-dimensional solution structure of conotoxin TVIIA, a 30-residue polypeptide from the venom of the piscivorous cone snail Conus tulipa, has been determined using 2D H-1 NMR spectroscopy. TVIIA contains six cysteine residues which form a 'four-loop' structural framework common to many peptides from Conus venoms including the omega-, delta-, kappa-, and mu O-conotoxins. However, TVIIA does not belong to these well-characterized pharmacological classes of conotoxins, but displays high sequence identity with conotoxin GS, a muscle sodium channel blocker from Conus geographus. Structure calculations were based on 562 interproton distance restraints inferred from NOE data, together with 18 backbone and nine side-chain torsion angle restraints derived from spin-spin coupling constants. The final family of 20 structures had mean pairwise rms differences over residues 2-27 of 0.18 +/- 0.05 Angstrom for the backbone atoms and 1.39 +/- 0.33 Angstrom for all heavy atoms. The structure consists of a triple-stranded, antiparallel beta sheet with +2x, -1 topology (residues 7-9, 16-20 and 23-27) and several beta turns. The core of the molecule is formed by three disulfide bonds which form a cystine knot motif common to many toxic and inhibitory polypeptides. The global fold, molecular shape and distribution of amino-acid sidechains in TVIIA is similar to that previously reported for conotoxin GS, and comparison with other four-loop conotoxin structures provides further indication that TVIIA and GS represent a new and distinct subgroup of this structural family. The structure of TVIIA determined in this study provides the basis for determining a structure-activity relationship for these molecules and their interaction with target receptors.

Conservation of the sequence and temporal expression of let-7 heterochronic regulatory RNA

Two small RNAs regulate the timing of Caenorhabditis elegans development(1,2). Transition from the first to the second larval stage fates requires the 22-nucleotide lin-4 RNA(1,3,4), and transition from late larval to adult cell fates requires the 21-nucleotide let-7 RNA 2. The lin-4 and let-7 RNA genes are not homologous to each other, but are each complementary to sequences in the 3' untranslated regions of a set of protein-coding target genes that are normally negatively regulated by the RNAs1,2,5,6. Here we have detected let-7 RNAs of similar to 21 nucleotides in samples from a wide range of animal species, including vertebrate, ascidian, hemichordate, mollusc, annelid and arthropod, but not in RNAs from several cnidarian and poriferan species, Saccharomyces cerevisiae, Escherichia coli or Arabidopsis. We did not detect lin-4 RNA in these species. We found that let-7 temporal regulation is also conserved: let-7 RNA expression is first detected at late larval stages in C. elegans and Drosophila, at 48 hours after fertilization in zebrafish, and in adult stages of annelids and molluscs. The let-7 regulatory RNA may control late temporal transitions during development across animal phylogeny.

Three-dimensional structure of RK-1: A novel alpha-defensin peptide

NMR spectroscopy and simulated annealing calculations have been used to determine the three-dimensional structure of RK-1, an antimicrobial peptide from rabbit kidney recently discovered from homology screening based on the distinctive physicochemical properties of the corticostatins/defensins. RK-1 consists of 32 residues, including six cysteines arranged into three disulfide bonds. It exhibits antimicrobial activity against Escherichia coli and activates Ca2+ channels in vitro. Through its physicochemical similarity, identical cysteine spacing, and linkage to the corticostatins/defensins, it was presumed to be a member of this family. However, RK-1 lacks both a large number of arginines in the primary sequence and a high overall positive charge, which are characteristic of this family of peptides. The three-dimensional solution structure, determined by NMR, consists of a triple-stranded antiparallel beta -sheet and a series of turns and is similar to the known structures of other alpha -defensins. This has enabled the definitive classification of RK-1 as a member of this family of antimicrobial peptides. Ultracentrifuge measurements confirmed that like rabbit neutrophil defensins, RK-1 is monomeric in solution, in contrast to human neutrophil defensins, which are dimeric.

Neuronal sodium-channel alpha 1-subunit mutations in generalized epilepsy with febrile seizures plus

Generalized epilepsy with febrile seizures plus (GEFS+) is a familial epilepsy syndrome characterized by the presence of febrile and afebrile seizures. The first gene, GEFS1, was mapped to chromosome 19q and was identified as the sodium-channel beta1-subunit, SCN1B. A second locus on chromosome 2q, GEFS2, was recently identified as the sodium-channel alpha1-subunit, SCN1A. Single-stranded conformation analysis (SSCA) of SCN1A was performed in 53 unrelated index cases to estimate the frequency of mutations in patients with GEFS+. No mutations were found in 17 isolated cases of GEFS+. Three novel SCN1A mutations-D188V, V1353L, and I1656M-were found in 36 familial cases; of the remaining 33 families, 3 had mutations in SCN1B. On the basis of SSCA, the combined frequency of SCN1A and SCN1B mutations in familial cases of GEFS+ was found to be 17%.