The cDNA cloning and mRNA expression of heat shock protein 70 gene in the haemocytes of bay scallop (Argopecten irradians, Lamarck 1819) responding to bacteria challenge and naphthalin stress

Heat shock protein 70 (HSP70) is an important member of the heat shock protein superfamily, and it plays a key role in the process of protecting cells, facilitating the folding of nascent peptides and responding to stress. The cDNA of bay scallop Argopecten irradians HSP70 (designated AIHSP70) was cloned by the techniques of homological cloning and rapid amplification of cDNA end (RACE). The full length of AIHSP70 cDNA was 2651 bp in length, having a 5' untranslated region (UTR) of 96 bp, a 3' UTR of 575 bp, and an open reading frame (ORF) of 1980 bp encoding a polypeptide of 659 amino acids with an estimated molecular mass of 71.80 kDa and an estimated isoelectric point of 5.26. BLAST analysis revealed that the AIHSP70 gene shared high identity with other known HSP70 genes. Three classical HSP signature motifs were detected in AIHSP70 by InterPro, analysis. 3-D structural prediction of AIHSP70 showed that its N terminal ATPase activity domain and,C terminal substrate-binding domain shared high similarity with that in human heat shock protein 70. The results indicated that the AIHSP70 was a member of the heat shock protein 70 family. A semi-quantitive RT-PCR method was used to analyse the expression of AIHSP70 gene after the treatment of naphthalin which is one kind of polycyclic aromatic hydrocarbon (PAH) and the challenge of bacteria. mRNA expression of AIHSP70 in scallop was up-regulated significantly after the stimulation of naphthalin and increased with increasing naphthalin concentration. A clearly time-dependent expression pattern of AIHSP70 was observed after the scallops were infected by Vibrio anguillarum, and the mRNA expression reached a maximum level at 8 h and lasted to 16 h, and then dropped progressively. The results indicated that AIHSP70 could play an important role in mediating the environmental stress and immune response in scallop. (c) 2006 Elsevier Ltd. All rights reserved.

Effects of Charge Location on the Absorptions and Lifetimes of Protonated Tyrosine Peptides in Vacuo

Nearby charges affect the electronic energy levels of chromophores, with the extent of the effect being determined by the magnitude of the charge and degree of charge-chromophore separation. The molecular configuration dictates the charge chromophore distance. Hence, in this study, we aim to assess how the location of the charge influences the absorption of a set of model protonated and diprotonated peptide ions, and whether spectral differences are large enough to be identified. The studied ions were the dipeptide YK, the tripeptide KYK (Y = tyrosine; K = lysine) and their complexes with 18-crown-6-ether (CE). The CE targets the ammonium group by forming internal ionic hydrogen bonds and limits the folding of the peptide. In the tripeptide, the distance between the chromophore and the backbone ammonium is enlarged relative to that in the dipeptide. Experiments were performed in an electrostatic ion storage ring using a tunable laser system, and action spectra based on lifetime measurements were obtained in the range from 210 to 310 nm. The spectra are all quite similar though there seems to be some changes in the absorption band between 210 and 250 nm, while in the lower energy band all ions had a maximum absorption at similar to 275 nm. Lifetimes after photoexcitation were found to shorten upon protonation and lengthen upon CE complexation, in accordance with the increased number of degrees of freedom and an increase in activation energies for dissociation as the mobile proton model is no longer operative.

Octave Uzanne’s L’Éventail: a literary mythology of the fan

In the last half of the nineteenth century, the folding fan was phenomenally popular in France. The accessory was a ubiquitous component of women’s dress, yet it also attracted the attention of some prominent collectors and Orientalists as well as acquiring an importance in the art and literature of the period. In many plastic works and literary texts devoted to it, the fan retains a link with femininity, and particularly with feminine sexuality, even as its identity as an art object is emphasized. Octave Uzanne’s L’Éventail (1882), a self-professed literary history of the fan, exemplifies this dualistic treatment as it presents the fan both as a titillating intimate companion of women and as a literary and (although to a lesser extent) art historical subject. This article focuses on Uzanne’s treatment of the fan’s early history in the Far and Middle East. By comparing his text with other contemporary histories of the fan, it demonstrates that the “history” of the accessory may be more accurately described as a mythology.

Letter regarding a special service held by DeVeaux School to re-inter the body of Mrs. Maria Woodruff

Letter (2 page typed form letter with a handwritten note from Don Loker) regarding a special service held by DeVeaux School to re-inter the body of Mrs. Maria Woodruff, the first wife of Judge Samuel DeVeaux. Her body was originally buried at DeVeaux School in Niagara Falls, New York. The location of the ceremony was the cemetery in St. Davids, Ontario. Maria died on April 23, 1815 at the age of 19. This notice was sent by Rev. Alec Pudwell, chaplain of DeVeaux School, Niagara Falls, New York, May 6, 1963.

Spiral jetty, thirty seven years later: the cinematic time of James Benning

This article considers cinematic time in James Benning’s film, casting a glance (2007), in relation to its subject, Robert Smithson’s 1970 earthwork Spiral Jetty, and his film of the same name. The radicalism of Smithson’s thinking on time has been widely acknowledged, and his influence continues to pervade contemporary artistic practice. The relationship of Benning’s films with this legacy may appear somewhat oblique, given their apparent phenomenological rendition of ‘real time’. However, closer examination of Benning’s formal strategies reveals a more complex temporal construction, characterized by uncertain intervals that interrupt the folding of cinematic time into the flow of consciousness. Smithson’s film uses cinematic analogy to gesture towards vast reaches of geological time; Benning’s film creates a simulated timescale to evoke the short history of the earthwork itself. Smithson’s embrace of the entropic was a counter-cultural stance at the end of the1960s, but under the shadow of ecological disaster, this orientation has come to appear melancholy and romantic rather than radical. Benning’s film returns the jetty to anthropic time, but raises questions about the ways we inhabit time. His practice of working with ‘borrowed time’ is particularly suited to the cultural and historical moment of his later work.

Stabilization of the i-motif structure by the intra-strand cross-link formation

The i-motif structures are formed by oligonucleotides containing cytosine tracts under acidic conditions. The folding of the i-motif under physiological conditions is of great interest because of its biological role. In this study, we investigated the effect of the intra-strand cross-link on the stability of the i-motif structure. The 4-vinyl-substituted analog of thymidine (T-vinyl) was incorporated into the 5′-end of the human telomere complementary strand, which formed the intra-strand cross-link with the internal adenine. The intra-strand cross-linked i-motif displayed CD spectra similar to that of the natural i-motif at acidic pH, which was transformed into a random coil with the increasing pH. The pH midpoint for the transition from the i-motif to random coil increased from pH 6.1 for the natural one to pH 6.8 for the cross-linked one. The thermodynamic parameters were obtained by measuring the thermal melting behaviors by CD and UV, and it was determined that the intra-strand cross-linked i-motif is stabilized due to a favorable entropy effect. Thus, this study has clearly indicated the validity of the intra-strand cross-linking for stabilization of the i-motif structure.

On the Denaturation Mechanisms of the Ligand Binding Domain of Thyroid Hormone Receptors

The ligand binding domain (LBD) of nuclear hormone receptors adopts a very compact, mostly alpha-helical structure that binds specific ligands with very high affinity. We use circular dichroism spectroscopy and high-temperature molecular dynamics Simulations to investigate unfolding of the LBDs of thyroid hormone receptors (TRs). A molecular description of the denaturation mechanisms is obtained by molecular dynamics Simulations of the TR alpha and TR beta LBDs in the absence and in the presence of the natural ligand Triac. The Simulations Show that the thermal unfolding of the LBD starts with the loss of native contacts and secondary Structure elements, while the Structure remains essentially compact, resembling a molten globule state. This differs From most protein denaturation simulations reported to date and suggests that the folding mechanism may start with the hydrophobic collapse of the TR LBDs. Our results reveal that the stabilities of the LBDs of the TR alpha and TR beta Subtypes are affected to different degrees by the binding of the isoform selective ligand Triac and that ligand binding confers protection against thermal denaturation and unfolding in a subtype specific manner. Our Simulations indicate two mechanisms by which the ligand stabilizes the LBD: (1) by enhancing the interactions between H8 and H 11, and the interaction of the region between H I and the Omega-loop with the core of the LBD, and (2) by shielding the hydrophobic H6 from hydration.

Coordinate and time-dependent diffusion dynamics in protein folding

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Computational studies of the reversible domain swapping of p13suc1

A minimalist representation of protein structures using a Go- like potential for interactions is implemented to investigate the mechanisms of the domain swapping of p13suc1, a protein that exists in two native conformations: a monomer and a domain- swapped dimer formed by the exchange of a beta- strand. Inspired by experimental studies which showed a similarity of the transition states for folding of the monomer and the dimer, in this study we justify this similarity in molecular descriptions. When intermediates are populated in the simulations, formation of a domain- swapped dimer initiates from the ensemble of unfolded monomers, given by the fact that the dimer formation occurs at the folding/ unfolding temperature of the monomer ( T-f). It is also shown that transitions, leading to a dimer, involve the presence of two intermediates, one of them has a dimeric form and the other is monomeric; the latter is much more populated than the former. However, at temperatures lower than T-f, the population of intermediates decreases. It is argued that the two folded forms may coexist in absence of intermediates at a temperature much lower than T-f. Computational simulations enable us to find a mechanism, `` lock- and- dock'', for domain swapping of p13suc1. To explore the route toward dimer formation, the folding of unstructured monomers must be retarded by first locking one of the free ends of each chain. Then, the other free termini could follow and dock at particular regions, where most intrachain contacts are formed, and thus de. ne the transition states of the dimer. The simulations also showed that a decrease in the maximum distance between monomers increased their stability, which is explained based on confinement arguments. Although the simulations are based on models extracted from the native structure of the monomer and the dimer of p13suc1, the mechanism of the domain- swapping process could be general, not only for p13suc1.

Structures of the noncovalent complexes of human and bovine prothrombin fragment 2 with human PPACK-thrombin

Both human and bovine prothrombin fragment 2 (the second kringle) have been cocrystallized separately with human PPACK (D-Phe-Pro-Arg)-thrombin, and the structures of these noncovalent complexes have been determined and refined (R = 0.155 and 0.157, respectively) at 3.3-Å resolution using X-ray crystallographic methods. The kringles interact with thrombin at a site that has previously been proposed to be the heparin binding region. The latter is a highly electropositive surface near the C-terminal helix of thrombin abundant in arginine and lysine residues. These form salt bridges with acidic side chains of kringle 2. Somewhat unexpectedly, the negative groups of the kringle correspond to an enlarged anionic center of the lysine binding site of lysine binding kringles such as plasminogens K1 and K4 and TPA K2. The anionic motif is DGDEE in prothrombin kringle 2. The corresponding cationic center of the lysine binding site region has an unfavorable Arg70Asp substitution, but Lys35 is conserved. However, the folding of fragment 2 is different from that of prothrombin kringle 1 and other kringles: the second outer loop possesses a distorted two-turn helix, and the hairpin β-turn of the second inner loop pivots at Val64 and Asp70 by 60°. Lys35 is located on a turn of the helix, which causes it to project into solvent space in the fragment 2-thrombin complex, thereby devastating any vestige of the cationic center of the lysine binding site. Since fragment 2 has not been reported to bind lysine, it most likely has a different inherent folding conformation for the second outer loop, as has also been observed to be the case with TPA K2 and the urokinase kringle. The movement of the Val64-Asp70 β-turn is most likely a conformational change accompanying complexation, which reveals a new heretofore unsuspected flexibility in kringles. The fragment 2-thrombin complex is only the second cassette module-catalytic domain structure to be determined for a multidomain blood protein and only the third domain-domain interaction to be described among such proteins, the others being factor Xa without a Gla domain and Ca2+ prothrombin fragment 1 with a Gla domain and a kringle. © 1993 American Chemical Society.

Specific and nonspecific collapse in protein folding funnels

Experiments with fast folding proteins are beginning to address the relationship between collapse and folding. We investigate how different scenarios for folding can arise depending on whether the folding and collapse transitions are concurrent or whether a nonspecific collapse precedes folding. Many earlier studies have focused on the limit in which collapse is fast compared to the folding time; in this work we focus on the opposite limit where, at the folding temperature, collapse and folding occur simultaneously. Real proteins exist in both of these limits. The folding mechanism varies substantially in these two regimes. In the regime of concurrent folding and collapse, nonspecific collapse now occurs at a temperature below the folding temperature (but slightly above the glass transition temperature).

Steric constraints as folding coadjuvant

Through the analyses of the Miyazawa-Jernigan matrix it has been shown that the hydrophobic effect generates the dominant driving force for protein folding. By using both lattice and off-lattice models, it is shown that hydrophobic-type potentials are indeed efficient in inducing the chain through nativelike configurations, but they fail to provide sufficient stability so as to keep the chain in the native state. However, through comparative Monte Carlo simulations, it is shown that hydrophobic potentials and steric constraints are two basic ingredients for the folding process. Specifically, it is shown that suitable pairwise steric constraints introduce strong changes on the configurational activity, whose main consequence is a huge increase in the overall stability condition of the native state; detailed analysis of the effects of steric constraints on the heat capacity and configurational activity are provided. The present results support the view that the folding problem of globular proteins can be approached as a process in which the mechanism to reach the native conformation and the requirements for the globule stability are uncoupled.

Visualization of Protein Folding Funnels in Lattice Models

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ab initio protein folding simulations using atomic burials as informational intermediates between sequence and structure

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Characterization of fold-related fractures in the carbonate rocks of the Cingoli anticline, northern Apennines, Italy

Thrust fault-related folds in carbonate rocks are characterized by deformation accommodated by different structures, such as joints, faults, pressure solution seams, and deformation bands. Defining the development of fracture systems related to the folding process is significant both for theoretical and practical purposes. Fracture systems are useful constrains in order to understand the kinematical evolution of the fold. Furthermore, understanding the relationships between folding and fracturing provides a noteworthy contribution for reconstructing the geodynamic and the structural evolution of the studied area. Moreover, as fold-related fractures influence fluid flow through rocks, fracture systems are relevant for energy production (geothermal studies, methane and CO2 , storage and hydrocarbon exploration), environmental and social issues (pollutant distribution, aquifer characterization). The PhD project shows results of a study carried out in a multilayer carbonate anticline characterized by different mechanical properties. The aim of this study is to understand the factors which influence the fracture formation and to define their temporal sequence during the folding process. The studied are is located in the Cingoli anticline (Northern Apennines), which is characterized by a pelagic multilayer characterized by sequences with different mechanical stratigraphies. A multi-scale analysis has been made in several outcrops located in different structural positions. This project shows that the conceptual sketches proposed in literature and the strain distribution models outline well the geometrical orientation of most of the set of fractures observed in the Cingoli anticline. On the other hand, the present work suggests the relevance of the mechanical stratigraphy in particular controlling the type of fractures formed (e.g. pressure solution seams, joints or shear fractures) and their subsequent evolution. Through a multi-scale analysis, and on the basis of the temporal relationship between fracture sets and their orientation respect layering, I also suggest a conceptual model for fracture systems formation.