Isolation, properties and spatial site analysis of gamma subunits of B-phycoerythrin and R-phycoerythrin

Polysiphonia urceolata R-phycoerythrin and Porphyridium cruentum B-phycoerythrin were degraded with proteinaseK, and then the nearly native gamma subunits were isolated from the reaction mixture. The process of degradation of phycoerythrin with proteinaseK showed that the gamma subunit is located in the central cavity of (alpha beta)(6) hexamer of phycoerythrin. Comparative analysis of the spectra of the native phycoerythrin, the phycoerythrin at pH 12 and the isolated gamma subunit showed that the absorption peaks of phycoerythrobilins on alpha or beta subunit are at 535 nm (or 545 nm) and 565 nm, the fluorescence emission maximum at 580 nm; the absorption peak of phycoerythrobilins on the isolated gamma subunit is at 589 nm, the fluorescence emission peak at 620 nm which overlaps the absorption maximum of C-phycocyanin and perhaps contributes to the energy transfer with high efficiency between phycoerythrin and phycocyanin in phycobilisome; the absorption maximum of phycourobilin on the isolated gamma subunit is at 498 nm, which is the same as that in native phycoerythrin, and the fluorescence emission maximum at 575 nm.

Molecular cloning and characterization of a catalase gene from Zhikong scallop Chlamys farreri

Catalase is one of the central enzymes involved in scavenging the high level of reactive oxygen species (ROS) by degradation of hydrogen peroxide to oxygen and water. The full-length catalase cDNA of Zhikong scallop Chlamys farreri (denoted as CfCAT) was identified from hemocytes by expressed sequence tag (EST) and rapid amplification of cDNA ends (RACE) approaches. The nucleotide sequence of CfCAT cDNA consisted of 3146 bp with a 5' UTR of 103 bp, an unusually long 31 UTR of 1519 bp with a canonical polyadenylation signal sequence AATAAA and a potyA tail, and an open reading frame (ORF) of 1521 bp encoding a polypeptide of 507 amino acids with predicted molecular weight of 57.5 kDa. The deduced amino acid sequence of CfCAT has significant homology to catalases from animals, plants and bacteria. Several highly conserved motifs including the proximal heme-ligand signature sequence RLFSYNDTH, the proximal active site signature FNRERIPERVVHAKGGGA, and the three catalytic amino acid residues of His(72), Asn(145) and Tyr(355) were identified in the deduced amino acid sequence of CfCAT. The CfCAT was demonstrated to be a peroxisomal glycoprotein with two potential glycosylation sites and a peroxisome targeting signal of ANL that was consistent with human, mouse and rat catalases. The time-course expression of CfCAT in hemocytes was measured by quantitative real-time PCR. The expression of CfCAT increased gradually and reached the highest point at 12 h post-Vibrio infection, then recovered to the original level at 24 h. All these results indicate that CfCAT, a constitutive and inducible protein, is a member of the catalase family and is involved in the process against ROS in scallop. (c) 2007 Published by Elsevier Ltd.

Three dimensional fluorescence excitation-emission on matrix spectrum of dissolved organic substance in marine microalgaes growth process

The three-dimensional fluorescence spectrum was used to detect the changes in dissolved organic substances from the cultured Skeletonema costatum, Alexandrium tamarense, Alexandrium mimutum, Scrippsiella trochodea, Prorocentrum donghaiense and Prorocentrum micans. The result indicates that all of the microalgaes can produce FDOM in the growth courses. Diatom such as Skeletonema costatum can produce humic-like FDOM. However dinoflagellate can produce protein-like FDOM at exponential growth phase. When the algae grows into decadency phase, the intensity of humic-like and protein-like fluorescence augments rapidly, which may be due to a mass of FDOM realeased by the old or dead cell fragmentation and the degradation of bacteria by using non-FDOM. The fluorescent intensity of Alexandrium tamarense, Alexandrium mimutum, Prorocentrum donghaiense and Prorocentrum micans can reduce at anaphase of decadency phase because of the degradation of bacteria and light. The same genus of algae can produce similar FDOM, for example: Alexandrium tamarense, and Alexandrium mimutum, Prorocentrum donghaiense and Prorocentrum micans, but the positions of the fluorescence peaks are different.

Salt-assisted acid hydrolysis of chitosan to oligomers under microwave irradiation

The effect of inorganic salts such as sodium chloride on the hydrolysis of chitosan in a microwave field was investigated. While it is known that microwave heating is a convenient way to obtain a wide range of products of different molecular weights only by changing the reaction time and/or the radiation power, the addition of some inorganic salts was shown to effectively accelerate the degradation of chitosan under microwave irradiation. The molecular weight of the degraded chitosan obtained by microwave irradiation was considerably lower than that obtained by traditional heating. Moreover, the molecular weight of degraded chitosan obtained by microwave irradiation assisted under the conditions of added salt was considerably lower than that obtained by microwave irradiation without added salt. Furthermore, the effect of ionic strength of the added salts was not linked with the change of molecular weight. FTIR spectral analyses demonstrated that a significantly shorter time was required to obtain a satisfactory molecular weight by the microwave irradiation-assisted inorganic salt method than by microwave irradiation without inorganic salts and conventional technology. (C) 2005 Elsevier Ltd. All rights reserved.

Preparation of low-molecular-weight and high-sulfate-content chitosans under microwave radiation and their potential antioxidant activity in vitro

In the present paper microwave radiation has been used to introduce N-sulfo and O-sulfo groups into chitosan with a thigh degree of substitution and low-molecular weight. The sulfation of chitosan was performed in microwave ovens. It was found that microwave heating is a convenient way to obtain a wide range of products of different degrees of substitution and molecular weight only by changing reaction time or/and radiation power. Moreover, microwave radiation accelerated the degradation of sulfated chitosan, and the molecular weight of sulfated chitosan was considerably lower than that obtained by traditional heating. There are no differences in the chemical structure of sulfated chitosan obtained by microwave and by conventional technology. FTIR and C-13 NMR spectral analyses demonstrated that a significantly shorter time is required to obtain a satisfactory degree of substitution and molecular weight by microwave radiation than by conventional technology. In this present paper, we also determined antioxidant activity of low-molecular-weight and high-sulfate-content chitosans (LCTS). The results showed LCTS could scavenge superoxide and hydroxyl radical. Its IC50 is 0.025 and 1.32mg/mL, respectively. It is a potential antioxidant in vitro. (C) 2004 Published by Elsevier Ltd.

Livestock grazing, plateau pikas and the conservation of avian biodiversity on the Tibetan plateau

On the Qinghai-Tibet plateau increased livestock numbers have resulted in degradation of the grasslands with potential impacts on native biodiversity. Concurrently, perceived increases in populations of native small mammals such as plateau pikas (Ochotona curzoniae) have led to poisoning programs, with uncertain impacts on species such as ground-nesting birds. We explored the relationships between the local seasonal abundance of small birds and (1) the density of pika burrows; (2) livestock grazing practices; and (3) local poisoning of pikas. Around Naqu prefecture, central Tibet, we used a nested experimental design to collect data from areas rested from grazing over summer, nearby areas with year-round grazing and areas subjected to pika poisoning. Additional data were collected from a site where grazing had not occurred for at least 4 years prior to the study. Poisoning pikas in spring had no detectable effect on the local abundance of birds the following autumn. However, two ground-nesting species, white-rumped and rufous-necked snowfinches, showed positive associations with the density of pika burrows, indicating that long-term 'pika poisoning could reduce the density of these species by reducing the density of pika burrows. Rufous-necked snowfinches and non ground-nesting species including horned larks and common hoopoes showed positive responses to reduced grazing pressure from livestock, particularly in the long-rested site, indicating current grazing levels could be having a negative impact on these species. Conservation of small passerine biodiversity in this system will require changed management practices for livestock and pikas that consider the complex three-way interaction between livestock grazing, pikas and small birds. (C) 2008 Elsevier Ltd. All rights reserved.

Influence of different neuroprotective drugs on dopamine neurotoxicity induced by 3,4-methylenedioxymethamphetamine and MPTP in mice

Introduction: Parkinson‟s disease (PD) is characterized by a chronic progressive loss of nigrostriatal dopaminergic neurons that is associated with chronic neuroinflammation. Current treatments for PD can significantly improve symptoms but do not cure the disease or slow its progression. An approach used in existing therapies is based on the inhibition of monoamine oxidase (MAO), enzyme involved in the metabolic degradation of dopamine. Although, preclinical studies showed that MAO-B inhibitors have neuroprotective activity in cellular and animal models of PD, clinical trials did not completely confirm this result. Therefore a large number of new molecules, with more potent MAO-B inhibitory activity and a possible neuroprotective effect, have been proposed to replace the pre-existing MAO-B inhibitors. The profile of the recent MAO inhibitor, SZV558, appears to be particularly interesting because of its pharmacodynamic, favorable for disease-modifying properties and its irreversible MAO-B enzyme bind. The enhancement of adult neurogenesis could be of great clinical interest in the management of neurodegenerative disorders. In line with this, the metformin, a well-known antidiabetic drug, has recently been proposed to promote neurogenesis and to have a neuroprotective effect on the neurodegenerative processes induced by the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in a mice PD model. Although, PD has multiple origins, one hypothesis is that amphetamine-related drugs may be part of the wide array of factors leading to the dopaminergic neuron degeneration that causes the disease. These hypothesis are supported by different results that showed a persistent, long-term dopaminergic toxicity induced by 3,4-methylenedioxymethamphetamine (MDMA) in mice. Moreover, the MDMA, altering the dopaminergic transmission, may affect neurogenesis and synaptogenesis. On these basis, considering that the young brain is particularly sensitive to drug-induced neurotoxicity, the consumption of MDMA during the adolescence might increase the vulnerability of dopaminergic neurons. However, the use of amphetamine-related drugs by adolescent and young people is often combined with caffeinated energy drinks in order to amplify their stimulant actions. Although caffeine use is safe, the combined treatment of caffeine and MDMA increases not only the DA release but also the microglia and astroglia activation. Aims: During my Ph.D. I studied the influence of neuroprotective drugs, such as MAO inhibitors and metformin, or substances, such as caffeine, on the neurodegenerative effects of two dopaminergic toxins, MDMA and MPTP, in mice. 1. In the first phase of my study, I evaluated the neuroprotective activity of the new MAO-B inhibitor SZV558, compared with well-known rasagiline, in a chronic mouse model of MPTP plus probenecid (MPTPp), which induces a progressive loss of nigrostriatal dopaminergic neurons. 2. Previous results showed that when MDMA is associated with caffeine, a more pronounced degeneration in adolescent compared with adult mice was observed. To better clarify the molecular mechanism at the base of the different neurotoxic effect of this drug association at different ages, I evaluated the neuronal nitric oxide synthase (nNOS) expression, which plays a critical role in the integration of dopaminergic and glutamatergic transmissions, in the CPu of adolescent or adult mice treated with MDMA, alone or in combination with caffeine. 3. Finally, I investigated the neuroprotective effect of metformin against dopaminergic neurotoxicity induced by MDMA in the CPu and SNc of adult mice. Conclusions: These results demonstrated that the dopaminergic neurodegenerative process may be induced or conditioned by environment stressors or substances which influence, through different ways, the development of neurodegenerative mechanisms. In the present study I evaluated the effects of 3 substances, known as potentially neuroprotective, in combination with two different neurotoxins that affect the nigrostriatal dopaminergic system. The SZV558 MAO-B inhibitor and the metformin protected the nigrostriatal pathway, usually affected in PD, by MPTP- and MDMA- induced neurotoxicity, respectively. On the other hand, caffeine, administrated with MDMA, showed a neurotoxic potential depending on the age of consumers, confirming the vulnerability of adolescent brain to consumption of drug and substances that affected the dopaminergic system. In conclusion, the study of neurodegenerative processes may be relevant to understand the human pharmacology, the origin and development of neurodegenerative disease and to predict the neurotoxic effect of drug abuse.

Using a logical model to predict the growth of yeast

Whelan, K. E. and King, R. D. Using a logical model to predict the growth of yeast. BMC Bioinformatics 2008, 9:97

Evidence in support of a role for plant-mediated proteolysis in the rumens of grazing animals

Kingston-Smith, A. H., Merry, R. J., Leemans, D. K., Thomas, Howard, Theodorou, M. K. (2005). Evidence in support of a role for plant-mediated proteolysis in the rumens of grazing animals. British Journal of Nutrition, 93(1), 73-79. Sponsorship: DEFRA / BBSRC RAE2008

Nonlinear Acoustics in Underwater and Biomedical Applications: Array Performance Degradation and Time Reversal Invariance

This dissertation describes a model for acoustic propagation in inhomogeneous flu- ids, and explores the focusing by arrays onto targets under various conditions. The work explores the use of arrays, in particular the time reversal array, for underwater and biomedical applications. Aspects of propagation and phasing which can lead to reduced focusing effectiveness are described. An acoustic wave equation was derived for the propagation of finite-amplitude waves in lossy time-varying inhomogeneous fluid media. The equation was solved numerically in both Cartesian and cylindrical geometries using the finite-difference time-domain (FDTD) method. It was found that time reversal arrays are sensitive to several debilitating factors. Focusing ability was determined to be adequate in the presence of temporal jitter in the time reversed signal only up to about one-sixth of a period. Thermoviscous absorption also had a debilitating effect on focal pressure for both linear and nonlinear propagation. It was also found that nonlinearity leads to degradation of focal pressure through amplification of the received signal at the array, and enhanced absorption in the shocked waveforms. This dissertation also examined the heating effects of focused ultrasound in a tissue-like medium. The application considered is therapeutic heating for hyperther- mia. The acoustic model and a thermal model for tissue were coupled to solve for transient and steady temperature profiles in tissue-like media. The Pennes bioheat equation was solved using the FDTD method to calculate the temperature fields in tissue-like media from focused acoustic sources. It was found that the temperature-dependence of the medium's background prop- erties can play an important role in the temperature predictions. Finite-amplitude effects contributed excess heat when source conditions were provided for nonlinear ef- fects to manifest themselves. The effect of medium heterogeneity was also found to be important in redistributing the acoustic and temperature fields, creating regions with hotter and colder temperatures than the mean by local scattering and lensing action. These temperature excursions from the mean were found to increase monotonically with increasing contrast in the medium's properties.

Safe Composition of Web Communication Protocols for Extensible Edge Services

As new multi-party edge services are deployed on the Internet, application-layer protocols with complex communication models and event dependencies are increasingly being specified and adopted. To ensure that such protocols (and compositions thereof with existing protocols) do not result in undesirable behaviors (e.g., livelocks) there needs to be a methodology for the automated checking of the "safety" of these protocols. In this paper, we present ingredients of such a methodology. Specifically, we show how SPIN, a tool from the formal systems verification community, can be used to quickly identify problematic behaviors of application-layer protocols with non-trivial communication models—such as HTTP with the addition of the "100 Continue" mechanism. As a case study, we examine several versions of the specification for the Continue mechanism; our experiments mechanically uncovered multi-version interoperability problems, including some which motivated revisions of HTTP/1.1 and some which persist even with the current version of the protocol. One such problem resembles a classic degradation-of-service attack, but can arise between well-meaning peers. We also discuss how the methods we employ can be used to make explicit the requirements for hardening a protocol's implementation against potentially malicious peers, and for verifying an implementation's interoperability with the full range of allowable peer behaviors.

Impact of non-optimal grounding of the CC2420 RFIC on a 802.15.4 Tyndall sensor wireless mote

The performance of an RF output matching network is dependent on integrity of the ground connection. If this connection is compromised in anyway, additional parasitic elements may occur that can degrade performance and yield unreliable results. Traditionally, designers measure Constant Wave (CW) power to determine that the RF chain is performing optimally, the device is properly matched and by implication grounded. It is shown that there are situations where modulation quality can be compromised due to poor grounding that is not apparent using CW power measurements alone. The consequence of this is reduced throughput, range and reliability. Measurements are presented on a Tyndall Mote using a CC2420 RFIC todemonstrate how poor solder contact between the ground contacts and the ground layer of the PCB can lead tothe degradation of modulated performance. Detailed evaluation that required the development of a new measurement definition for 802.15.4 and analysis is presented to show how waveform quality is affected while the modulated output power remains within acceptable limits.

Systematic delay-driven power optimisation and power-driven delay optimisation of combinational circuits

With the proliferation of mobile wireless communication and embedded systems, the energy efficiency becomes a major design constraint. The dissipated energy is often referred as the product of power dissipation and the input-output delay. Most of electronic design automation techniques focus on optimising only one of these parameters either power or delay. Industry standard design flows integrate systematic methods of optimising either area or timing while for power consumption optimisation one often employs heuristics which are characteristic to a specific design. In this work we answer three questions in our quest to provide a systematic approach to joint power and delay Optimisation. The first question of our research is: How to build a design flow which incorporates academic and industry standard design flows for power optimisation? To address this question, we use a reference design flow provided by Synopsys and integrate in this flow academic tools and methodologies. The proposed design flow is used as a platform for analysing some novel algorithms and methodologies for optimisation in the context of digital circuits. The second question we answer is: Is possible to apply a systematic approach for power optimisation in the context of combinational digital circuits? The starting point is a selection of a suitable data structure which can easily incorporate information about delay, power, area and which then allows optimisation algorithms to be applied. In particular we address the implications of a systematic power optimisation methodologies and the potential degradation of other (often conflicting) parameters such as area or the delay of implementation. Finally, the third question which this thesis attempts to answer is: Is there a systematic approach for multi-objective optimisation of delay and power? A delay-driven power and power-driven delay optimisation is proposed in order to have balanced delay and power values. This implies that each power optimisation step is not only constrained by the decrease in power but also the increase in delay. Similarly, each delay optimisation step is not only governed with the decrease in delay but also the increase in power. The goal is to obtain multi-objective optimisation of digital circuits where the two conflicting objectives are power and delay. The logic synthesis and optimisation methodology is based on AND-Inverter Graphs (AIGs) which represent the functionality of the circuit. The switching activities and arrival times of circuit nodes are annotated onto an AND-Inverter Graph under the zero and a non-zero-delay model. We introduce then several reordering rules which are applied on the AIG nodes to minimise switching power or longest path delay of the circuit at the pre-technology mapping level. The academic Electronic Design Automation (EDA) tool ABC is used for the manipulation of AND-Inverter Graphs. We have implemented various combinatorial optimisation algorithms often used in Electronic Design Automation such as Simulated Annealing and Uniform Cost Search Algorithm. Simulated Annealing (SMA) is a probabilistic meta heuristic for the global optimization problem of locating a good approximation to the global optimum of a given function in a large search space. We used SMA to probabilistically decide between moving from one optimised solution to another such that the dynamic power is optimised under given delay constraints and the delay is optimised under given power constraints. A good approximation to the global optimum solution of energy constraint is obtained. Uniform Cost Search (UCS) is a tree search algorithm used for traversing or searching a weighted tree, tree structure, or graph. We have used Uniform Cost Search Algorithm to search within the AIG network, a specific AIG node order for the reordering rules application. After the reordering rules application, the AIG network is mapped to an AIG netlist using specific library cells. Our approach combines network re-structuring, AIG nodes reordering, dynamic power and longest path delay estimation and optimisation and finally technology mapping to an AIG netlist. A set of MCNC Benchmark circuits and large combinational circuits up to 100,000 gates have been used to validate our methodology. Comparisons for power and delay optimisation are made with the best synthesis scripts used in ABC. Reduction of 23% in power and 15% in delay with minimal overhead is achieved, compared to the best known ABC results. Also, our approach is also implemented on a number of processors with combinational and sequential components and significant savings are achieved.

VCP/p97 is essential for maturation of ubiquitin-containing autophagosomes and this function is impaired by mutations that cause IBMPFD.

VCP (VCP/p97) is a ubiquitously expressed member of the AAA(+)-ATPase family of chaperone-like proteins that regulates numerous cellular processes including chromatin decondensation, homotypic membrane fusion and ubiquitin-dependent protein degradation by the proteasome. Mutations in VCP cause a multisystem degenerative disease consisting of inclusion body myopathy, Paget disease of bone, and frontotemporal dementia (IBMPFD). Here we show that VCP is essential for autophagosome maturation. We generated cells stably expressing dual-tagged LC3 (mCherry-EGFP-LC3) which permit monitoring of autophagosome maturation. We determined that VCP deficiency by RNAi-mediated knockdown or overexpression of dominant-negative VCP results in significant accumulation of immature autophagic vesicles, some of which are abnormally large, acidified and exhibit cathepsin B activity. Furthermore, expression of disease-associated VCP mutants (R155H and A232E) also causes this autophagy defect. VCP was found to be essential to autophagosome maturation under basal conditions and in cells challenged by proteasome inhibition, but not in cells challenged by starvation, suggesting that VCP might be selectively required for autophagic degradation of ubiquitinated substrates. Indeed, a high percentage of the accumulated autophagic vesicles contain ubiquitin-positive contents, a feature that is not observed in autophagic vesicles that accumulate following starvation or treatment with Bafilomycin A. Finally, we show accumulation of numerous, large LAMP-1 and LAMP-2-positive vacuoles and accumulation of LC3-II in myoblasts derived from patients with IBMPFD. We conclude that VCP is essential for maturation of ubiquitin-containing autophagosomes and that defect in this function may contribute to IBMPFD pathogenesis.

The prevalence and regulation of antisense transcripts in Schizosaccharomyces pombe.

A strand-specific transcriptome sequencing strategy, directional ligation sequencing or DeLi-seq, was employed to profile antisense transcriptome of Schizosaccharomyces pombe. Under both normal and heat shock conditions, we found that polyadenylated antisense transcripts are broadly expressed while distinct expression patterns were observed for protein-coding and non-coding loci. Dominant antisense expression is enriched in protein-coding genes involved in meiosis or stress response pathways. Detailed analyses further suggest that antisense transcripts are independently regulated with respect to their sense transcripts, and diverse mechanisms might be potentially involved in the biogenesis and degradation of antisense RNAs. Taken together, antisense transcription may have profound impacts on global gene regulation in S. pombe.