Both DNA gyrase and reverse gyrase are present in the hyperthermophilic bacterium Thermotoga maritima

Like all hyperthermophiles yet tested, the bacterium Thermotoga maritima contains a reverse gyrase. Here we show that it contains also a DNA gyrase. The genes top2A and top2B encoding the two subunits of a DNA gyrase-like enzyme have been cloned and sequenced. The Top2A (type II DNA topoisomerase A protein) is more similar to GyrA (DNA gyrase A protein) than to ParC [topoisomerase IV (Topo IV) C protein]. The difference is especially striking at the C-terminal domain, which differentiates DNA gyrases from Topo IV. DNA gyrase activity was detected in T. maritima and purified to homogeneity using a novobiocin-Sepharose column. This hyperhermophilic DNA gyrase has an optimal activity around 82–86°C. In contrast to plasmids from hyperthermophilic archaea, which are from relaxed to positively supercoiled, we found that the plasmid pRQ7 from Thermotoga sp. RQ7 is negatively supercoiled. pRQ7 became positively supercoiled after addition of novobiocin to cell cultures, indicating that its negative supercoiling is due to the DNA gyrase of the host strain. The findings concerning DNA gyrase and negative supercoiling in Thermotogales put into question the role of reverse gyrase in hyperthermophiles.

A two-subunit type I DNA topoisomerase (reverse gyrase) from an extreme hyperthermophile.

A recently described reverse gyrase from the hyperthermophilic methanogen Methanopyrus kandleri is the only known example of a heterodimeric type I topoisomerase. The enzyme is made up of a 42-kDa subunit which covalently interacts with DNA (RgyA) and a 138-kDa subunit which binds ATP (RgyB). We have now cloned and sequenced the genes for both subunits of this enzyme. Surprisingly, the universally conserved type I topoisomerase domain [Lima, C. D., Wang, J. C. & Mondragon, A. (1994) Nature (London) 367, 138-146] which has been found as a contiguous polypeptide in the prokaryotes and eukaryotes is shared between the protomers. The subdomain with the active-site tyrosine is entirely within RgyA, whereas the subdomain implicated in noncovalent binding of the cleaved DNA strand is contained entirely in RgyB. The appearance of this unique structure in a highly conserved enzyme family supports the hypothesis that the methanogens branched from other prokaryotes and eukaryotes very early in evolution.

Analyse de la corrélation conditionnelle dérivée de la coévolution d’un système de trois gènes par un modèle du maximum de vraisemblance

Les gènes codant pour des protéines peuvent souvent être regroupés et intégrés en modules fonctionnels par rapport à un organelle. Ces modules peuvent avoir des composantes qui suivent une évolution corrélée pouvant être conditionnelle à un phénotype donné. Les gènes liés à la motilité possèdent cette caractéristique, car ils se suivent en cascade en réponse à des stimuli extérieurs. L’hyperthermophilie, d’autre part, est interreliée à la reverse gyrase, cependant aucun autre élément qui pourrait y être associé avec certitude n’est connu. Ceci peut être dû à un déplacement de gènes non orthologues encore non résolu. En utilisant une approche bio-informatique, une modélisation mathématique d’évolution conditionnelle corrélée pour trois gènes a été développée et appliquée sur des profils phylétiques d’archaea. Ceci a permis d’établir des théories quant à la fonction potentielle du gène du flagelle FlaD/E ainsi que l’histoire évolutive des gènes lui étant liés et ayant contribué à sa formation. De plus, une histoire évolutive théorique a été établie pour une ligase liée à l’hyperthermophilie.

Reverse immigration

This book (256 pages, written in Korean) is a critical essay that reviews, questions, and criticises Korean and Eastern immigrants’ thinking and behaviour styles in Australia from their cultural perspectives, and discuss and proposes a creative cultural dimension for their better life in a multicultural context. Multiculturalism is not supportive of Eastern cultures because of individualistic collection of cultures, while transculturalism facilitates nurture of their culture in a community-oriented way within multicultural circumstances. Korean and Eastern immigrants, sharing oriental cultural systems and values, should approach to the Australian multicultural context with transculturalism which allows creating new cultural values in collaboration with and by participation into local communities. ------------------------------------------------------------ Many Eastern immigrants live in their own ethnic communities without or less interacting with Australian (communities). The author defines this phenomenon as “reverse immigration”. Reverse immigration refers to re-immigrating to their ethnic community in Australia or to their birth country despite they did not anticipate that this would happen to them before immigration to Australia. The author argues that Easterners’ collectivistic culture often devalues individuality and vice versa. Cultural clash between West and East often forces the immigrants to choose reverse immigration because of their lack of understanding of Western culture and their cultural characteristics such as low individuality, high power distance, and high uncertainty avoidance. For example, a vague boundary between individualist and collectivist in a collectivistic context (within their ethnic group) often leads to maladjustment to local communities and enhancement of cultural conservatism. The author proposes that the cultural clash can be overcome by cross-cultural activities named “transculturalism”. To Eastern immigrants, transculturalism can be achieved by acculturation of their two predominant cultures, the third-person perspective and generalised others. In a multicultural context, the former refers to the ability to share another person's feelings and emotions as if they were your own, and the latter does the ability to manage community and public expectations. When both cultural values are used for quality interactions between East and West, they allow Eastern immigrants to be more creative and critical and Australian to be more socially inclusive and culturally tolerant. With these discussions, the author discusses cultural differences throughout the book with four topics (chapters) and proposes transculturalism as a solution to the reverse immigration. ------------------------------------------------------------ Chapter 1 criticises Koreans’ attitudes and methods towards learning English that is less pragmatic and practical, but more likely to be a scholarly study. The author explains that Koreans’ non-pragmatic towards learning English has been firmly built based on their traditional systems and values that Koreans view English as a discipline and an aim of academic achievements rather than a means of communication. Within their cultural context, English can be perceived as more than a language, but something like vastly superior to their language and culture. Their collectivistic culture regards English as an unreachable and heterogeneous one that may threaten their cultural identity, so that “scholarly studying” is only the way to achieve (not learn) it. This discourages the immigrants to engage and involve in daily dialogues by “using” English as a second language. The author further advises the readers to be aware of Eastern collectivistic culture in communication and interaction that sometimes completely reverses private and public topics in a Western context. This leads them to feel that they have no content to talk to natives. ------------------------------------------------------------ Chapter 2 compares between Korea and Australia in terms of their educational systems and values, and proposes how Eastern overseas students can achieve critical and creative thinking within a Western educational setting. Interestingly, this chapter includes an explanation of why Eastern overseas students easily fail assessments including essay writing, oral presentations and discussions. One of the reasons the author explains is that Eastern students are not familiar to criticise others and think creatively, especially when they recognise that their words and ideas may harm the collectivistic harmony. Western educational systems focuses on enhancement of individuality such as self-confidence, self-esteem, and self-expression, while Eastern educational systems foster group-oriented values such as interpersonal relationship, and strong moral and spiritual values. Yet, the author argues that the collectivistic approach to criticism and creativity is often more critical and creative than Western individuals when they know what they are supposed to do for a group (or a community). Therefore, Eastern students need to think their cultural merits and demerits by using an individual perspective rather than generalised others’ perspective. The latter often discourages individual participation in a community, and the generalised others in a Western culture is weaker than Eastern. Furthermore, Western educational systems do not educate students to transform (loose) their individuality to fit into a group or a community. Rather they cultivate individuality for community prosperity. ------------------------------------------------------------ Chapter 3 introduces various cases of reverse immigration in workplaces that many immigrants return to their country or their ethnic community after many trials for acculturation. Reverse immigration is unexpected and not planned before immigration, so that its emotional embarrassment increases such severe social loneliness. Most Eastern immigrant workers have tried to adjust themselves in this new cultural environment at the early stages of immigration. However, their cultural features of collectivism, high power distance, high uncertainty avoidance, and long-term oriented cultures suppress individual initiative and eliminate the space for experiments in ways of acculturation. The author argues that returning to their ethnic community (physically and psychologically) leads to two significant problems: their distorted parenting and becoming more conservatives. The former leads the first generation of immigrants to pressure their children to pursue extrinsic or materialist values, such as financial success, fame and physical appearance, rather than on intrinsic values, while the latter refers to their isolated conservative characters because of their remoteness from the changes of their own country. The author also warns that their ethnic and religious groups actively strengthens immigrants’ social loneliness and systematically discourages immigrants’ interests and desire to be involving into local communities. The ethnic communities and leaders have not been interacting with Australian local communities and, as a result, are eager to conserve outdated cultural systems values. Even they have a tendency to weed out those people who wish to settle down within Australian local communities. They believe that those people can threaten their community’s survival and continuity. ------------------------------------------------------------ Chapter 4 titled multiculturalism argues that Korean and Eastern immigrants should more precisely understand Australia as a multicultural society in a way of collaboratively creating new cultural values. The author introduces multiculturalism with its definitions and history in Australia and argues the limitations of multiculturalism from an Easterner’s perspective. With well known tragedies of the second generations of U.S. immigrants, Cho Seung-Hui, a university student, massacred 32 people on the Virginia Tech before committing suicide and Hidal Hassan, an Army psychiatrist, killed 13 people at Fort Hood and the responses of ethnic community, the author explains that their mental illness may be derived from their parents’ (or ethnic group) culturally isolated attitude and socially static viewpoint of U.S. (Western system and values). The author insists that multiculturalism may restrict Eastern immigrants’ engagement and involvement in local communities. Multiculturalism has been systematically and historically developed based on Western systems and cultural values. In other words, multiculturalism requires high self-confidence and self-esteem that Eastern immigrants less prioritise them. It has been generally known that Easterners put more weight on human relationship than Westerners, but the author claims that this is not true. Within an individualistic culture, Westerners are more interested in building person-to-person connections and relationships. While Easterners are more interested in how individuals can achieve a sense of belonging within a group and a community. Therefore, multiculturalism is an ideology which forces Eastern immigrants to discard their strong desire to be part of a group and does not give a sense of belonging. In a consequence, the author advises that Eastern immigrants should aim towards “transculturalism” which allows them to actively participate in and contribute to their multicultural community. Transculturalism does not ask Easterners to discard their cultural values, but enables them to be a collectivistic individualist (a community leader) who is capable of developing new cultural values in a more creative and productive way. Furthermore, transculturalism encourages Western Australians in a multicultural context to collaborate with ethnic minorities to build a better community.

Reverse engineering and finite element analysis of a 3-wheeler automotive structure

Auto rickshaws (3-wheelers) are the most sought after transport among the urban and rural poor in India. The assembly of the vehicle involves assemblies of several major components. The L-angle is the component that connects the front panel with the vehicle floor. Current L-angle part has been observed to experience permanent deformation failure over period of time. This paper studies the effect of the addition of stiffeners on the L-angle to increase the strength of the component. A physical model of the L-angle was reversed engineered and modelled in CAD before static loading analysis were carried out on the model using finite element analysis. The modified L-angle fitted with stiffeners was shown to be able to withstand more load compare to previous design.

Removal of toxic arsenic and antimony from groundwater Spiro Tunnel Bulkhead in Park City Utah using colloidal iron hydroxide : comparison with reverse ssmosis

Verification testing of two model technologies in pilot scale to remove arsenic and antimony based on reverse osmosis and chemical coagulation/filtration systems was conducted in Spiro Tunnel Water Filtration Plant located in Park City, Utah, US. The source water was groundwater in abandoned silver mine, naturally contaminated by 60-80 ppb of arsenic and antimony below 10 ppb. This water represents one of the sources of drinking water for Park City and constitutes about 44% of the water supply. The failure to remove antimony efficiently by coagulation/filtration (only 4.4% removal rate) under design conditions is discussed in terms of the chemistry differences between Sb (III, V) and As (III, V). Removal of Sb(V) at pH > 7, using coagulation/filtration technology, requires much higher (50 to 80 times) concentration of iron (III) than As. The stronger adsorption of arsenate over a wider pH range can be explained by the fact that arsenic acid is tri-protic, whereas antimonic acid is monoprotic. This difference in properties of As(V) and Sb(V) makes antimony (V) more difficult to be efficiently removed in low concentrations of iron hydroxide and alkaline pH waters, especially in concentration of Sb < 10 ppb.

Reverse biased Pt/nanostructured MoO3/SiC Schottky diode based hydrogen gas sensors

Pt/nanostructured molybdenum oxide (MoO3) /SiC Schottky diode based gas sensors were fabricated for hydrogen (H2) gas sensing. Due to the enhanced performance, which is ascribed to the application of MoO3 nanostructures, these devices were used in reversed bias. MoO3 characterization by scanning electron microscopy showed morphology of randomly orientated nanoplatelets with thicknesses between 50 and 500 nm. An α-Β mixed phase crystallographic structure of MoO3 was characterized by x-ray diffraction. At 180 °C, 1.343 V voltage shift in the reverse I-V curve and a Pt/ MoO3 barrier height change of 20 meV were obtained after exposure to 1% H2 gas in synthetic air. © 2009 American Institute of Physics.

Conceptualisation of articular cartilage as a giant reverse micelle: A hypothetical mechanism for joint biocushioning and lubrication

Phospholipid (PL) molecules form the main structure of the membrane that prevents the direct contact of opposing articular cartilage layers. In this paper we conceptualise articular cartilage as a giant reverse micelle (GRM) in which the highly hydrated three-dimensional network of phospholipids is electrically charged and able to resist compressive forces during joint movement, and hence loading. Using this hypothetical base, we describe a hydrophilic-hydrophilic (HL-HL) biopair model of joint lubrication by contacting cartilages, whose mechanism is reliant on lamellar cushioning. To demonstrate the viability of our concept, the electrokinetic properties of the membranous layer on the articular surface were determined by measuring via microelectrophoresis, the adsorption of ions H, OH, Na and Cl on phospholipid membrane of liposomes, leading to the calculation of the effective surface charge density. The surface charge density was found to be -0.08 ± 0.002 cm-2 (mean ± S.D.) for phospholipid membranes, in 0.155 M NaCl solution and physiological pH. This value was approximately five times less than that measured in 0.01 M NaCl. The addition of synovial fluid (SF) to the 0.155 M NaCl solution reduced the surface charge density by 30% which was attributed to the binding of synovial fluid macromolecules to the phospholipid membrane. Our experiments show that particles charge and interact strongly with the polar core of RM. We demonstrate that particles can have strong electrostatic interactions when ions and macromolecules are solubilized by reverse micelle (RM). Since ions are solubilized by reverse micelle, the surface entropy influences the change in the charge density of the phospholipid membrane on cartilage surfaces. Reverse micelles stabilize ions maintaining equilibrium, their surface charges contribute to the stability of particles, while providing additional screening for electrostatic processes. © 2008 Elsevier Ireland Ltd. All rights reserved.

Identification of early-stage colorectal cancer patients at risk of relapse post-resection by immunobead reverse transcription-PCR analysis of peritoneal lavage fluid for malignant cells

Purpose: Colorectal cancer patients diagnosed with stage I or II disease are not routinely offered adjuvant chemotherapy following resection of the primary tumor. However, up to 10% of stage I and 30% of stage II patients relapse within 5 years of surgery from recurrent or metastatic disease. The aim of this study was to determine if tumor-associated markers could detect disseminated malignant cells and so identify a subgroup of patients with early-stage colorectal cancer that were at risk of relapse. Experimental Design: We recruited consecutive patients undergoing curative resection for early-stage colorectal cancer. Immunobead reverse transcription-PCR of five tumor-associated markers (carcinoembryonic antigen, laminin γ2, ephrin B4, matrilysin, and cytokeratin 20) was used to detect the presence of colon tumor cells in peripheral blood and within the peritoneal cavity of colon cancer patients perioperatively. Clinicopathologic variables were tested for their effect on survival outcomes in univariate analyses using the Kaplan-Meier method. A multivariate Cox proportional hazards regression analysis was done to determine whether detection of tumor cells was an independent prognostic marker for disease relapse. Results: Overall, 41 of 125 (32.8%) early-stage patients were positive for disseminated tumor cells. Patients who were marker positive for disseminated cells in post-resection lavage samples showed a significantly poorer prognosis (hazard ratio, 6.2; 95% confidence interval, 1.9-19.6; P = 0.002), and this was independent of other risk factors. Conclusion: The markers used in this study identified a subgroup of early-stage patients at increased risk of relapse post-resection for primary colorectal cancer. This method may be considered as a new diagnostic tool to improve the staging and management of colorectal cancer. © 2006 American Association for Cancer Research.

Inhibitors of human immunodeficiency virus type 1 reverse transcriptase target distinct phases of early reverse transcription

Early HIV-1 reverse transcription can be separated into initiation and elongation phases. Here we show, using PCR analysis of negative-strand strong-stop DNA [(−)ssDNA] synthesis in intact virus, that different reverse transcriptase (RT) inhibitors affect distinct phases of early natural endogenous reverse transcription (NERT). The effects of nevirapine on NERT were consistent with a mechanism of action including both specific and nonspecific binding events. The nonspecific component of this inhibition targeted the elongation reaction, whereas the specific effect seemed principally to be directed at very early events (initiation or the initiation-elongation switch). In contrast, foscarnet and the nucleoside analog ddATP inhibited both early and late (−)ssDNA synthesis in a similar manner. We also examined compounds that targeted other viral proteins and found that Ro24-7429 (a Tat antagonist) and rosmarinic acid (an integrase inhibitor) also directly inhibited RT. Our results indicate that NERT can be used to identify and evaluate compounds that directly target the reverse transcription complex.