Inhibition of archaeal growth and DNA topoisomerase VI activities by the Hsp90 inhibitor radicicol

Type II DNA topoisomerases have been classified into two families, Topo IIA and Topo IIB, based on structural and mechanistic dissimilarities. Topo IIA is the target of many important antibiotics and antitumoural drugs, most of them being inactive on Topo IIB. The effects and mode of action of Topo IIA inhibitors in vitro and in vivo have been extensively studied for the last twenty-five years. In contrast, studies of Topo IIB inhibitors were lacking. To document this field, we have studied two Hsp90 inhibitors (radicicol and geldanamycin), known to interact with the ATP-binding site of Hsp90 (the Bergerat fold), which is also present in Topo IIB. Here, we report that radicicol inhibits the decatenation and relaxation activities of Sulfolobus shibatae DNA topoisomerase VI (a Topo IIB) while geldanamycin does not. In addition, radicicol has no effect on the Topo IIA Escherichia coli DNA gyrase. In agreement with their different effects on DNA topoisomerase VI, we found that radicicol can theoretically fit in the ATP-binding pocket of the DNA topoisomerase VI ‘Bergerat fold’, whereas geldanamycin cannot. Radicicol inhibited growths of Sulfolobus acidocaldarius (a crenarchaeon) and of Haloferax volcanii (a euryarchaeon) at the same doses that inhibited DNA topoisomerase VI in vitro. In contrast, the bacteria E.coli was resistant to this drug. Radicicol thus appears to be a very promising compound to study the mechanism of Topo IIB in vitro, as well as the biological roles of these enzymes in vivo

Detection of changes in gene regulatory patterns, elicited by perturbations of the Hsp90 molecular chaperone complex, by visualizing multiple experiments with an animation

<p>Abstract</p> <p>Background</p> <p>To make sense out of gene expression profiles, such analyses must be pushed beyond the mere listing of affected genes. For example, if a group of genes persistently display similar changes in expression levels under particular experimental conditions, and the proteins encoded by these genes interact and function in the same cellular compartments, this could be taken as very strong indicators for co-regulated protein complexes. One of the key requirements is having appropriate tools to detect such regulatory patterns.</p> <p>Results</p> <p>We have analyzed the global adaptations in gene expression patterns in the budding yeast when the Hsp90 molecular chaperone complex is perturbed either pharmacologically or genetically. We integrated these results with publicly accessible expression, protein-protein interaction and intracellular localization data. But most importantly, all experimental conditions were simultaneously and dynamically visualized with an animation. This critically facilitated the detection of patterns of gene expression changes that suggested underlying regulatory networks that a standard analysis by pairwise comparison and clustering could not have revealed.</p> <p>Conclusions</p> <p>The results of the animation-assisted detection of changes in gene regulatory patterns make predictions about the potential roles of Hsp90 and its co-chaperone p23 in regulating whole sets of genes. The simultaneous dynamic visualization of microarray experiments, represented in networks built by integrating one's own experimental with publicly accessible data, represents a powerful discovery tool that allows the generation of new interpretations and hypotheses.</p

Structural Insights into the Quinolone Resistance Mechanism of Mycobacterium tuberculosis DNA Gyrase

Mycobacterium tuberculosis DNA gyrase, an indispensable nanomachine involved in the regulation of DNA topology, is the only type II topoisomerase present in this organism and is hence the sole target for quinolone action, a crucial drug active against multidrug-resistant tuberculosis. To understand at an atomic level the quinolone resistance mechanism, which emerges in extensively drug resistant tuberculosis, we performed combined functional, biophysical and structural studies of the two individual domains constituting the catalytic DNA gyrase reaction core, namely the Toprim and the breakage-reunion domains. This allowed us to produce a model of the catalytic reaction core in complex with DNA and a quinolone molecule, identifying original mechanistic properties of quinolone binding and clarifying the relationships between amino acid mutations and resistance phenotype of M. tuberculosis DNA gyrase. These results are compatible with our previous studies on quinolone resistance. Interestingly, the structure of the entire breakage-reunion domain revealed a new interaction, in which the Quinolone-Binding Pocket (QBP) is blocked by the N-terminal helix of a symmetry-related molecule. This interaction provides useful starting points for designing peptide based inhibitors that target DNA gyrase to prevent its binding to DNA

In-situ Combustion Pilot Basic Design and Laboratory Experiments

When it must be decided to develop a field with an enhanced oil recovery method, first it is needed to have a reservoir characterization model of high quality. Then the choice of the best suited method has to be carried out. For any method, a preliminary study has to be performed in order to help to decide. In the case of an in-situ combustion field development, various patterns are considered; at the same time, duration for the combustion front to move from the injector to a producer is analyzed. Field examples of various patterns are presented. The amount of air to inject in case of dry combustion and of air and water in case of wet combustion has to be determined in order to design air compressors and water pumps. The amount of air is a function of the volumetric sweep efficiency and of the oil and the matrix from the reservoir. Lab experiments must be performed in the reservoir matrix with the reservoir oil to determine the air requirement, which is the amount of air needed to burn a unit volume of reservoir. The amount of water is also determined by lab tests. Then the flows of air and of water are determined, which allows the design of compressors and pumps. The amount of oil produced is calculated taking into account the sweep efficiency in the different zones in front of the combustion. Production of oil, water and gas and their compositions obtained at the lab scale are presented. A scheme of the production, treatment and storage for a pilot field test is shown. In conclusion, a diagram shows the general guidelines for the preparation and implementation of field experiments using in-situ combustion

La gazéification souterraine profonde du charbon en France. L'expérience de Bruay-en-Artois

Le projet français de gazéification souterraine du charbon, dont l'objectif à terme est la production d'un substitut de gaz naturel, vise la gazéification de veines de charbon de faible épaisseur, situées à grande profondeur et non exploitables par les méthodes classiques. De nombreuses difficultés sont liées à la profondeur (plus de 1000 m), en particulier la nécessité d'opérer sous pression élevée dans un charbon très peu perméable. Les premiers essais, effectués sur un pilote à 1200 m de la surface sur le site de Bruay-en-Artois (nord de la France), ont consisté à :- relier deux puits distants de 65 m par fracturation hydraulique, un test de la liaison par injection d'azote indiquant un taux de récupération du gaz injecté proche de 50 % ; - allumer le charbon au moyen d'un allumeur électrique ; - initier et tenter de propager une combustion à contre-courant avec injection d'air à faible débit. Des expériences de laboratoire, consécutives à cet essai pilote, ont mis en évidence les risques d'allumage spontané du charbon lorsque la pression partielle d'oxygène dans le gaz injecté est élevée. Un nouvel essai de combustion à contre-courant a alors été entrepris sur le site de Bruay-en-Artois, avec injection d'un gaz contenant 5 % d'oxygène et 95 % d'azote. Les résultats de cette expérimentation arrêtée en juillet 1981 sont en cours d'exploitation

Acetal and ester protecting-groups in the hydrogen fluoride-catalysed synthesis of D-fructose and L-sorbose difuranose dianhydrides

International audienc

The behaviour of D-fructose and inulin towards anhydrous hydrogen fluoride

International audienc