Chimeric streptavidins as host proteins for artificial metalloenzymes

The streptavidin scaffold was expanded with well-structured naturally occurring motifs. These chimeric scaffolds were tested as hosts for biotinylated catalysts as artificial metalloenzymes (ArM) for asymmetric transfer hydrogenation, ring-closing metathesis and anion−π catalysis. The additional second coordination sphere elements significantly influence both the activity and the selectivity of the resulting hybrid catalysts. These findings lead to the identification of propitious chimeric streptavidins for future directed evolution efforts of artificial metalloenzymes

Potent interaction of flavopiridol with MRP1

The multidrug resistance protein 1 (MRP1) is an ATP-dependent transport protein for organic anions, as well as neutral or positively charged anticancer agents. In this study we show that flavopiridol, a synthetic flavonoid currently studied in phase 1 trials for its anti-proliferative characteristics, interacts with MRP1 in a potent way. Flavopiridol, as well as other (iso)flavonoids stimulate the ATPase activity of MRP1 in a dose-dependent way at low micromolar concentrations. A new specific monoclonal antibody against MRP1 (MIB6) inhibits the (iso)flavonoid-induced ATPase activity of plasma membrane vesicles prepared from the MRP1 overexpressing cell line GLC4/ADR. The accumulation of daunorubicin in GLC4/ADR cells is increased by flavopiridol and by other non-glycosylated (iso)flavonoids that interact with MRP1 ATPase activity. However, flavopiridol is the only tested compound that affects the daunorubicin accumulation when present at concentrations below 1 μM. Glycosylated (iso)flavonoids do not affect MRP1-mediated transport or ATPase activity. Finally, MRP1 overexpressing and transfected cells are resistant to flavopiridol, but not to other (iso)flavonoids tested. These findings may be of relevance for the development of anticancer therapies with flavopiridol. © 1999 Cancer Research Campaig

Kavezno izlaganje lubina (Dicentrarchus labrax) u procjeni genotoksičnog utjecaja onečišćenja

Genotoxic effects are often the earliest signs of pollution-related environmental disturbance. In this study, we used the comet assay and micronucleus test to assess DNA damage in the erythrocytes of the European sea bass (Dicentrarchus labrax) exposed to environmental pollution in situ. Fish were collected from a fi sh farm in the Trogir Bay and their cages placed at an unpolluted reference site Šolta (Nečujam Bay) and a polluted site Vranjic (Kaštela Bay) for four weeks. A group of fi sh which remained at the fi sh farm Trogir Bay were used as the second control group. Fish exposed at the Vranjic site showed a signifi cantly higher erythrocyte DNA damage, measured by the comet assay, than either control group. Micronucleus induction showed a similar gradient of DNA damage, but did not reach statistical signifi cance. Our results show that cage exposure of a marine fi sh D. labrax can be useful in environmental biomonitoring and confi rm the comet assay as a suitable tool for detecting pollution-related genotoxicity.Genotoksični učinak često je jedan od najranijih pokazatelja štetnog djelovanja onečišćenja okoliša. U ovom radu procijenjeno je oštećenje DNA u eritrocitima lubina (Dicentrarchus labrax) izloženima okolišnom onečišćenju s pomoću komet-testa i mikronukleus-testa. Lubini su prikupljeni na ribogojilištu i kavezno izloženi u periodu od četiri tjedna na dvije postaje različitog stupnja onečišćenja na jadranskoj obali: na kontrolnoj postaji Šolta (zaljev Nečujam) i na onečišćenoj postaji Vranjic (Kaštelanski zaljev). Zasebna skupina lubina skupljena na ribogojilištu poslužila je kao druga kontrola. Rezultati komet-testa pokazali su statistički značajan porast oštećenja DNA na postaji Vranjic u usporedbi s obje kontrolne postaje. Rezultati mikronukleus-testa pokazali su sličan gradijent onečišćenja, iako nisu dosegli statističku značajnost. Ovi rezultati upućuju na primjenjivost kaveznog izlaganja lubina D. labrax u biomonitoringu vodenog okoliša te potvrđuju korisnost komet-testa kao prikladne metode za detekciju genotoksičnog utjecaja onečišćenja

The Zea mays mutants opaque-2 and opaque-7 disclose extensive changes in endosperm metabolism as revealed by protein, amino acid, and transcriptome-wide analyses

<p>Abstract</p> <p>Background</p> <p>The changes in storage reserve accumulation during maize (<it>Zea mays </it>L.) grain maturation are well established. However, the key molecular determinants controlling carbon flux to the grain and the partitioning of carbon to starch and protein are more elusive. The <it>Opaque-2 </it>(<it>O2</it>) gene, one of the best-characterized plant transcription factors, is a good example of the integration of carbohydrate, amino acid and storage protein metabolisms in maize endosperm development. Evidence also indicates that the <it>Opaque-7 </it>(<it>O7</it>) gene plays a role in affecting endosperm metabolism. The focus of this study was to assess the changes induced by the <it>o2 </it>and <it>o7 </it>mutations on maize endosperm metabolism by evaluating protein and amino acid composition and by transcriptome profiling, in order to investigate the functional interplay between these two genes in single and double mutants.</p> <p>Results</p> <p>We show that the overall amino acid composition of the mutants analyzed appeared similar. Each mutant had a high Lys and reduced Glx and Leu content with respect to wild type. Gene expression profiling, based on a unigene set composed of 7,250 ESTs, allowed us to identify a series of mutant-related down (17.1%) and up-regulated (3.2%) transcripts. Several differentially expressed ESTs homologous to genes encoding enzymes involved in amino acid synthesis, carbon metabolism (TCA cycle and glycolysis), in storage protein and starch metabolism, in gene transcription and translation processes, in signal transduction, and in protein, fatty acid, and lipid synthesis were identified. Our analyses demonstrate that the mutants investigated are pleiotropic and play a critical role in several endosperm-related metabolic processes. Pleiotropic effects were less evident in the <it>o7 </it>mutant, but severe in the <it>o2 </it>and <it>o2o7 </it>backgrounds, with large changes in gene expression patterns, affecting a broad range of kernel-expressed genes.</p> <p>Conclusion</p> <p>Although, by necessity, this paper is descriptive and more work is required to define gene functions and dissect the complex regulation of gene expression, the genes isolated and characterized to date give us an intriguing insight into the mechanisms underlying endosperm metabolism.</p

An EPR study of the radicals issued from nitrocatechols and nitrohydroquinone: a new method of production of nitroaromatic anion radicals

Nitrocatechols and nitrohydroquinone have been treated by means of sodium borohydride in oxygenated aqueous solution in a pH range between 9 to 12, and the EPR spectra have been recorded. In all cases, the nitroaromatic anion radical is observed. At pH=12, a semiquinonic radical is observed in the case of nitrohydroquinone and 4- nitrocatechol

Active site model peptides of glutathione peroxidase : a

Nous avons synthétisé et enregistré les spectres RMN 1H et 77Se de peptides modèles du site actif de la glutathion peroxydase possédant un résidu sélénocystéine afin d’envisager l’utilisation de la RMN du sélénium-77 pour la détermination du mécanisme d’action de l’enzyme. Nous avons montré que l’oxydation de la forme réduite sélénol par un hydroperoxyde en présence de thiol conduit à un sélénénamide cyclique et à un diséléniure. Les formes acide sélénénique, acide séléninique, thioséléninate et séléninamide n’ont pas été détectées au cours de la réaction. La réduction du sélénénamide par un thiol conduit au sélénosulfure et au diséléniure. La forme sélénol n’a pas été observée dans ce cas