Characterization of TsaR,an oxygen-sensitive LysR-type regulator for the degradation of p-toluenesulfonate in Comamonas testosteroni T-2

TsaR is the putative LysR-type regulator of the tsa operon (tsaMBCD) which encodes the first steps in the degradation of p-toluenesulfonate (TSA) in Comamonas testosteroni T-2. Transposon mutagenesis was used to knock out tsaR. The resulting mutant lacked the ability to grow with TSA and p-toluenecarboxylate (TCA). Reintroduction of tsaR in trans on an expression vector reconstituted growth with TSA and TCA. The tsaR gene was cloned into Escherichia coli with a C-terminal His tag and overexpressed as TsaR(His). TsaR(His) was subject to reversible inactivation by oxygen, which markedly influenced the experimental approaches used. Gel filtration showed TsaR(His) to be a monomer in solution. Overexpressed TsaR(His) bound specifically to three regions within the promoter between the divergently transcribed tsaR and tsaMBCD. The dissociation constant (K(D)) for the whole promoter region was about 0.9 micro M, and the interaction was a function of the concentration of the ligand TSA. A regulatory model for this LysR-type regulator is proposed on the basis of these data.     

Gentiana purpurea L. als Vertreter der skandinavischen Bergflora

Ohne Zusammenfassung     

Conformationally constrained NR2B selective NMDA receptor antagonists derived from ifenprodil: Synthesis and biological evaluation of tetrahydro-3-benzazepine-1,7-diols

NR2B selective NMDA receptor antagonists with tetrahydro-3-benzazepine-1,7-diol scaffold have been designed by formal cleavage and reconstitution of the piperidine ring of the lead compound ifenprodil (1). The secondary amine 10 represents the central building block for the synthesis of more than 25 tetrahydro-3-benzazepin-1-ols. Generally 7-hydroxy derivatives display higher NR2B receptor affinities than the corresponding 7-benzyloxy compounds. A distance of four atoms (five bond lengths) between the basic amino group and the terminal aryl moiety led to highest NR2B affinity. 3-(4-Phenylbutyl)-2,3,4,5-tetrahydro-1H-3-benzazepine-1,7-diol (WMS-1410, 25) represents the most promising NR2B antagonist of this series showing a K(i)-value of 14nM. Compound 25 reveals excellent selectivity over more than 100 further relevant target proteins, antagonizes glutamate induced excitotoxicity (IC(50)=18.4nM) and is metabolically more stable than ifenprodil. Up to a dose of 100mg/kg 25 is well tolerated by mice and it shows dose dependent analgesic activity in the late neuropathic pain phase of the formalin assay.     

Interleukin-1 signaling is dependent on free thiols

Stimulation of the Interleukin-1 receptor type I (IL-1-RI) with IL-1 activates an associated serine/threonine kinase, IRAK, which phosphorylates downstream targets, resulting in NFkappaB activation. The signaling cascade is accompanied by oxidative processes and contains putative targets for redox regulation. Preincubation of the murine T cell line EL-4 and the human umbilical cord vein endothelial cell line ECV 304 with thiol modifying compounds like diamide, menadione or phenylarsine oxide inhibited the IL-1-induced phosphorylation of an endogenous substrate with a molecular mass of 60 kD. In the endothelial cell line, a second target of about 85 kD was phosphorylated after IL-1 stimulation, which was also inhibited by thiol modification. These data suggest that IL-1 signal transduction depends on free thiols which might be targets for redox regulation not only in lymphocytes, but also in endothelial cells.     

Detection of Rev peptides with impedance-sensors--comparison of device-geometries

Two different impedance-sensor geometries have been compared for the detection of Rev peptides with a molecular weight of 2.4 kDa. Planar, two-dimensional interdigitated capacitor (IDC) sensors with electrode separations of 1.1 microm as well as three-dimensional nanogap-sensors with an electrode separation of 75 nm have been used. Both sensors have been operated at a fixed frequency of 980 MHz. We discuss the specific interaction of the Rev peptide to an immobilized RNA anti-Rev aptamer (9.2 kDa) for peptide concentrations in the range of 100 nM-2 microM. For the IDC sensor, only peptide concentrations above 500 nM gave detectable signals. For the nanogap sensor, the binding process was clearly visible for all concentrations applied. The higher sensitivity of the nanogap compared to the IDC is ascribed to the improved surface-to-volume ratio.     

Immobilization of wild ocelots with tiletamine and zolazepam in southern Texas

Telazol was used to immobilize nine wild ocelots (Leopardus pardalis) captured in box-traps in southern Texas (USA) between May 1997 and April 1998. Mean (+/- SD) intramuscular dosage rate of 5.05 (+/- 0.76) mg/kg produced an induction time of 3.7 +/- 1.8 min. Duration of cataleptic anesthesia was 67.4 +/- 19.8 min and ocelots stood 50.0 +/- 30.7 min after emergence from cataleptic anesthesia. Ocelots recovered to their preinjection condition 129.7 +/- 28.8 min after first standing and 250.8 +/- 55.1 min after initial injection. We observed no adverse reactions to Telazol aside from minor loss of thermoregulatory control. Telazol administered at 5 mg/kg was an effective and safe immobilizing agent for wild ocelots.     

Circadian rhythmicity during prolonged chemostat cultivation of Neurospora crassa

Following exposure to light and attainment of steady-state in the chemostat, Neurospora was grown in constant conditions of darkness at 25 degrees C for 6 days. Biomass samples were taken every 4h for the extraction of RNA and protein, and the state of the circadian clock was assessed by assaying the levels of three rhythmically expressed mRNAs; frequency (frq), antisense frq (qrf) and clock-controlled gene-14 (ccg-14), and by monitoring the clock-controlled rhythm of sporulation. Our results indicate that the Neurospora clock continued to run in the chemostat. This is the longest reported time that Neurospora has been grown in a chemostat in filamentous form and opens up the possibility of studying the response of Neurospora to a range of stimuli in the absence of confounding effects due to; alterations in growth rate, aging, and changing conditions of the growth medium.     

Survival of radiocollared adult leopard cats<Emphasis Type="Italic">Prionailurus bengalensis</Emphasis> in Thailand

From December 1999 to December 2002, we radiocollared 25 adult (17 M, 8 F) leopard catsPrionailurus bengalensis Kerr, 1792 in Phu Khieo Wildlife Sanctuary, Thailand. We found no difference in annual survival rate between adult males (0.96 ±0.05) and females (0.87 ± 0.13), and we found no difference in mean seasonal survival rates between the dry (0.95 ± 0.04) and wet season (0.97 ± 0.04). We estimated a pooled mean annual survival of 0.92 ± 0.06 for adult leopard cats, and also estimated annual survival rates of 0.83 ± 0.24 and 0.53 ± 0.21 for leopard cats in 2 other studies. We believe our high mean annual survival estimate of adult leopard cats was related to our study site being located within a remote wildlife sanctuary with minimal anthropogenic effects.