2,270 research outputs found
Structure of the pheromone peptide of the Staphylococcus epidermidis agr system
AbstractThe agr quorum-sensing system is responsible for the regulation of several virulence factors in staphylococci, with an extracellular pheromone peptide as signalling molecule. By monitoring the biological activity of synthetic peptides, it could be demonstrated that the pheromone of the agr system in Staphylococcus epidermidis is an octapeptide containing a thiolester linkage between the central cysteine and the C-terminal carboxyl group. The peptide was active at nanomolar concentrations. The N-terminus of the peptide pheromone, which is encoded as part of a protein precursor, proved to be crucial for biological activity
Monitoring the expression of purinoceptors and nucleotide-metabolizing ecto-enzymes with antibodies directed against proteins in native conformation
Following their release from cells, ATP and NAD, the universal currencies of energy metabolism, function as extracellular signalling molecules. Mammalian cells express numerous purinoceptors, i.e., the nucleotide-gated P2X ion channels and the G-protein-coupled P2Y receptors. Signalling through purinoceptors is controlled by nucleotide-metabolizing ecto-enzymes, which regulate the availability of extracellular nucleotides. These enzymes include ecto-nucleoside triphosphate diphosphohydrolases (ENTPD, CD39 family) and ecto-nucleotide pyrophosphatase/phosphodiesterases (ENPP, CD203 family). Investigation of these receptors and enzymes has been hampered by the lack of available antibodies, especially ones that recognize these proteins in their native conformation. This study reports the use of genetic immunization to generate such antibodies against P2X1, P2X4, P2X7, ENTPD1, ENPTD2, ENPTD5, ENPTD6, ENPP2, ENPP3, ENPP4, ENPP5, and ENPP6. Genetic immunization ensures expression of the native protein by the cells of the immunized animal and yields antibodies directed against proteins in native conformation (ADAPINCs). Such antibodies are especially useful for immunofluorescence and immunoprecipitation analyses, whereas antibodies against synthetic peptides usually function well only in Western-blot analyses. Here we illustrate the utility of the new antibodies to monitor the cell surface expression of and to purify some key players of purinergic signalling
Stable Chinese Hamster Ovary Suspension Cell Lines Harboring Recombinant Human Cytochrome P450 Oxidoreductase and Human Cytochrome P450 Monooxygenases as Platform for In Vitro Biotransformation Studies
In the liver, phase-1 biotransformation of drugs and other xenobiotics is largely facilitated by enzyme complexes consisting of cytochrome P450 oxidoreductase (CPR) and cytochrome P450 monooxygenases (CYPs). Generated from human liver-derived cell lines, recombinant in vitro cell systems with overexpression of defined phase-1 enzymes are widely used for pharmacological and toxicological drug assessment and laboratory-scale production of drug-specific reference metabolites. Most, if not all, of these cell lines, however, display some background activity of several CYPs, making it difficult to attribute effects to defined CYPs. The aim of this study was to generate cell lines with stable overexpression of human phase-1 enzymes based on Chinese hamster ovary (CHO) suspension cells. Cells were sequentially modified with cDNAs for human CPR in combination with CYP1A2, CYP2B6, or CYP3A4, using lentiviral gene transfer. In parallel, CYP-overexpressing cell lines without recombinant CPR were generated. Successful recombinant expression was demonstrated by mRNA and protein analyses. Using prototypical CYP-substrates, generated cell lines proved to display specific enzyme activities of each overexpressed CYP while we did not find any endogenous activity of those CYPs in parental CHO cells. Interestingly, cell lines revealed some evidence that the dependence of CYP activity on CPR could vary between CYPs. This needs to be confirmed in further studies. Recombinant expression of CPR was also shown to enhance CYP3A4-independent metabolisation of testosterone to androstenedione in CHO cells. We propose the novel serum-free CHO suspension cell lines with enhanced CPR and/or defined CYP activity as a promising “humanised” in vitro model to study the specific effects of those human CYPs. This could be relevant for toxicology and/or pharmacology studies in the pharmaceutical industry or medicine
topology of bismuth
While first-principles calculations with different levels of sophistication
predict a topologically trivial state for bulk bismuth, some
photoemission experiments show surface states consistent with the
interpretation of bismuth being in a topologically non-trivial state. We
resolve this contradiction between theory and experiment by showing, based on
quasiparticle self-consistent calculations, that the experimental surface
states interpreted as supporting a non-trivial phase are actually consistent
with a trivial invariant. We identify this contradiction as the result of
a crosstalk effect arising from the extreme penetration depth of the surface
states into the bulk of Bi. A film of Bi can be considered bulk-like only for
thicknesses of about 1000 bilayers ( 400 nm) and more
Quantification of dynamic contrast-enhanced ultrasound (CEUS) in non-cystic breast lesions using external perfusion software
The aim of this present clinical pilot study is the display of typical perfusion results in patients with solid, non-cystic breast lesions. The lesions were characterized using contrast enhanced ultrasound (CEUS) with (i) time intensity curve analyses (TIC) and (ii) parametric color maps. The 24 asymptomatic patients included were genetically tested for having an elevated risk for breast cancer. At a center of early detection of familial ovary and breast cancer, those patients received annual MRI and grey-scale ultrasound. If lesions remained unclear or appeared even suspicious, those patients also received CEUS. CEUS was performed after intravenous application of sulfur hexafluoride microbubbles. Digital DICOM cine loops were continuously stored for one minute in PACS (picture archiving and communication system). Perfusion images and TIC analyses were calculated off-line with external perfusion software (VueBox). The lesion diameter ranged between 7 and 15 mm (mean 11 ± 3 mm). Five hypoechoic irregular lesions were scars, 6 lesions were benign and 12 lesions were highly suspicious for breast cancer with irregular enhancement at the margins and a partial wash out. In those 12 cases, histopathology confirmed breast cancer. All the suspicious lesions were correctly identified visually. For the perfusion analysis only Peak Enhancement (PE) and Area Under the Curve (AUC) added more information for correctly identifying the lesions. Typical for benign lesions is a prolonged contrast agent enhancement with lower PE and prolonged wash out, while scars are characterized typically by a reduced enhancement in the center. No differences (p = 0.428) were found in PE in the center of benign lesions (64.2 ± 28.9 dB), malignant lesions (88.1 ± 93.6 dB) and a scar (40.0 ± 17.0 dB). No significant differences (p = 0.174) were found for PE values at the margin of benign lesions (96.4 ± 144.9 dB), malignant lesions (54.3 ± 86.2 dB) or scar tissue (203.8 ± 218.9 dB). Significant differences (p < 0.001) were found in PE of the surrounding tissue when comparing benign lesions (33.6 ± 25.2 dB) to malignant lesions (15.7 ± 36.3 dB) and scars (277.2 ± 199.9 dB). No differences (p = 0.821) were found in AUC in the center of benign lesions (391.3 ± 213.7), malignant lesions (314.7 ± 643.9) and a scar (213.1 ± 124.5). No differences (p = 0.601) were found in AUC values of the margin of benign lesions (313.3 ± 372.8), malignant lesions (272.6 ± 566.4) or scar tissue (695.0 ± 360.6). Significant differences (p < 0.01) were found in AUC of the surrounding tissue for benign lesions (151.7 ± 127.8), malignant lesions (177.9 ± 1345.6) and scars (1091 ± 693.3). There were no differences in perfusion evaluation for mean transit time (mTT), rise time (RT) and time to peak (TTP) when comparing the center to the margins and the surrounding tissue. The CEUS perfusion parameters PE and AUC allow a very good assessment of the risk of malignant breast lesions and thus a downgrading of BI-RADS 4 lesions. The use of the external perfusion software (VueBox, Bracco, Milan, Italy) did not lead to any further improvement in the diagnosis of suspicious breast lesions and does appears not to have any additional diagnostic value in breast lesions
Statistisches Material zur Beleuchtung livländischer Bauerverhältnisse
http://tartu.ester.ee/record=b2414192~S1*es
Нові інтелектуальні технології та методи оптимізації для дослідження складних систем
Protein transport into the endoplasmic reticulum (ER) is essential for all eukaryotic cells and evolutionary related to protein transport into and across the cytoplasmic membrane of eubacteria and archaea. It is based on amino-terminal signal peptides in the precursor polypeptides plus various transport components in cytosol plus ER and can occur either cotranslationally or posttranslationally. The two mechanisms merge at the heterotrimeric Sec61 complex in the ER membrane, which forms an aqueous polypeptide-conducting channel. Since the mammalian ER is also the main intracellular calcium storage organelle, the Sec61 complex is tightly regulated in its dynamics between the open and closed conformations by various ligands, such as precursor polypeptides at the cytosolic face and the Hsp70-type molecular chaperone BiP at the ER lumenal face (Hsp, heat shock protein). Furthermore, BiP binding to the incoming precursor polypeptide contributes to unidirectionality and efficiency of transport. Recent insights into the structural dynamics of the Sec61 complex and related complexes in eubacteria and archaea have various mechanistic and functional implications
First experimental evidence for quantum echoes in scattering systems
A self-pulsing effect termed quantum echoes has been observed in experiments
with an open superconducting and a normal conducting microwave billiard whose
geometry provides soft chaos, i.e. a mixed phase space portrait with a large
stable island. For such systems a periodic response to an incoming pulse has
been predicted. Its period has been associated to the degree of development of
a horseshoe describing the topology of the classical dynamics. The experiments
confirm this picture and reveal the topological information.Comment: RevTex 4.0, 5 eps-figure
A turbulent model of torque in von Karman swirling flow
A stochastic model is derived to predict the turbulent torque produced by a
swirling flow. It is a simple Langevin process, with a colored noise. Using the
unified colored noise approximation, we derive analytically the PDF of the
fluctuations of injected power in two forcing regimes: constant angular
velocity or constant applied torque. In the limit of small velocity
fluctuations and vanishing inertia, we predict that the injected power
fluctuates twice less in the case of constant torque than in the case of
constant angular velocity forcing. The model is further tested against
experimental data in a von Karman device filled with water. It is shown to
allow for a parameter-free prediction of the PDF of power fluctuations in the
case where the forcing is made at constant torque. A physical interpretation of
our model is finally given, using a quasi-linear model of turbulence
- …