Regulation of bacterial sugar-H+ symport by phosphoenolpyruvate-dependent enzyme I/HPr-mediated phosphorylation

The lactose-H+ symport protein (LacS) of Streptococcus thermophilus has a C-terminal hydrophilic domain that is homologous to IIA protein(s) domains of the phosphoenolpyruvate:sugar phosphotransferase system (PTS). C-terminal truncation mutants were constructed and expressed in Escherichia coli and their properties were analyzed. Remarkably, the entire IIA domain (160 amino acids) could be deleted without significant effect on lactose-H+ symport and galactoside equilibrium exchange. Analysis of the LacS mutants in S. thermophilus cells suggested that transport is affected by PTS-mediated phosphorylation of the IIA domain. For further studies, membrane vesicles of S. thermophilus were fused with cytochrome c oxidase-containing liposomes, and, when appropriate, phosphoenolpyruvate (PEP) plus purified enzyme I and heat-stable protein HPr were incorporated into the hybrid membranes. Generation of a protonmotive force (Delta p) in the hybrid membranes resulted in accumulation of lactose, whereas uptake of the PTS sugar sucrose was not observed. With PEP and the energy-coupling proteins enzyme I and HPr of the PTS on the inside, high rates of sucrose uptake were observed, whereas Delta-driven lactose uptake by wild-type LacS was inhibited. This inhibition was not observed with LacS(Delta 160) and LacS(H552R), indicating that PEP-dependent enzyme I/HPr-mediated phosphorylation of the IIA domain (possibly the conserved His-552 residue) modulates lactose-H+ symport activity.</p

Cell sorting in a Petri dish controlled by computer vision.

Fluorescence-activated cell sorting (FACS) applying flow cytometry to separate cells on a molecular basis is a widespread method. We demonstrate that both fluorescent and unlabeled live cells in a Petri dish observed with a microscope can be automatically recognized by computer vision and picked up by a computer-controlled micropipette. This method can be routinely applied as a FACS down to the single cell level with a very high selectivity. Sorting resolution, i.e., the minimum distance between two cells from which one could be selectively removed was 50-70 micrometers. Survival rate with a low number of 3T3 mouse fibroblasts and NE-4C neuroectodermal mouse stem cells was 66 +/- 12% and 88 +/- 16%, respectively. Purity of sorted cultures and rate of survival using NE-4C/NE-GFP-4C co-cultures were 95 +/- 2% and 62 +/- 7%, respectively. Hydrodynamic simulations confirmed the experimental sorting efficiency and a cell damage risk similar to that of normal FACS

A scale-down mimic for mapping the process performance of centrifugation, depth and sterile filtration

In the production of biopharmaceuticals disk-stack centrifugation is widely used as a harvest step for the removal of cells and cellular debris. Depth filters followed by sterile filters are often then employed to remove residual solids remaining in the centrate. Process development of centrifugation is usually conducted at pilot-scale so as to mimic the commercial scale equipment but this method requires large quantities of cell culture and significant levels of effort for successful characterization. A scale-down approach based upon the use of a shear device and a bench-top centrifuge has been extended in this work towards a preparative methodology that successfully predicts the performance of the continuous centrifuge and polishing filters. The use of this methodology allows the effects of cell culture conditions and large-scale centrifugal process parameters on subsequent filtration performance to be assessed at an early stage of process development where material availability is limited

YAG nano-light sources with high Ce concentration

We investigate the luminescence properties of 10 nm YAG nanoparticles doped with Ce ions at 0.2%, 4% and 13% that are designed as active probes for Scanning Near field Optical Microscopy. They are produced by a physical method without any subsequent treatment, which is imposed by the desired application. The structural analysis reveals the amorphous nature of the particles, which we relate to some compositional defect as indicated by the elemental analysis. The optimum emission is obtained with a doping level of 4%. The emission of the YAG nanoparticles doped at 0.2% is strongly perturbed by the crystalline disorder whereas the 13% doped particles hardly exhibit any luminescence. In the latter case, the presence of Ce4+ ions is confirmed, indicating that the Ce concentration is too high to be incorporated efficiently in YAG nanoparticles in the trivalent state. By a unique procedure combining cathodoluminescence and Rutherford backscattering spectrometry, we demonstrate that the enhancement of the particles luminescence yield is not proportional to the doping concentration, the emission enhancement being larger than the Ce concentration increase. Time-resolved photoluminescence reveals the presence of quenching centres likely related to the crystalline disorder as well as the presence of two distinct Ce ions populations. Eventually, nano-cathodoluminescence indicates that the emission and therefore the distribution of the doping Ce ions and of the defects are homogeneous

Evidence for Color Fluctuations in Hadrons from Coherent Nuclear Diffraction}

A QCD-based treatment of projectile size fluctuations is used to compute inelastic diffractive cross sections $\sigma_{diff}$ for coherent hadron-nuclear processes. We find that fluctuations near the average size give the major contribution to the cross section with $\le few \%$ contribution from small size configurations. The computed values of $\sigma_{diff}$ are consistent with the limited available data. The importance of coherent diffraction studies for a wide range of projectiles for high energy Fermilab fixed target experiments is emphasized. The implications of these significant color fluctuations for relativistic heavy ion collisions are discussed.Comment: Report number DOE/ER 40427-13-N93 11 pages, 3 figures available from author Mille

Correlations and Fluctuations in High-Energy Nuclear Collisions

Nucleon correlations in the target and projectile nuclei are shown to reduce significantly the fluctuations in multiple nucleon-nucleon collisions, total multiplicity and transverse energy in relativistic heavy-ion collisions, in particular for heavy projectile and target. The interplay between cross-section fluctuations, from color transparency and opacity, and nuclear correlations is calculated and found to be able to account for large fluctuations in transverse energy spectra. Numerical implementation of correlations and cross-section fluctuations in Monte-Carlo codes is discussed.Comment: 30 pages, in Revtex, plus 4 figures. Figures and preprint can be obtained by mailing address to: [email protected]

Evolutionary impact assessment: accounting for evolutionary consequences of fishing in an ecosystem approach to fisheries management

Managing fisheries resources to maintain healthy ecosystems is one of the main goals of the ecosystem approach to fisheries (EAF). while the number of international treaties call for the implementation of EAF, there are still gaps in the underlying methodology. One aspect that has received substantial scientific attention recently in fisheries-induced evolution (FIE). Increasing evidence indicates that intensive fishing has the potential to exert strong directional selection on life-history traits, behavior, physiology, and morphology of exploited fish. Of particular concern is that reversing evolutionary responses to fishing can be much more difficult than reversing demographic or phenotypically plastic responses. Furthermore, like climate change, multiple agents cause fisheries-induced evolution with effects accumulating over time. Consequently, FIE may alter then utility derived from fish stocks, which in turn can modify the monetary value living aquatic resources provide to society. Quantifying and predicting the evolutionary effects of fishing is therefore important for both ecological and economic reasons, An important reason this is not happening is the lack of an appropriate assessment framework. We therefor describe the evolutionary impact assessment (EvoIA) as a structured approach for assessing the evolutionary outcomes of alternative management options. EvoIA can contribute to the ecosystem approach to fisheries management by clarifying how evolution may alter stock properties and ecological relations, support the precautionary approach to fisheries management by addressing a previously overlooked source of uncertainty and risk, and thus contribute to sustainable fisheries

Prevalence of anatomical variants and coronary anomalies in 543 consecutive patients studied with 64-slice CT coronary angiography

The aim of our study was to assess the prevalence of variants and anomalies of the coronary artery tree in patients who underwent 64-slice computed tomography coronary angiography (CT-CA) for suspected or known coronary artery disease. A total of 543 patients (389 male, mean age 60.5 ± 10.9) were reviewed for coronary artery variants and anomalies including post-processing tools. The majority of segments were identified according to the American Heart Association scheme. The coronary dominance pattern results were: right, 86.6%; left, 9.2%; balanced, 4.2%. The left main coronary artery had a mean length of 112 ± 55 mm. The intermediate branch was present in the 21.9%. A variable number of diagonals (one, 25%; two, 49.7%; more than two, 24%; none, 1.3%) and marginals (one, 35.2%; two, 46.2%; more than two, 18%; none, 0.6%) was visualized. Furthermore, CT-CA may visualize smaller branches such as the conus branch artery (98%), the sinus node artery (91.6%), and the septal branches (93%). Single or associated coronary anomalies occurred in 18.4% of the patients, with the following distribution: 43 anomalies of origin and course, 68 intrinsic anomalies (59 myocardial bridging, nine aneurisms), three fistulas. In conclusion, 64-slice CT-CA provides optimal visualization of the variable and complex anatomy of coronary arteries because of the improved isotropic spatial resolution and flexible post-processing tool