Isolation of an ovine pulmonary surfactant-associated anionic peptide bactericidal for Pasteurella haemolytica.

Ovine pulmonary surfactant is bactericidal for Pasteurella haemolytica when surfactant and bacteria mixtures are incubated with normal ovine serum. To isolate this component, surfactant (1 mg/ml) was centrifuged at 100,000 x gav, and the supernatant was fractionated by HPLC. Fractions were eluted with acetonitrile (10-100%)/0.1% trifluoracetic acid and tested for bactericidal activity. Amino acid and sequence analysis of three bactericidal fractions showed that fraction 2 contained H-GDDDDDD-OH, fraction 3 contained H-DDDDDDD-OH, and fraction 6 contained H-GADDDDD-OH. Peptides in 0.14 M NaCl/10 microM ZnCl2 (zinc saline solution) induced killing of P. haemolytica and other bacteria comparable to defensins and beta-defensins [minimal bactericidal concentration (MBC)50 range, 0.01-0.06 mM] but not in 0.14 M NaCl/10 mM sodium phosphate buffer, pH 7.2/0.5 mM CaCl2/0.15 mM MgCl2 (MBC50 range, 2.8-11.5 mM). Bactericidal activity resided in the core aspartate hexapeptide homopolymeric region, and MBC50 values of aspartate dipeptide-to-heptapeptide homopolymers were inversely proportional to the number of aspartate residues in the peptide. P. haemolytica incubated with H-DDDDDD-OH in zinc saline solution was killed within 30 min. Ultrastructurally, cells contained flocculated intracellular constituents. In contrast to cationic defensins and beta-defensins, surfactant-associated anionic peptides are smaller in size, opposite in charge, and are bactericidal in zinc saline solution. They are members of another class of peptide antibiotics containing aspartate, which when present in pulmonary secretions may help clear bacteria as a part of the innate pulmonary defense system.

Vector-mediated delivery of 125I-labeled beta-amyloid peptide A beta 1-40 through the blood-brain barrier and binding to Alzheimer disease amyloid of the A beta 1-40/vector complex.

The brain amyloid of Alzheimer disease (AD) may potentially be imaged in patients with AD by using neuroimaging technology and a radiolabeled form of the 40-residue beta-amyloid peptide A beta 1-40 that is enabled to undergo transport through the brain capillary endothelial wall, which makes up the blood-brain barrier (BBB) in vivo. Transport of 125I-labeled A beta 1-40 (125I-A beta 1-40) through the BBB was found to be negligible by experiments with both an intravenous injection technique and an internal carotid artery perfusion method in anesthetized rats. In addition, 125I-A beta 1-40 was rapidly metabolized after either intravenous injection or internal carotid artery perfusion. BBB transport was increased and peripheral metabolism was decreased by conjugation of monobiotinylated 125I-A beta 1-40 to a vector-mediated drug delivery system, which consisted of a conjugate of streptavidin (SA) and the OX26 monoclonal antibody to the rat transferrin receptor, which undergoes receptor-mediated transcytosis through the BBB. The brain uptake, expressed as percent of injected dose delivered per gram of brain, of the 125I,bio-A beta 1-40/SA-OX26 conjugate was 0.15 +/- 0.01, a level that is 2-fold greater than the brain uptake of morphine. The binding of the 125I,bio-A beta 1-40/SA-OX26 conjugate to the amyloid of AD brain was demonstrated by both film and emulsion autoradiography performed on frozen sections of AD brain. Binding of the 125I,bio-A beta 1-40/SA-OX26 conjugate to the amyloid of AD brain was completely inhibited by high concentrations of unlabeled A beta 1-40. In conclusion, these studies show that BBB transport and access to amyloid within brain may be achieved by conjugation of A beta 1-40 to a vector-mediated BBB drug delivery system.

Rapid mass spectrometric peptide sequencing and mass matching for characterization of human melanoma proteins isolated by two-dimensional PAGE.

We report a general mass spectrometric approach for the rapid identification and characterization of proteins isolated by preparative two-dimensional polyacrylamide gel electrophoresis. This method possesses the inherent power to detect and structurally characterize covalent modifications. Absolute sensitivities of matrix-assisted laser desorption ionization and high-energy collision-induced dissociation tandem mass spectrometry are exploited to determine the mass and sequence of subpicomole sample quantities of tryptic peptides. These data permit mass matching and sequence homology searching of computerized peptide mass and protein sequence data bases for known proteins and design of oligonucleotide probes for cloning unknown proteins. We have identified 11 proteins in lysates of human A375 melanoma cells, including: alpha-enolase, cytokeratin, stathmin, protein disulfide isomerase, tropomyosin, Cu/Zn superoxide dismutase, nucleoside diphosphate kinase A, galaptin, and triosephosphate isomerase. We have characterized several posttranslational modifications and chemical modifications that may result from electrophoresis or subsequent sample processing steps. Detection of comigrating and covalently modified proteins illustrates the necessity of peptide sequencing and the advantages of tandem mass spectrometry to reliably and unambiguously establish the identity of each protein. This technology paves the way for studies of cell-type dependent gene expression and studies of large suites of cellular proteins with unprecedented speed and rigor to provide information complementary to the ongoing Human Genome Project.

Noise and PSRR improvement technique for TPC readout front-end in CMOS. technology.

ALICE is one of four major experiments of particle accelerator LHC installed in the European laboratory CERN. The management committee of the LHC accelerator has just approved a program update for this experiment. Among the upgrades planned for the coming years of the ALICE experiment is to improve the resolution and tracking efficiency maintaining the excellent particles identification ability, and to increase the read-out event rate to 100 KHz. In order to achieve this, it is necessary to update the Time Projection Chamber detector (TPC) and Muon tracking (MCH) detector modifying the read-out electronics, which is not suitable for this migration. To overcome this limitation the design, fabrication and experimental test of new ASIC named SAMPA has been proposed . This ASIC will support both positive and negative polarities, with 32 channels per chip and continuous data readout with smaller power consumption than the previous versions. This work aims to design, fabrication and experimental test of a readout front-end in 130nm CMOS technology with configurable polarity (positive/negative), peaking time and sensitivity. The new SAMPA ASIC can be used in both chambers (TPC and MCH). The proposed front-end is composed of a Charge Sensitive Amplifier (CSA) and a Semi-Gaussian shaper. In order to obtain an ASIC integrating 32 channels per chip, the design of the proposed front-end requires small area and low power consumption, but at the same time requires low noise. In this sense, a new Noise and PSRR (Power Supply Rejection Ratio) improvement technique for the CSA design without power and area impact is proposed in this work. The analysis and equations of the proposed circuit are presented which were verified by electrical simulations and experimental test of a produced chip with 5 channels of the designed front-end. The measured equivalent noise charge was <550e for 30mV/fC of sensitivity at a input capacitance of 18.5pF. The total core area of the front-end was 2300?m × 150?m, and the measured total power consumption was 9.1mW per channel.

Efficient tool path computation using multi-core GPUs

Tool path generation is one of the most complex problems in Computer Aided Manufacturing. Although some efficient strategies have been developed, most of them are only useful for standard machining. However, the algorithms used for tool path computation demand a higher computation performance, which makes the implementation on many existing systems very slow or even impractical. Hardware acceleration is an incremental solution that can be cleanly added to these systems while keeping everything else intact. It is completely transparent to the user. The cost is much lower and the development time is much shorter than replacing the computers by faster ones. This paper presents an optimisation that uses a specific graphic hardware approach using the power of multi-core Graphic Processing Units (GPUs) in order to improve the tool path computation. This improvement is applied on a highly accurate and robust tool path generation algorithm. The paper presents, as a case of study, a fully implemented algorithm used for turning lathe machining of shoe lasts. A comparative study will show the gain achieved in terms of total computing time. The execution time is almost two orders of magnitude faster than modern PCs.