Characterization of the Biosynthesis, Processing and Kinetic Mechanism of Action of the Enzyme Deficient in Mucopolysaccharidosis IIIC

Heparin acetyl-CoA:alpha-glucosaminide N-acetyltransferase (N-acetyltransferase, EC 2.3.1.78) is an integral lysosomal membrane protein containing 11 transmembrane domains, encoded by the HGSNAT gene. Deficiencies of N-acetyltransferase lead to mucopolysaccharidosis IIIC. We demonstrate that contrary to a previous report, the N-acetyltransferase signal peptide is co-translationally cleaved and that this event is required for its intracellular transport to the lysosome. While we confirm that the N-acetyltransferase precursor polypeptide is processed in the lysosome into a small amino-terminal alpha- and a larger ß- chain, we further characterize this event by identifying the mature amino-terminus of each chain. We also demonstrate this processing step(s) is not, as previously reported, needed to produce a functional transferase, i.e., the precursor is active. We next optimize the biochemical assay procedure so that it remains linear as N-acetyltransferase is purified or protein-extracts containing N-acetyltransferase are diluted, by the inclusion of negatively charged lipids. We then use this assay to demonstrate that the purified single N-acetyltransferase protein is both necessary and sufficient to express transferase activity, and that N-acetyltransferase functions as a monomer. Finally, the kinetic mechanism of action of purified N-acetyltransferase was evaluated and found to be a random sequential mechanism involving the formation of a ternary complex with its two substrates; i.e., N-acetyltransferase does not operate through a ping-pong mechanism as previously reported. We confirm this conclusion by demonstrating experimentally that no acetylated enzyme intermediate is formed during the reaction

Regular consumption of vitamin D-fortified yogurt drink (Doogh) improved endothelial biomarkers in subjects with type 2 diabetes: a randomized double-blind clinical trial

<p>Abstract</p> <p>Background</p> <p>Endothelial dysfunction has been proposed as the underlying cause of diabetic angiopathy that eventually leads to cardiovascular disease, the major cause of death in diabetes. We recently demonstrated the ameliorating effect of regular vitamin D intake on the glycemic status of patients with type 2 diabetes (T2D). In this study, the effects of improvement of vitamin D status on glycemic status, lipid profile and endothelial biomarkers in T2D subjects were investigated.</p> <p>Methods</p> <p>Subjects with T2D were randomly allocated to one of the two groups to receive either plain yogurt drink (PYD; containing 170 mg calcium and no vitamin D/250 mL, n₁= 50) or vitamin D3-fortified yogurt drink (FYD; containing 170 mg calcium and 500 IU/250 mL, n₂= 50) twice a day for 12 weeks. Anthropometric measures, glycemic status, lipid profile, body fat mass (FM) and endothelial biomarkers including serum endothelin-1, E-selectin and matrix metalloproteinase (MMP)-9 were evaluated at the beginning and after the 12-week intervention period.</p> <p>Results</p> <p>The intervention resulted in a significant improvement in fasting glucose, the Quantitative Insulin Check Index (QUICKI), glycated hemoglobin (HbA1c), triacylglycerols, high-density lipoprotein cholesterol (HDL-C), endothelin-1, E-selectin and MMP-9 in FYD compared to PYD (<it>P </it>< 0.05, for all). Interestingly, difference in changes of endothelin-1, E-selectin and MMP-9 concentrations in FYD compared to PYD (-0.35 ± 0.63 versus -0.03 ± 0.55, <it>P </it>= 0.028; -3.8 ± 7.3 versus 0.95 ± 8.3, <it>P </it>= 0.003 and -2.3 ± 3.7 versus 0.44 ± 7.1 ng/mL, respectively, <it>P </it>< 0.05 for all), even after controlling for changes of QUICKI, FM and waist circumference, remained significant for endothelin-1 and MMP-9 (<it>P </it>= 0.009 and <it>P </it>= 0.005, respectively) but disappeared for E-selectin (<it>P </it>= 0.092). On the contrary, after controlling for serum 25(OH)D, the differences disappeared for endothelin-1(<it>P </it>= 0.066) and MMP-9 (<it>P </it>= 0.277) but still remained significant for E-selectin (<it>P </it>= 0.011).</p> <p>Conclusions</p> <p>Ameliorated vitamin D status was accompanied by improved glycemic status, lipid profile and endothelial biomarkers in T2D subjects. Our findings suggest both direct and indirect ameliorating effects of vitamin D on the endothelial biomarkers.</p> <p>Trial registration</p> <p>ClinicalTrials.gov: <a href="http://www.clinicaltrials.gov/ct2/show/NCT01236846">NCT01236846</a></p

Magnetized plasma implosion in a snail target driven by a moderateintensity laser pulse

Optical generation of compact magnetized plasma structures is studied in the moderate intensity domain. A sub-ns laser beam irradiated snail-shaped targets with the intensity of about 10(16) W/cm(2). With a neat optical diagnostics, a sub-megagauss magnetized plasmoid is traced inside the target. On the observed hydrodynamic time scale, the hot plasma formation achieves a theta-pinch-like density and magnetic field distribution, which implodes into the target interior. This simple and elegant plasma magnetization scheme in the moderate-intensity domain is of particular interest for fundamental astrophysical-related studies and for development of future technologies © The Author(s) 201

Short-wavelength experiments on laser pulse interaction with extended pre-plasma at the PALS-installation

The paper is a continuation of research carried out at Prague Asterix Laser System (PALS) related to the shock ignition (SI) approach in inertial fusion, which was carried out with use of 1ω main laser beam as the main beam generating a shock wave. Two-layer targets were used, consisting of Cu massive planar target coated with a thin polyethylene layer, which, in the case of two-beam irradiation geometry, simulate conditions related to the SI scenario. The investigations presented in this paper are related to the use of 3ω to create ablation pressure for high-power shock wave generation. The interferometric studies of the ablative plasma expansion, complemented by measurements of crater volumes and K α emission, clearly demonstrate the effect of changing the incident laser intensity due to changing the focal radius on efficiency of laser energy transfer to a shock wave and fast electron emission. The efficiency of the energy transfer increases with the radius of the focused laser beam. The pre-plasma does not significantly change the character of this effect. However, it unambiguously results in the increasing temperature of fast electrons, the total energy of which remains very small (<0.1% of the laser energy). This study shows that the optimal radius from the point of view of 3ω radiation energy transfer to the shock wave is the maximal one used in these experiments and equal to 200 m that corresponds to the minimal effect of two-dimensional (2D)-expansion. Such a result is typical for the ablation process determined by electron conductivity energy transfer under the conditions of one-dimensional or 2D matter expansion without any appreciable effect due to energy transfer by fast electrons. The 2D simulations based on application of the ALANT-HE code and an analytical model that includes generation and transport of hot electrons has been used to support of experimental data

Pre-plasma effect on laser beam energy transfer to a dense target under conditions relevant to shock ignition

This paper reports on properties of a plasma formed by sequential action of two laser beams on a flat target, simulating the conditions of shock-ignited inertial confinement fusion target exposure. The experiments were performed using planar targets consisting of a massive copper (Cu) plate coated with a thin plastic (CH) layer, which was irradiated by the 1ω PALS laser beam (λ = 1.315 μm) at the energy of 250 J. The intensity of the fixed-energy laser beam was scaled by varying the focal spot radius. To imitate shock ignition conditions, the lower-intensity auxiliary 1ω beam created CH-pre-plasma which was irradiated by the main beam with a delay of 1.2 ns, thus generating a shock wave in the massive part of the target. To study the parameters of the plasma treated by the two-beam irradiation of the targets, a set of various diagnostics was applied, namely: (i) Two-channel polaro-interferometric system irradiated by the femtosecond laser (∼40 fs), (ii) spectroscopic measurements in the X-ray range, (iii) two-dimensional (2D)-resolved imaging of the Kα line emission from Cu, (iv) measurements of the ion emission by means of ion collectors, and (v) measurements of the volume of craters produced in a massive target providing information on the efficiency of the laser energy transfer to the shock wave. The 2D numerical simulations have been used to support the interpretation of experimental data. The general conclusion is that the fraction of the main laser beam energy deposited into the massive copper at two-beam irradiation decreases in comparison with the case of pre-plasma. The reason is that the pre-formed and expanding plasma deteriorates the efficiency of the energy transfer from the mai Copyright © Cambridge University Press 2015