Biosynthesis and NMR-studies of a double transmembrane domain from the Y4 receptor, a human GPCR

The human Y4 receptor, a class A G-protein coupled receptor (GPCR) primarily targeted by the pancreatic polypeptide (PP), is involved in a large number of physiologically important functions. This paper investigates a Y4 receptor fragment (N-TM1-TM2) comprising the N-terminal domain, the first two transmembrane (TM) helices and the first extracellular loop followed by a (His)6 tag, and addresses synthetic problems encountered when recombinantly producing such fragments from GPCRs in Escherichia coli. Rigorous purification and usage of the optimized detergent mixture 28 mM dodecylphosphocholine (DPC)/118 mM% 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-rac-(1-glycerol)] (LPPG) resulted in high quality TROSY spectra indicating protein conformational homogeneity. Almost complete assignment of the backbone, including all TM residue resonances was obtained. Data on internal backbone dynamics revealed a high secondary structure content for N-TM1-TM2. Secondary chemical shifts and sequential amide proton nuclear Overhauser effects defined the TM helices. Interestingly, the properties of the N-terminal domain of this large fragment are highly similar to those determined on the isolated N-terminal domain in the presence of DPC micelles

Detection of Quadruplex DNA by Gold Nanoparticles

Gold nanoparticles have been used as a probe to detect low (<10 ppb) concentrations of quadruplex DNA. These nanoparticles display a tendency to form aggregates in the presence of certain quadruplex forms, as observed via enhanced plasmon resonance light scattering (PRLS) signals. These nanoparticles showed differing degrees of interactions with different types of quadruplex and mixed sequences but no interaction with duplex DNA. Enhancement of PRLS signals greater than 50% was observed at nanomolar DNA concentration, and a lower limit of detection of 2.1 nM was established for three different quadruplex DNA sequences, including the thrombin-inhibiting single-stranded 15 mer aptamer DNA, d(GGTTGGTGTGGTTGG), and the double-stranded 12 mer DNA, d(G4T4G4). Two different sample preparation protocols were used for the PRLS experiments, and they yielded similar results

Productivity of reed (Phragmites australis Trin. ex Steud.) in continental-arid NW China in relation to soil, groundwater, and land-use

Reed (Phragmites australis Trin. ex Steud.) is a cosmopolitan plant species which can build up large stands in wetlands, floodplains, and on sites where groundwater is available. Phragmites australis provides many ecosystem services, such as the production of raw material (e.g. house construction or organic fuel). In the desert regions of Central Asia, reed occurs along river, e.g. the Tarim, Syr Darya, Amu Darya, and serves as fodder plant and raw material for construction and paper production. In those arid regions, reed occurs on submerged sites as well as non-flooded sites in a wide variety of phenotypes, ranging from so-called „giant reed“ (2-4 m high) to dwarf-like thorny reed not exceeding 40 cm stem length. We investigated the net primary production of the different phenotypes and their distribution with regard to soil and groundwater salt content and regarding grazing. The phenotypes were characterized through stem length, stem diameter, number of leaves per stem length, leaf weight ratio, leaf length, and leaf width. The net primary production reached 6,004 kg/ha·a on a non-grazed site, which is submerged for one month in late summer. The depth of the closed capillary fringe before onset of the flood was 2.2 m. The electric conductivity at the closed capillary fringe (determined from a water saturated soil extract) was 2 mS/cm. Stem length and stem diameter did not decrease with increasing soil and groundwater salt content, as expected. Conversely, stem length and stem diameter decreased and leaf weight ratio increased with increasing grazing intensity. Thus, grazing turned reed into dwarf-like thorny phenotypes. Non-grazed reed stands are the most productive ecosystems of the riparian vegetation at the Tarim and have a high potential to be used as raw material plant. We conclude that biomass harvesting could be an alternative to grazing with regard to sustainable land use

IMMUNOLOGICAL ASPECTS OF CROHN'S DISEASE: A REGULATORY FUNCTION OF TIM-3/GALECTIN-9 IN LYTH1.

Crohn's disease (CD) is a type of inflammatory bowel disease (IBD) and its etiology is multifactorial and involves a combination of genetic environmental factors. The interaction of these factors causes an imbalance of the microbiota, leading to the activation of several immunological and inflammatory mechanism. From an immunological point of view, there seems to be an involvement of the TIM-3/GALECTIN-9 pathway and of the autoregulation of lyTh1. These studies show that in patients with CD the autoregulation of lyth1 is lost due to a reduced concentration of galectin-9 and a reduced TIM-3 expression in LyTh1. This could be one of the reason for the state of perpetual activation of lyTh1, resulting in the chronic inflammation process

Floquet-Markov description of the parametrically driven, dissipative harmonic quantum oscillator

Using the parametrically driven harmonic oscillator as a working example, we study two different Markovian approaches to the quantum dynamics of a periodically driven system with dissipation. In the simpler approach, the driving enters the master equation for the reduced density operator only in the Hamiltonian term. An improved master equation is achieved by treating the entire driven system within the Floquet formalism and coupling it to the reservoir as a whole. The different ensuing evolution equations are compared in various representations, particularly as Fokker-Planck equations for the Wigner function. On all levels of approximation, these evolution equations retain the periodicity of the driving, so that their solutions have Floquet form and represent eigenfunctions of a non-unitary propagator over a single period of the driving. We discuss asymptotic states in the long-time limit as well as the conservative and the high-temperature limits. Numerical results obtained within the different Markov approximations are compared with the exact path-integral solution. The application of the improved Floquet-Markov scheme becomes increasingly important when considering stronger driving and lower temperatures.Comment: 29 pages, 7 figure

Decoherence and Initial Correlations in Quantum Brownian Motion

We analyze the evolution of a quantum Brownian particle starting from an initial state that contains correlations between this system and its environment. Using a path integral approach, we obtain a master equation for the reduced density matrix of the system finding relatively simple expressions for its time dependent coefficients. We examine the evolution of delocalized initial states (Schr\"odinger's cats) investigating the effectiveness of the decoherence process. Analytic results are obtained for an ohmic environment (Drude's model) at zero temperature.Comment: 15 pages, RevTex, 5 figures included. Submitted to Phys. Rev.

In vitro and in vivo evaluation of a somatostatin analogue released from PLGA microspheres

The purpose of this study was to design poly(lactide-co-glycolide) (PLGA) microspheres for the continuous delivery of the somatostatin analogue, vapreotide, over 2–4 weeks. The microspheres were produced by spray-drying and the desired characteristics, i.e. high encapsulation efficiency and controlled release over 2–4 weeks, achieved through optimizing the type of polymer, processing solvent, and co-encapsulated additive. The in vitro release was tested in fetal bovine serum preserved with 0.02% of thiomersal. Furthermore, formulations were injected intramuscularly into rats to obtain pharmacokinetic profiles. Encapsulation efficiency was between 34 and 91%, depending on the particular formulation. The initial peptide release (within 6 h) was lowest, i.e. 1 ng/ml) over 21–28 days in rats was the one made with end-group uncapped PLGA 50:50, the solvent acetic acid and the additive polyethyleneglycol. In conclusion, the optimization of formulation parameters allowed us to produce vapreotide-loaded PLGA microspheres of suitable characteristics for therapeutic use