998 resultados para M2


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In the ionotropic glutamate receptor, the global conformational changes induced by partial agonists are smaller than those induced by full agonists. However, in the pentameric ligand-gated ion channel receptor family, the structural basis of partial agonism is not understood. This study investigated whether full and partial agonists induce different conformation changes in the glycine receptor chloride channel ( GlyR). A substituted cysteine accessibility analysis demonstrated previously that glycine binding induced an increase in surface accessibility of all residues from Arg(271) to Lys(276) in the M2-M3 domain of the homomeric alpha1 GlyR. Here we compare the surface accessibility changes induced by the full agonist, glycine, and the partial agonist, taurine. In GlyRs incorporating the A272C, S273C, L274C, or P275C mutation, the reaction rate of the cysteine-specific compound, methanethiosulfonate ethyltrimethylammonium, depended on how strongly the receptors were activated but was agonist-independent. Reaction rates could not be compared in the R271C and K276C mutant GlyRs because methanethiosulfonate ethyltrimethylammonium did not modify the extremely small currents induced by saturating taurine or equivalent low glycine concentrations. The results indicate that bound taurine and glycine molecules impose identical conformational changes to the M2-M3 domain. We therefore conclude that the higher efficacy of glycine is due to an increased ability to stabilize a common activated configuration.

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The external loop linking the M2 and M3 transmembrane domains is crucial for coupling agonist binding to channel gating in the glycine receptor chloride channel (GlyR). A substituted cysteine accessibility scan previously showed that glycine activation increased the surface accessibility of 6 contiguous residues (Arg(271) Lys(276)) toward the N-terminal end of the homomeric alpha 1 GlyR M2 - M3 loop. In the present study we used a similar approach to determine whether the allosteric antagonist, picrotoxin, could impose conformational changes to this domain that cannot be induced by varying agonist concentrations alone. Picrotoxin slowed the reaction rate of a sulfhydryl-containing compound ( MTSET) with A272C, S273C, and L274C. Before interpreting this as a picrotoxin-specific conformational change, it was necessary to eliminate the possibility of steric competition between picrotoxin and MTSET. Accordingly, we showed that picrotoxin and the structurally unrelated blocker, bilobalide, were both trapped in the R271C GlyR in the closed state and that a point mutation to the pore-lining Thr(6') residue abolished inhibition by both compounds. We also demonstrated that the picrotoxin dissociation rate was linearly related to the channel open probability. These observations constitute a strong case for picrotoxin binding in the pore. We thus conclude that the picrotoxin-specific effects on the M2 - M3 loop are mediated allosterically. This suggests that the M2 - M3 loop responds differently to the occupation of different binding sites.

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The aim of the work described in this paper was two-fold: (1) the purification of the hydroxylase component of the MSAMO to electrophoretic homogeneity using a four-step chromatographic strategy and (2) the crystallization of the two-component hydroxylase of the MSAMO in order to enhance our understanding of the precise three-dimensional structure of the MSAMO, thus yielding an insight into the nature of the active site of this enzyme. Optimised crystallization conditions were identified allowing growth of crystals of the hydroxylase component of the MSAMO within five days. Crystals exhibited a brown colour suggesting the presence on an intact Rieske-iron sulfur centre and diffracted to 7.0 Å when a few degrees of data were evaluated on a beam line X11. © 2006 Elsevier Inc. All rights reserved.

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Aims: Pulmonary arterial hypertension [1] is a proliferative disorder associated with enhanced proliferation and suppressed apoptosis of pulmonary artery smooth muscle cells (PASMCs). Reactive oxygen species (ROS) is implicated in the development of PAH and regulates the vascular tone and functions. However, which cellular signaling mechanisms are triggered by ROS in PAH is still unknown. Hence, here we wished to characterize the signaling mechanisms triggered by ROS. Methods and Results: By Western blots, we showed that increased intracellular ROS caused inhibition of the glycolytic pyruvate kinase M2 (PKM2) activity through promoting the phosphorylation of PKM2. Monocrotaline (MCT)-induced rats developed severe PAH and right ventricular hypertrophy, with a significant increase in the P-PKM2 and decrease in pyruvate kinase activity which could be attenuated with the treatments of PKM2 activators, FBP and l-serine. The antioxidant NAC, apocynin and MnTBAP had the similar protective effects in the development of PAH. In vitro assays confirmed that inhibition of PKM2 activity could modulate the flux of glycolytic intermediates in support of cell proliferation through the increased pentose phosphate pathway (PPP). Increased ROS and decreased PKM2 activity also promoted the Cav1.2 expression and intracellular calcium. Conclusion: Our data provide new evidence that PKM2 makes a critical regulatory contribution to the PAHs for the first time. Decreased pyruvate kinase M2 activity confers additional advantages to rat PASMCs by allowing them to sustain anti-oxidant responses and thereby support cell survival in PAH. It may become a novel treatment strategy in PAH by using of PKM2 activators.

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This article provides necessary and sufficient conditions for both of the Diophantine equations X^2 − DY^2 = m1 and x^2 − Dy^2 = m2 to have primitive solutions when m1 , m2 ∈ Z, and D ∈ N is not a perfect square. This is given in terms of the ideal theory of the underlying real quadratic order Z[√D].

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Many applications of high-power laser diodes demand tight focusing. This is often not possible due to the multimode nature of semiconductor laser radiation possessing beam propagation parameter M2 values in double-digits. We propose a method of 'interference' superfocusing of high-M2 diode laser beams with a technique developed for the generation of Bessel beams based on the employment of an axicon fabricated on the tip of a 100 μm diameter optical fiber with highprecision direct laser writing. Using axicons with apex angle 140º and rounded tip area as small as 10 μm diameter, we demonstrate 2-4 μm diameter focused laser 'needle' beams with approximately 20 μm propagation length generated from multimode diode laser with beam propagation parameter M2=18 and emission wavelength of 960 nm. This is a few-fold reduction compared to the minimal focal spot size of 11 μm that could be achieved if focused by an 'ideal' lens of unity numerical aperture. The same technique using a 160º axicon allowed us to demonstrate few-μm-wide laser 'needle' beams with nearly 100 μm propagation length with which to demonstrate optical trapping of 5-6 μm rat blood red cells in a water-heparin solution. Our results indicate the good potential of superfocused diode laser beams for applications relating to optical trapping and manipulation of microscopic objects including living biological objects with aspirations towards subsequent novel lab-on-chip configurations.

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The focusing of multimode laser diode beams is probably the most significant problem that hinders the expansion of the high-power semiconductor lasers in many spatially-demanding applications. Generally, the 'quality' of laser beams is characterized by so-called 'beam propagation parameter' M2, which is defined as the ratio of the divergence of the laser beam to that of a diffraction-limited counterpart. Therefore, M2 determines the ratio of the beam focal-spot size to that of the 'ideal' Gaussian beam focused by the same optical system. Typically, M2 takes the value of 20-50 for high-power broad-stripe laser diodes thus making the focal-spot 1-2 orders of magnitude larger than the diffraction limit. The idea of 'superfocusing' for high-M2 beams relies on a technique developed for the generation of Bessel beams from laser diodes using a cone-shaped lens (axicon). With traditional focusing of multimode radiation, different curvatures of the wavefronts of the various constituent modes lead to a shift of their focal points along the optical axis that in turn implies larger focal-spot sizes with correspondingly increased values of M2. In contrast, the generation of a Bessel-type beam with an axicon relies on 'self-interference' of each mode thus eliminating the underlying reason for an increase in the focal-spot size. For an experimental demonstration of the proposed technique, we used a fiber-coupled laser diode with M2 below 20 and an emission wavelength in ~1μm range. Utilization of the axicons with apex angle of 140deg, made by direct laser writing on a fiber tip, enabled the demonstration of an order of magnitude decrease of the focal-spot size compared to that achievable using an 'ideal' lens of unity numerical aperture. © 2014 SPIE.

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2010 Mathematics Subject Classification: 05C50.

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We provide new evidence on sea surface temperature (SST) variations and paleoceanographic/paleoenvironmental changes over the past 1500 years for the north Aegean Sea (NE Mediterranean). The reconstructions are based on multiproxy analyses, obtained from the high resolution (decadal to multi-decadal) marine record M2 retrieved from the Athos basin. Reconstructed SSTs show an increase from ca. 850 to 950 AD and from ca. 1100 to 1300 AD. A cooling phase of almost 1.5 °C is observed from ca. 1600 AD to 1700 AD. This seems to have been the starting point of a continuous SST warming trend until the end of the reconstructed period, interrupted by two prominent cooling events at 1832 ± 15 AD and 1995 ± 1 AD. Application of an adaptive Kernel smoothing suggests that the current warming in the reconstructed SSTs of the north Aegean might be unprecedented in the context of the past 1500 years. Internal variability in atmospheric/oceanic circulations systems as well as external forcing as solar radiation and volcanic activity could have affected temperature variations in the north Aegean Sea over the past 1500 years. The marked temperature drop of approximately ~2 °C at 1832 ± 15 yr AD could be related to the 1809 ?D 'unknown' and the 1815 AD Tambora volcanic eruptions. Paleoenvironmental proxy-indices of the M2 record show enhanced riverine/continental inputs in the northern Aegean after ca. 1450 AD. The paleoclimatic evidence derived from the M2 record is combined with a socio-environmental study of the history of the north Aegean region. We show that the cultivation of temperature-sensitive crops, i.e. walnut, vine and olive, co-occurred with stable and warmer temperatures, while its end coincided with a significant episode of cooler temperatures. Periods of agricultural growth in Macedonia coincide with periods of warmer and more stable SSTs, but further exploration is required in order to identify the causal links behind the observed phenomena. The Black Death likely caused major changes in agricultural activity in the north Aegean region, as reflected in the pollen data from land sites of Macedonia and the M2 proxy-reconstructions. Finally, we conclude that the early modern peaks in mountain vegetation in the Rhodope and Macedonia highlands, visible also in the M2 record, were very likely climate-driven.