Spectrophotometric analysis of Cabergoline in pharmaceutical preparations

Cabergoline (CAB) is a synthetic ergoline dopamine agonist with high affinity to D2 receptors, thus used for the treatment of Parkinson's patients and hyperprolactinemia disorders. In this study, simple, fast, reliable and validated UV-VIS and 2nd order derivative spectrophotometric methods for determination of CAB in pharmaceutical preparations were developed without any previous sample preparation step. Determination of CAB was performed at 280 nm wavelength by UV-VIS spectrophotometry and in the range of 227-232 nm by 2nd order derivative spectrophotometry. Developed both spectrophotometric methods were linear over the range of CAB concentrations from 1 to 125 µg mL-1-. The highest relative error and relative standard deviation in within-day and day-to-day study for UV-VIS spectrophotometry were found to be 1.10% and 0.63%, respectively. The relative error and relative standard deviation in within-day and day-to-day study for 2nd order derivative spectrophotometry were not higher than 1.10% and 0.70%, respectively.Colegio de Farmacéuticos de la Provincia de Buenos Aire

Mechanism for Adhesion G Protein-Coupled Receptor GPR56-Mediated RhoA Activation Induced By Collagen III Stimulation

GPR56 is a member of the adhesion G protein-coupled receptor (GPCR) family. Despite the importance of GPR56 in brain development, where mutations cause a devastating human brain malformation called bilateral frontoparietal polymicrogyria (BFPP), the signaling mechanism(s) remain largely unknown. Like many other adhesion GPCRs, GPR56 is cleaved via a GPCR autoproteolysis-inducing (GAIN) domain into N- and C-terminal fragments (GPR56N and GPR56C); however, the biological significance of this cleavage is elusive. Taking advantage of the recent identification of a GPR56 ligand and the presence of BFPP-associated mutations, we investigated the molecular mechanism of GPR56 signaling. We demonstrate that ligand binding releases GPR56N from the membrane-bound GPR56C and triggers the association of GPR56C with lipid rafts and RhoA activation. Furthermore, one of the BFPP-associated mutations, L640R, does not affect collagen III-induced lipid raft association of GPR56. Instead, it specifically abolishes collagen III-mediated RhoA activation. Together, these findings reveal a novel signaling mechanism that may apply to other members of the adhesion GPCR family

A Potent Peptidomimetic Inhibitor of Botulinum Neurotoxin Serotype A Has a Very Different Conformation than SNAP-25 Substrate

SummaryBotulinum neurotoxin serotype A is the most lethal of all known toxins. Here, we report the crystal structure, along with SAR data, of the zinc metalloprotease domain of BoNT/A bound to a potent peptidomimetic inhibitor (Ki = 41 nM) that resembles the local sequence of the SNAP-25 substrate. Surprisingly, the inhibitor adopts a helical conformation around the cleavage site, in contrast to the extended conformation of the native substrate. The backbone of the inhibitor's P1 residue displaces the putative catalytic water molecule and concomitantly interacts with the “proton shuttle” E224. This mechanism of inhibition is aided by residue contacts in the conserved S1′ pocket of the substrate binding cleft and by the induction of new hydrophobic pockets, which are not present in the apo form, especially for the P2′ residue of the inhibitor. Our inhibitor is specific for BoNT/A as it does not inhibit other BoNT serotypes or thermolysin

A Comparative Study of Drosophila and Human A-Type Lamins

Nuclear intermediate filament proteins, called lamins, form a meshwork that lines the inner surface of the nuclear envelope. Lamins contain three domains: an N-terminal head, a central rod and a C-terminal tail domain possessing an Ig-fold structural motif. Lamins are classified as either A- or B-type based on structure and expression pattern. The Drosophila genome possesses two genes encoding lamins, Lamin C and lamin Dm0, which have been designated A- and B-type, respectively, based on their expression profile and structural features. In humans, mutations in the gene encoding A-type lamins are associated with a spectrum of predominantly tissue-specific diseases known as laminopathies. Linking the disease phenotypes to cellular functions of lamins has been a major challenge. Drosophila is being used as a model system to identify the roles of lamins in development. Towards this end, we performed a comparative study of Drosophila and human A-type lamins. Analysis of transgenic flies showed that human lamins localize predictably within the Drosophila nucleus. Consistent with this finding, yeast two-hybrid data demonstrated conservation of partner-protein interactions. Drosophila lacking A-type lamin show nuclear envelope defects similar to those observed with human laminopathies. Expression of mutant forms of the A-type Drosophila lamin modeled after human disease-causing amino acid substitutions revealed an essential role for the N-terminal head and the Ig-fold in larval muscle tissue. This tissue-restricted sensitivity suggests a conserved role for lamins in muscle biology. In conclusion, we show that (1) localization of A-type lamins and protein-partner interactions are conserved between Drosophila and humans, (2) loss of the Drosophila A-type lamin causes nuclear defects and (3) muscle tissue is sensitive to the expression of mutant forms of A-type lamin modeled after those causing disease in humans. These studies provide new insights on the role of lamins in nuclear biology and support Drosophila as a model for studies of human laminopathies involving muscle dysfunction

Discovery of very high energy gamma rays from PKS 1424+240 and multiwavelength constraints on its redshift

We report the first detection of very-high-energy (VHE) gamma-ray emission above 140 GeV from PKS 1424+240, a BL Lac object with an unknown redshift. The photon spectrum above 140 GeV measured by VERITAS is well described by a power law with a photon index of 3.8 +- 0.5_stat +- 0.3_syst and a flux normalization at 200 GeV of (5.1 +- 0.9_stat +- 0.5_syst) x 10^{-11} TeV^-1 cm^-2 s^-1, where stat and syst denote the statistical and systematical uncertainty, respectively. The VHE flux is steady over the observation period between MJD 54881 and 55003 (2009 February 19 to June 21). Flux variability is also not observed in contemporaneous high energy observations with the Fermi Large Area Telescope (LAT). Contemporaneous X-ray and optical data were also obtained from the Swift XRT and MDM observatory, respectively. The broadband spectral energy distribution (SED) is well described by a one-zone synchrotron self-Compton (SSC) model favoring a redshift of less than 0.1. Using the photon index measured with Fermi in combination with recent extragalactic background light (EBL) absorption models it can be concluded from the VERITAS data that the redshift of PKS 1424+240 is less than 0.66.Comment: accepted for publication, Ap

VERITAS Search for VHE Gamma-ray Emission from Dwarf Spheroidal Galaxies

Indirect dark matter searches with ground-based gamma-ray observatories provide an alternative for identifying the particle nature of dark matter that is complementary to that of direct search or accelerator production experiments. We present the results of observations of the dwarf spheroidal galaxies Draco, Ursa Minor, Bootes 1, and Willman 1 conducted by VERITAS. These galaxies are nearby dark matter dominated objects located at a typical distance of several tens of kiloparsecs for which there are good measurements of the dark matter density profile from stellar velocity measurements. Since the conventional astrophysical background of very high energy gamma rays from these objects appears to be negligible, they are good targets to search for the secondary gamma-ray photons produced by interacting or decaying dark matter particles. No significant gamma-ray flux above 200 GeV was detected from these four dwarf galaxies for a typical exposure of ~20 hours. The 95% confidence upper limits on the integral gamma-ray flux are in the range 0.4-2.2x10^-12 photons cm^-2s^-1. We interpret this limiting flux in the context of pair annihilation of weakly interacting massive particles and derive constraints on the thermally averaged product of the total self-annihilation cross section and the relative velocity of the WIMPs. The limits are obtained under conservative assumptions regarding the dark matter distribution in dwarf galaxies and are approximately three orders of magnitude above the generic theoretical prediction for WIMPs in the minimal supersymmetric standard model framework. However significant uncertainty exists in the dark matter distribution as well as the neutralino cross sections which under favorable assumptions could further lower the limits.Comment: 21 pages, 2 figures, updated to reflect version published in ApJ. NOTE: M.D. Wood added as autho

The Interdomain Linker of AAV-2 Rep68 Is an Integral Part of Its Oligomerization Domain: Role of a Conserved SF3 Helicase Residue in Oligomerization

The four Rep proteins of adeno-associated virus (AAV) orchestrate all aspects of its viral life cycle, including transcription regulation, DNA replication, virus assembly, and site-specific integration of the viral genome into the human chromosome 19. All Rep proteins share a central SF3 superfamily helicase domain. In other SF3 members this domain is sufficient to induce oligomerization. However, the helicase domain in AAV Rep proteins (i.e. Rep40/Rep52) as shown by its monomeric characteristic, is not able to mediate stable oligomerization. This observation led us to hypothesize the existence of an as yet undefined structural determinant that regulates Rep oligomerization. In this document, we described a detailed structural comparison between the helicase domains of AAV-2 Rep proteins and those of the other SF3 members. This analysis shows a major structural difference residing in the small oligomerization sub-domain (OD) of Rep helicase domain. In addition, secondary structure prediction of the linker connecting the helicase domain to the origin-binding domain (OBD) indicates the potential to form α-helices. We demonstrate that mutant Rep40 constructs containing different lengths of the linker are able to form dimers, and in the presence of ATP/ADP, larger oligomers. We further identified an aromatic linker residue (Y224) that is critical for oligomerization, establishing it as a conserved signature motif in SF3 helicases. Mutation of this residue critically affects oligomerization as well as completely abolishes the ability to produce infectious virus. Taken together, our data support a model where the linker residues preceding the helicase domain fold into an α-helix that becomes an integral part of the helicase domain and is critical for the oligomerization and function of Rep68/78 proteins through cooperative interaction with the OBD and helicase domains

Discovery of VHE γ\gamma-ray emission from the SNR G54.1+0.3

We report the discovery of very high energy gamma-ray emission from the direction of the SNR G54.1+0.3 using the VERITAS ground-based gamma-ray observatory. The TeV signal has an overall significance of 6.8$\sigma$ and appears point-like given the 5$^{arcminute}$ resolution of the instrument. The integral flux above 1 TeV is 2.5% of the Crab Nebula flux and significant emission is measured between 250 GeV and 4 TeV, well described by a power-law energy spectrum dN/dE $\sim$ E$^{-\Gamma}$ with a photon index $\Gamma= 2.39\pm0.23_{stat}\pm0.30_{sys}$. We find no evidence of time variability among observations spanning almost two years. Based on the location, the morphology, the measured spectrum, the lack of variability and a comparison with similar systems previously detected in the TeV band, the most likely counterpart of this new VHE gamma-ray source is the PWN in the SNR G54.1+0.3. The measured X-ray to VHE gamma-ray luminosity ratio is the lowest among all the nebulae supposedly driven by young rotation-powered pulsars, which could indicate a particle-dominated PWN.Comment: 5 pages, 2 figure, Latex, emulateapj style, accepted by the Astrophysical Journal Letter