Genome Sequence of Acinetobacter sp. Strain HA, Isolated from the Gut of the Polyphagous Insect Pest Helicoverpa armigera

In this study, Acinetobacter sp. strain HA was isolated from the midgut of a fifth-instar larva of Helicoverpa armigera. Here, we report the draft genome sequence (3,125,085 bp) of this strain that consists of 102 contigs, 2,911 predicted coding sequences, and a G+C content of 41%.     

Synthesis and in vitro evaluation of [18F](R)-FEPAQ: a potential PET ligand for VEGFR2

Synthesis and in vitro evaluation of [(18)F](R)-N-(4-bromo-2-fluorophenyl)-7-((1-(2-fluoroethyl)piperidin-3-yl)methoxy)-6-methoxyquinazolin-4-amine ((R)-[(18)F]FEPAQ or [(18)F]1), a potential imaging agent for the VEGFR2, using phosphor image autoradiography are described. Synthesis of 2, the desfluoroethyl precursor for (R)-FEPAQ was achieved from t-butyl 3-(hydroxymethyl)piperidine-1-carboxylate (3) in five steps and in 50% yield. [(18)F]1 was synthesized by reaction of sodium salt of compound 2 with [(18)F]fluoroethyl tosylate in DMSO. The yield of [(18)F]1 was 20% (EOS based on [(18)F]F(-)) with >99% radiochemical purity and specific activity of 1-2 Ci/μmol (n=10). The total synthesis time was 75 min. The radiotracer selectively labeled VEGFR2 in slide-mounted sections of human brain and higher binding was found in surgically removed human glioblastoma sections as demonstrated by in vitro phosphor imager studies. These findings suggest [(18)F]1 may be a promising radiotracer for imaging VEGFR2 in brain using PET.     

Iron-mediated retinal degeneration in haemojuvelin-knockout mice

Haemochromatosis is a genetic disorder of iron overload resulting from loss-of-function mutations in genes coding for the iron-regulatory proteins HFE (human leucocyte antigen-like protein involved in iron homoeostasis), transferrin receptor 2, ferroportin, hepcidin and HJV (haemojuvelin). Recent studies have established the expression of all of the five genes in the retina, indicating their importance in retinal iron homoeostasis. Previously, we demonstrated that HJV is expressed in RPE (retinal pigment epithelium), the outer and inner nuclear layers and the ganglion cell layer. In the present paper, we report on the consequences of Hjv deletion on the retina in mice. Hjv-/- mice at ≥18 months of age had increased iron accumulation in the retina with marked morphological damage compared with age-matched controls; these changes were not found in younger mice. The retinal phenotype in Hjv-/- mice included hyperplasia of RPE. We isolated RPE cells from wild-type and Hjv-/- mice and examined their growth patterns. Hjv-/- RPE cells were less senescent and exhibited a hyperproliferative phenotype. Hjv-/- RPE cells also showed up-regulation of Slc7a11 (solute carrier family 7 member 11 gene), which encodes the 'transporter proper' subunit xCT in the heterodimeric amino acid transporter xCT/4F2hc (cystine/glutamate exchanger). BMP6 (bone morphogenetic protein 6) could not induce hepcidin expression in Hjv-/- RPE cells, confirming that retinal cells require HJV for induction of hepcidin via BMP6 signalling. HJV is a glycosylphosphatidylinositol-anchored protein, and the membrane-associated HJV is necessary for BMP6-mediated activation of hepcidin promoter in RPE cells. Taken together, these results confirm the biological importance of HJV in the regulation of iron homoeostasis in the retina and in RPE.     

Role of the Met3534-Ala4271 region of the ryanodine receptor in the regulation of Ca2+ release induced by calmodulin binding domain peptide

CaMBP, a peptide corresponding to the 3614-3643 calmodulin (CaM) binding region of the ryanodine receptor (RyR1), is known to activate RyR1 Ca²⁺ channel. To analyze the mechanism of channel regulation by the CaMBP-RyR1 interaction, we investigated a), CaMBP binding to RyR1, b), induced local conformational changes in the CaMBP binding region of RyR1 using the fluorescent conformational probe badan attached to CaMBP (CaMBP-badan), and c), effects of “a” and “b” on SR Ca²⁺ release. We also monitored the interaction of CaMBP-badan with CaM and a peptide corresponding to the Met³⁵³⁴-Ala⁴²⁷¹ region of RyR1 (R3534-4271) as a control. At lower peptide concentrations (≤15 μM), CaMBP binding to RyR1 increased the intensity of badan fluorescence emission at a shorter wavelength (the state resembling CaMBP-badan/Ca-CaM) and induced Ca²⁺ release. Further increase in CaMBP concentration (up to ∼50 μM) produced more binding of CaMBP accompanied by further increase in the badan fluorescence emission but at a longer wavelength (the state resembling CaMBP-badan/apo-CaM) and inhibited Ca²⁺ release. Binding of CaMBP-badan to R3534-4271 increased the intensity of badan fluorescence, showing the similar concentration-dependent red-shift of the emission maximum. It is proposed that CaMBP interacts with two classes of binding sites located in the Met³⁵³⁴-Ala⁴²⁷¹ region of RyR1, which activate and inhibit the Ca²⁺ channel, respectively.     

Pd(II)-catalysed and Hg(II)-co-catalysed oxidation of D-glucose and D-fructose by N-bromoacetamide in the presence of perchloric acid: a kinetic and mechanistic study

The kinetics of Pd(II)-catalysed and Hg(II)-co-catalysed oxidation of D-glucose (Glc) and D-fructose (Fru) by N-bromoacetamide (NBA) in the presence of perchloric acid using mercury(II) acetate as a scavenger for Br- ions have been studied. The results show first-order kinetics with respect to NBA at low concentrations, tending to zero order at high concentrations. First-order kinetics with respect to Pd(II) and inverse fractional order in Cl- ions throughout their variation have also been noted. The observed direct proportionality between the first-order rate constant (k1) and the reducing sugar concentration shows departure from the straight line only at very higher concentration of sugar. Addition of acetamide (NHA) decreases the first-order rate constant while the oxidation rate is not influenced by the change in the ionic strength (mu) of the medium. Variation of [Hg(OAc)2] shows a positive effect on the rate of reaction. The observed negative effect in H+ at lower concentrations tends to an insignificant effect at its higher concentrations. The first-order rate constant decreases with an increase in the dielectric constant of the medium. The various activation parameters have also been evaluated. The products of the reactions were identified as arabinonic acid and formic acid for both the hexoses. A plausible mechanism involving HOBr as the reactive oxidising species, Hg(II) as co-catalyst, and [PdCl3.S]-1 as the reactive Pd(II)-sugar complex in the rate-controlling step is proposed.     

Mini-alphaB-crystallin: a functional element of alphaB-crystallin with chaperone-like activity

Alpha-crystallin is a member of the family of small heat-shock proteins (sHSP) and is composed of two subunits, alphaA-crystallin and alphaB-crystallin, which exhibit molecular chaperone-like properties. In a previous study, we found that residues 70-88 in alphaA-crystallin can function like a molecular chaperone by preventing the aggregation and precipitation of denaturing substrate proteins [Sharma, K. K., et al. (2000) J. Biol. Chem. 275, 3767-3771]. In this study, we show that the complementary sequence in alphaB-crystallin, residues 73-92 (DRFSVNLDVKHFSPEELKVK), is the functional chaperone site of alphaB-crystallin. Like the mini-alphaA-crystallin chaperone, the mini-alphaB-crystallin chaperone interacts with 1,1'-bi(4-anilino) naphthalene-5,5'-disulphonic acid (bis-ANS) and also possesses significant beta-sheet and random coil structure. Deletion of four residues (DRFS) from the N-terminus or deletion of C-terminus LKVK residues from the 73-92 peptide abolishes the chaperone-like activity against denaturing alcohol dehydrogenase. However, removal of DRFS or HFSPEELKVK is necessary to completely abolish the antiaggregation property of the peptide in insulin reduction assay. Substitution of Asp at a site corresponding to D80 in alphaB-crystallin with d-Asp or beta-Asp results in a significant loss of chaperone-like activity. Kynurenine modification of His in the peptide abolishes the antiaggregation property of the mini-chaperone. These data suggest that the 73-92 region in alphaB-crystallin is one of the substrate binding sites during chaperone activity.     

A novel low molecular weight phospholipase D from Streptomyces sp. CS684

With the aim of isolating economically viable enzymes from a microbial source, a novel phospholipase D (PLD) was purified from Streptomyces sp. CS684 (PLD(684)). PLD(684) had molecular weight of 29 kDa, which makes it the second smallest PLD reported so far. The enzyme activity was optimum at pH 6 and 45 degrees C, and enhanced by various detergents. It was stable from pH 7 to 9 and at or below 45 degrees C when assayed after 40 h and 2h, respectively. The K(m) and V(max) values for phosphatidylcholine were 1.16 mM and 1453.74 micromol min(-1)mg(-1), respectively. It catalyzed the transphosphatidylation of glycerol, but not that of l-serine, myo-inositol or ethanolamine. Low molecular weight PLD(684) with transphosphatidylation activity may be utilized in the industrial production of rare and commercially important phospholipids.     

Influenza A: From highly pathogenic H5N1 to pandemic 2009 H1N1. Epidemiology and clinical features

The last decade has seen the emergence of two new influenza A subtypes and they have become a cause of concern for the global community. These are the highly pathogenic H5N1 influenza A virus (H5N1) and the Pandemic 2009 influenza H1N1 virus. Since 2003 the H5N1 virus has caused widespread disease and death in poultry, mainly in south East Asia and Africa. In humans the number of cases infected with this virus is few but the mortality has been about 60%. Most patients have presented with severe pneumonia and acute respiratory distress syndrome. The second influenza virus, the pandemic H1N1 2009, emerged in Mexico in March this year. This virus acquired the ability for sustained human to human spread and within a few months spread throughout the world and infected over 4 lakh individuals. The symptoms of infection with this virus are similar to seasonal influenza but it currently affecting younger individuals more often. Fortunately the mortality has been low. Both these new influenza viruses are currently circulating and have different clinical and epidemiological characteristics.