Surge ammonium uptake in macroalgae from a coral atoll

Surge (non-linear) uptake of ammonium, measured by incorporation of ¹⁵N, was investigated in three species of macroalgae (Ulva lactuca Linnaeus (Chlorophyta), Soliera robusta (Greville) Kylin (Rhodophyta) and Dictyota dichotoma (Hudson) Lamouroux (Phaeophyta)) from Kavaratti atoll (Lakshadweep, India). Addition of ammonium (up to 20 μmol L^− 1) led to pronounced uptake within 4-6 min, with the amount of ammonium taken up during surge phase (< 4 min) accounting for from about half to 10 times that taken up during the remaining period of incubation (5-30 min). Amount of ammonium taken up during surge related linearly to the concentration of ammonium given. Surge uptake in the dark was also substantial, averaging 80% of that in light. Capability for rapid uptake of pulses of ammonium released by heterotrophs during the day or night could thus be an important mechanism of survival and proliferation of macroalgae in the N-impoverished atoll waters.     

UDP-Gal: GlcNAc-R β1,4-galactosyltransferase—a target enzyme for drug design. Acceptor specificity and inhibition of the enzyme

Galactosyltransferases are important enzymes for the extension of the glycan chains of glycoproteins and glycolipids, and play critical roles in cell surface functions and in the immune system. In this work, the acceptor specificity and several inhibitors of bovine β1,4-Gal-transferase T1 (β4GalT, EC 2.4.1.90) were studied. Series of analogs of N-acetylglucosamine (GlcNAc) and GlcNAc-carrying glycopeptides were synthesized as acceptor substrates. Modifications were made at the 3-, 4- and 6-positions of the sugar ring of the acceptor, in the nature of the glycosidic linkage, in the aglycone moiety and in the 2-acetamido group. The acceptor specificity studies showed that the 4-hydroxyl group of the sugar ring was essential for β4GalT activity, but that the 3-hydroxyl could be replaced by an electronegative group. Compounds having the anomeric β-configuration were more active than those having the α-configuration, and O-, S- and C-glycosyl compounds were all active as substrates. The aglycone was a major determinant for the rate of Gal-transfer. Derivatives containing a 2-naphthyl aglycone were inactive as substrates although quinolinyl groups supported activity. Several compounds having a bicyclic structure as the aglycone were found to bind to the enzyme and inhibited the transfer of Gal to control substrates. The best small hydrophobic GlcNAc-analog inhibitor was found to be 1-thio-N-butyrylGlcNβ-(2-naphthyl) with a K_i of 0.01 mM. These studies help to delineate β4GalT–substrate interactions and will aid in the development of biologically applicable inhibitors of the enzyme.     

Preoperative diagnosis of renal angiomyolipoma with fine needle aspiration cytology: a report of 3 cases

BACKGROUND: Angiomyolipoma is a rare benign neoplasm, commonly involving the kidney. Preoperative diagnosis on fine needle aspiration cytology can confirm the diagnosis, reducing the chances of unnecessary surgery. CASES: We studied cases of renal angiomyolipoma (AML) presenting within 1 year with palpable renal masses confirmed on ultrasound to be of renal origin. Fine needle aspiration cytology (FNAC) smears showed a few cohesive syncytial fragments with adipocytes, spindle cells and isolated single cells with foamy cytoplasm; a diagnosis of AML was made. The diagnosis was confirmed on histopathology, which showed mature adipose tissue, tortuous and thick-walled blood vessels lacking elastic tissue lamina and bundles of smooth muscles that seemed to emanate from the blood vessels. CONCLUSION: A preoperative diagnosis of renal AML is of great importance for correct management. In fact, when the lesion is small and asymptomatic, a conservative approach may be considered. Partial nephrectomy is possible with masses < 5 cm. Preoperative FNAC along with computed tomographic findings and immunocytochemical analysis of FNAC smears with HNB-45 can confirm the diagnosis of AML, thus preventing unindicated nephrectomy.     

The transferable tail: fusion of the N-terminal acidic extension of heparin cofactor II to alpha1-proteinase inhibitor M358R specifically increases the rate of thrombin inhibition

The conversion of the reactive center bond of the serpin alpha1-proteinase inhibitor (alpha1-PI, also known as alpha1-antitrypsin) from Met-Ser to Arg-Ser decreases the rate at which it inhibits neutrophil elastase and endows it with the ability to inhibit thrombin and activated protein C (APC). Another serpin, heparin cofactor II (HCII), contains a unique N-terminal extension that binds thrombin exosite 1. We fused residues 1-75 of HCII to the N-terminus of alpha1-PI M358R, forming an HCII-alpha1-PI chimera (HAPI M358R). It inhibited alpha-thrombin 21-fold faster than alpha1-PI M358R, with second-order rate constants of 2.3 x 10(8) M(-1) min(-1) versus 1.1 x 10(7) M(-1) min(-1), respectively. When gammaT-thrombin, which lacks an intact exosite 1, was substituted for alpha-thrombin, the kinetic advantage of HAPI M358R over alpha1-PI M358R was reduced to 9-fold, whereas APC and trypsin, proteases lacking exosite 1-like regions, were inhibited only 1.3- and 2-fold more rapidly by HAPI M358R than by alpha1-PI M358R, respectively. Maximal enhancement of alpha1-PI M358R activity required the acidic residues found between HCII residues 55 and 75, because no enhancement was observed either by fusion of residues 1-54 alone or by fusion of a mutated HCII acidic extension in which all Glu and Asp residues between positions 55 and 75 were neutralized by mutation. Fusing residues 55-75 to alpha1-PI M358R yielded a relative rate enhancement of only 6-fold, suggesting a need for the full tail region to achieve maximal enhancement. Our results suggest that transfer of the N-terminal acidic extension of HCII to alpha1-PI M358R enhanced its inhibition of thrombin by conferring the ability to bind exosite 1 on HAPI M358R. This enhancement may aid in efforts to tailor this inhibitor for therapeutic use.     

Extracellular nucleotide signaling in adult neural stem cells: synergism with growth factor-mediated cellular proliferation

We have previously shown that the extracellular nucleoside triphosphate-hydrolyzing enzyme NTPDase2 is highly expressed in situ by stem/progenitor cells of the two neurogenic regions of the adult murine brain: the subventricular zone (type B cells) and the dentate gyrus of the hippocampus (residual radial glia). We explored the possibility that adult multipotent neural stem cells express nucleotide receptors and investigated their functional properties in vitro. Neurospheres cultured from the adult mouse SVZ in the presence of epidermal growth factor and fibroblast growth factor 2 expressed the ecto-nucleotidases NTPDase2 and the tissue non-specific isoform of alkaline phosphatase, hydrolyzing extracellular ATP to adenosine. ATP, ADP and, to a lesser extent, UTP evoked rapid Ca(2+) transients in neurospheres that were exclusively mediated by the metabotropic P2Y(1) and P2Y(2) nucleotide receptors. In addition, agonists of these receptors and low concentrations of adenosine augmented cell proliferation in the presence of growth factors. Neurosphere cell proliferation was attenuated after application of the P2Y(1)-receptor antagonist MRS2179 and in neurospheres from P2Y(1)-receptor knockout mice. In situ hybridization identified P2Y(1)-receptor mRNA in clusters of SVZ cells. Our results infer nucleotide receptor-mediated synergism that augments growth factor-mediated cell proliferation. Together with the in situ data, this supports the notion that extracellular nucleotides contribute to the control of adult neurogenesis.     

Relationship of physicochemical properties of rhizosphere soils with native population of mineral phosphate solubilizing fungi

Phosphate solubilizing fungi (PSF) were enumerated in 78 rhizosphere soil samples collected from various sites of Bhavnagar district with agro climatic zone of hot, semi-arid region of Gujarat. 81% of samples were inhabited with indigenous PSF; the count varied ranging from 0.01 to 90 × 10³ PSF g⁻¹. PSF population in the rhizosphere soils were maximal at pH 7.4–8.4, electrical conductivity (EC) 1.2 dS m⁻¹ wherein PSF population decreased with a rise in EC. Optimal moisture content ranged ≤10–30% where the abundance of PSF always decreased with increase in moisture levels while suitable organic matter content ranged from ≤0.5–4.5%. Available P range 100–300 mg kg⁻¹ was the most suitable for occurrence of PSF as being noticed from number of soil samples harboring them in rhizosphere soils. Amongst the various rhizosphere soil properties studied, a highly signifi cant positive correlation was established between PSF and soil available P as well as pH. A signifi cant positive association observed between total fungal population and organic matter as well as soil available P. Both abundance and number of PSF were more pronounced in descending order in plant covers: oilseeds, flowers, orchards, vegetables, pulses and cereals.     

Scale up production of isoflavonoids in cell suspension cultures of Pueraria tuberosa grown in shake flasks and bioreactor

Cell cultures of Pueraria tuberosa were grown in vessels of different sizes and 2L stirred tank bioreactor containing modified MS medium with morphactin (0.1 mg l^?1) and 2iP (5.0 mg l^?1) and 20% inoculum. Stable growth and total isoflavonoid yield of 76.6 mg l^?1 were recorded in the cultures during scale up. This was in concordance with the persistent yield of the individual isoflavonoids regardless of the vessel size.