Construction of a fusion gene comprising the Taka-amylase A promoter and the Escherichia coli β-glucuronidase gene and analysis of its expression in Aspergillus oryzae

Summary Northern blot analysis of glucose-grown and starch-grown mycelia of Aspergillus oryzae R11340 was conducted using the cloned Taka-amylase A (TAA) gene as a probe. The amount of mRNA homologous to the TAA gene was increased when this fungus was grown with starch as a sole carbon source. In order to analyze the induction mechanism, we inserted the Escherichia coli uidA gene encoding -glucuronidase (GUS) downstream of the TAA promoter and introduced the resultant fusion gene into the A. oryzae genome. Production of a functional GUS protein was induced by starch, but not by glucose. When the effects of various sugars on expression of the fusion gene were examined, the results suggested that the expression of the fusion gene was under control of the TAA gene promoter.     

Changes in Nine Enzyme Markers for Neurons, Glia, and Endothelial Cells in Agonal State and Huntington''s Disease Caudate Nucleus

Enzymes considered to be markers for neurons (angiotensin converting enzyme, thermolysin-like metalloendopeptidase, alanine aminopeptidase, and glutamate-oxaloacetate transaminase), glia (glutamine synthetase, pyruvate carboxylase, and beta-glucuronidase), and endothelial cells (alkaline phosphatase and plasminogen activator) were measured in caudate nucleus from 10 sudden death controls, eight agonal state controls, and 16 Huntington's disease patients. Glutamate-oxaloacetate transaminase was slightly reduced by agonal state. The four enzymes with a neuronal distribution were all correlatively reduced in Huntington's disease caudate nucleus. Glutamine synthetase activity was reduced and beta-glucuronidase mean activity increased over twofold in Huntington's disease caudate nucleus, with the two enzyme activities being inversely related. Pyruvate carboxylase was markedly affected by agonal state and was very variable in Huntington's disease caudate nucleus. The two endothelial enzymes were unaltered in Huntington's disease caudate nucleus. The findings are indicative of neuronal loss, an increased proportion of altered glia, and also of maintained vasculature in Huntington's disease caudate nucleus. Measurement of enzyme activities can help to delineate the types of cell altered in Huntington's disease.     

Serum antibodies to commensal oral and gut bacteria vary with age

Abstract Pyelonephritis is the most common urinary tract infection affecting females of all age groups. Despite concerted efforts the mechanism of renal injury in pyelonephritis is not clearly understood. In the present study we have made an attempt to characterise the mediators of inflammatory insult in an experimental model of ascending pyelonephritis. Mice infected with Escherichia coli O6:K13:H1 were sacrificed at 2, 7 and 14 days post-infection. Luminol-dependent chemiluminescence response, NADPH oxidase, acid phosphatase, β-glucuronidase and N -acetyl-β- d -glucosaminidase activities were monitored in circulating as well as renal phagocytic cells in order to determine the role of reactive oxygen species and lysosomal enzymes in genesis of renal injury. We have demonstrated that reactive oxygen species are generated at the initiation of infection and the levels increase progressively during the course of infection. While intracellular release of lysosomal enzymes was seen in all groups, extracellular release was primarily observed at 7 and 14 days post-infection only. The results indicate that while reactive oxygen species play a significant role in tissue injury during all stages of infection, lysosomal enzyme release in extracellular milieu augments tissue destruction at later stages only.     

Evolution of 1, 3, 5-trisubstituted bipyrazole scaffold based platinum(II) complexes as a biological active agent

Abstract

Square planar mononuclear platinum(II) complexes having general formula [Pt(Lⁿ)Cl₂], (where, Lⁿ?=?L^1–4) were synthesized with neutral bidentate heterocyclic 1,3,5-trisubstituted bipyrazole based ligands. The synthesized compounds were characterized by physicochemical method such as TGA, molar conductance, micro-elemental analysis and magnetic moment, and spectroscopic method such as, FT-IR, UV–vis, ¹H NMR, ¹³C NMR and mass spectrometry. Biological applications of the compounds were carried out using in vitro brine shrimp lethality bioassay, in vitro antimicrobial study against five different pathogens, and cellular level cytotoxicity against Schizosaccharomyces pombe (S. Pombe) cells. Pt(II) complexes were tested for DNA interaction activities using electronic absorption titration, viscosity measurements study, fluorescence quenching technique and molecular docking assay. Binding constants (K_b) of ligands and complexes were observed in the range of 0.23–1.07?×?10⁵?M^?1 and 0.51–3.13?×?10⁵?M^?1, respectively. Pt(II) complexes (I–IV) display an excellent binding tendency to biomolecule (DNA) and possess comparatively high binding constant (K_b) values than the ligands. The DNA binding study indicate partial intercalative mode of binding in complex-DNA. The gel electrophoresis activity was carried out to examine DNA nuclease property of pUC19 plasmid DNA.     

Synthesis of pyrimidine-2,4,6-trione derivatives: Anti-oxidant,anti-cancer, α-glucosidase, β-glucuronidase inhibition and their molecular docking studies

This paper describes a facile protocol, efficient, and environmentally benign for the synthesis a series of barbiturate acid substituted at C5 position 3a–o. The desired compounds subjected in vitro for different set of bioassays including against anti-oxidant (DPPH and super oxide scavenger assays), anti-cancer, α-glucosidase and β-glucuronidase inhibitions. Compound 3m (IC₅₀ = 22.9 ± 0.5 μM) found to be potent α-glucosidase enzyme inhibitors and showed more activity than standard acarbose (IC₅₀ = 841 ± 1.73 μM). Compound 3f (IC₅₀ = 86.9 ± 4.33 μM) found to be moderate β-Glucuronidase enzyme inhibitors and showed activity comparatively less than the standard d-saccharic acid 1,4-lactone (IC₅₀ = 45.75 ± 2.16 μM). Furthermore, in sillico investigation was carried out to investigate bonding mode of barbiturate acid derivatives.     

Transformation of the nematode-trapping fungus Arthrobotrys oligospora

The nematode-trapping fungus Arthrobotrys oligospora was transformed to hygromycin resistance using the hygromycin-B phosphotransferase gene from Escherichia coli under the control of various heterologous fungal promoters. Plasmid DNA was introduced into fungal protoplasts by polyethylene glycol/CaCl2 treatment. Transformation frequencies varied between 1-6 transformants per microgram DNA. Seven out of 13 integration events analyzed from transformants were single copy integrations, whereas the remaining were multiple and more complex integrations. The addition of restriction enzymes during transformations increased the frequency of single copy integrations. Co-transformation, using the E. coli uidA gene encoding the beta-glucuronidase reporter gene under the control of an Aspergillus nidulans promoter, occurred at frequencies of up to 63%.