Reaction of liver pyruvate kinase with sulfhydryl reagents: effect on its activity

Homogeneous liver pyruvate kinase was reacted with different sulfhydryl reagents, which included o-iodosobenzoate, 5',5'-dithiobis(2-nitrobenzoic acid) and N-ethylmaleimide. Activity determinations of the treated enzyme made with and without Fru(1,6)P2 indicate that the protein contains two sulfhydryl groups per subunit important to its properties, one more accessible than the other. Fru(1,6)P2 added to mixtures prevented loss of activity obtained with o-iodosobenzoate and 5',5'-dithiobis(2-nitrobenzoic acid). It appears that Fru(1,6)P2 does not interfere with the reaction of the reagent with the sulfhydryl group, but prevents an ensuing conformational change, which leads to changes in the enzyme's properties.     

A comparison of immunomagnetic and surface adhesion immunofluorescent techniques for the rapid detection of Listeria monocytogenes and Listeria innocua in meat

an immunomagnetic immunofluorescent method was investigated for the rapid detection of Listeria monocytogenes and Listeria innouca . This technique involved enrichment of the suspect sample at 30°C overnight. Listeria monocytogenes cells were isolated from the enriched sample using immunomagnetic separation and Listeria were subsequently visualized using an immunofluorescent microscopy technique. This technique was used in the detection of Listeria cells from pure culture, inoculated beef mince samples and naturally contaminated retail beef mince samples. A detection level of approximately 1×10³ cfu ml⁻¹ was achieved. When compared with traditional detection methods no false negatives or positives were recorded for L. monocytogenes or L. innocua . The immunomagnetic immunofluorescent technique had a detection level similar to a previously described surface adhesion immunofluorescent technique. Isolation of the Listeria cells by surface adhesion involved dipping a membrane attached to a microscope slide into the enriched sample for 10 min. This was quicker and simpler to perform than the immunomagnetic separation technique which took 2 h to carry out.     

Inhibition of Mitochondrial Complex II by the Anticancer Agent Lonidamine

The antitumor agent lonidamine (LND; 1-(2,4-dichlorobenzyl)-1H-indazole-3-carboxylic acid) is known to interfere with energy-yielding processes in cancer cells. However, the effect of LND on central energy metabolism has never been fully characterized. In this study, we report that a significant amount of succinate is accumulated in LND-treated cells. LND inhibits the formation of fumarate and malate and suppresses succinate-induced respiration of isolated mitochondria. Utilizing biochemical assays, we determined that LND inhibits the succinate-ubiquinone reductase activity of respiratory complex II without fully blocking succinate dehydrogenase activity. LND also induces cellular reactive oxygen species through complex II, which reduced the viability of the DB-1 melanoma cell line. The ability of LND to promote cell death was potentiated by its suppression of the pentose phosphate pathway, which resulted in inhibition of NADPH and glutathione generation. Using stable isotope tracers in combination with isotopologue analysis, we showed that LND increased glutaminolysis but decreased reductive carboxylation of glutamine-derived α-ketoglutarate. Our findings on the previously uncharacterized effects of LND may provide potential combinational therapeutic approaches for targeting cancer metabolism.     

Cryptorchidism and Infertility in Rats with Targeted Disruption of the Adamts16 Locus

A Disintegrin And Metalloproteinase with ThromboSpondin motifs16 (ADAMTS-16) is a member of a family of metalloproteinases. Using a novel zinc-finger nuclease based gene-edited rat model harboring a targeted mutation of the Adamts16 locus, we previously reported this gene to be linked to blood pressure regulation. Here we document our observation with this model that Adamts16 is essential for normal development of the testis. Absence of Adamts16 in the homozygous Adamts16^mutant males resulted in cryptorchidism and male sterility. Heterozygous Adamts16^mutant males were normal, indicating that this is a recessive trait. Testes of homozygous Adamts16^mutant males were significantly smaller with significant histological changes associated with the lack of sperm production. Temporal histological assessments of the testis demonstrated that the seminiferous tubules did not support active spermatogenesis, but progressively lost germ cells, accumulated vacuoles and did not have any sperm. These observations, taken together with our previous report of renal abnormalities observed with the same Adamts16^mutant rats, suggest an important mechanistic link between Adamts16 and the functioning of the male genitourinary system.     

Synthesis of 5-Fluoro-1-(3-O-benzoyl-4, 6-dideoxy- β-L-GLYCERO-hex-3-enopyranos-2-ulosyl)uracil

Abstract

Synthesis of the title compound, an unsaturated ketohexo-pyranosyl nucleoside of 5-fluorouracil is reported. It was prepared by oxidation of the corresponding dibenzoylhexopyranosyl nucleoside with pyridinium dichromate/molecular sieves system.     

Fire and grazing impacts on silica production and storage in grass dominated ecosystems

Grassland ecosystems are an important terrestrial component of the global biogeochemical silicon cycle. Although the structure and ecological functioning of grasslands are strongly influenced by fire and grazing, the role of these key ecological drivers in the production and storage of silicon represents a significant knowledge gap, particularly since they are being altered worldwide by human activities. We evaluated the effects of fire and grazing on the range and variability of plant derived biogenic silica stored in plant biomass and soils by sampling plants and soils from long-term experimental plots with known fire and grazing histories. Overall, plants and soils from grazed sites in the South African ecosystems had up to 76 and 54% greater biogenic silica totals (kg ha^?1), respectively, than grazed North American sites. In North American soils, the combination of grazing and annual fire resulted in the greatest abundance of biogenic silica, whereas South African soils had the highest biogenic silica content where grazed regardless of burn frequency. These results as well as those that show greater Si concentrations in grazed South African plants indicate that South African plants and soils responded somewhat differently to fire and grazing with respect to silicon cycling, which may be linked to differences in the evolutionary history and in the grazer diversity and grazing intensity of these ecosystems. We conclude that although fire and grazing (as interactive and/or independent factors) do not affect the concentration of Si taken up by plants, they do promote increased silicon storage in aboveground biomass and soil as a result of directly affecting other site factors such as aboveground net primary productivity. Therefore, as management practices, fire and grazing have important implications for assessing global change impacts on the terrestrial biogeochemical cycling of silicon.