Apigenin 4′-O-β-d-glucoside 7-O-β-d)- glucuronide: The copigment in the blue pigment of Centaurea cyanus

The copigment present in the crystalline blue pigment isolated from Blue Boy cornflowers (Centaurea cyanus L.) was identified as apigenin 4′-O-β-glucoside 7-O-β-d-glucuronide. The NMR spectra of aryl glucuronides are discussed.     

6-Phosphofructo-2-kinase and D-fructose-2,6-bisphosphatase activities in mung bean seedlings

6-Phosphofructo-2-kinase (ATP: D-fructose-6-phosphate-2-phosphotransferase) and D-fructose-2,6-bisphosphatase activities have been found in extracts prepared from etiolated mung bean seedlings. The activity of 6-phosphofructo-2-kinase exhibits a sigmoidal shape in response to changes in concentrations of both substrates, D-fructose 6-phosphate and ATP (S_0.5 values of 1.8 and 1.2 mM, respectively). Inorganic orthophosphate (P_i) has a strong stimulating effect on the 2-kinase activity (A_0.5 at about 2 mM), moderately increasing the V_max and modifying the response into hyperbolic curves with K_m values of 0.4 and 0.2 mM for fructose 6-phosphate and ATP, respectively. 3-Phosphoglycerate (I_0.5 about 0.15 mM) partially inhibited the kinase activity by counteracting the P_i activation. In contrast, the activity of D-fructose-2,6-bisphosphatase (K_m 0.38 mM) is strongly inhibited by P_i (I_0.5 0.8 mM) lowering its affinity to fructose-2,6-P₂ (K_m 1.4 mM). 3-Phosphoglycerate activites the enzyme (A_0.5 at about 0.3 mM) without causing a significant change in its K_m for fructose-2,6-P₂. The activities of both of these enzymes in relationship to the metabolic role of D-fructose 2,6-bisphosphate in the germinating seed is discussed.     

PatU3 plays a central role in coordinating cell division and differentiation in pattern formation of filamentous cyanobacterium Nostoc sp. PCC 7120

Spatial periodic signal for cell differentiation in some multicellular organisms is generated according to Turing’s principle for pattern formation. How a dividing cell responds to the signal of differentiation is addressed with the filamentous cyanobacterium Nostoc sp. PCC 7120, which forms the patterned distribution of heterocysts. We show that differentiation of a dividing cell was delayed until its division was completed and only one daughter cell became heterocyst. A mutant of patU3, which ...     

Different kinetics of the regulation of respiration in permeabilized cardiomyocytes and in HL-1 cardiac cells: Importance of cell structure/organization for respiration regulation

The aim of this study was to investigate the mechanism of cellular regulation of mitochondrial respiration in permeabilized cardiac cells with clearly different structural organization: (i) in isolated rat cardiomyocytes with very regular mitochondrial arrangement, (ii) in HL-1 cells from mouse heart, and (iii) in non-beating (NB HL-1 cells) without sarcomeres with irregular and dynamic filamentous mitochondrial network. We found striking differences in the kinetics of respiration regulation by exogenous ADP between these cells: the apparent Km for exogenous ADP was by more than order of magnitude (14 times) lower in the permeabilized non-beating NB HL-1 cells without sarcomeres (25 ± 4 μM) and seven times lower in normally cultured HL-1 cells (47 ± 15 μM) than in permeabilized primary cardiomyocytes (360 ± 51 μM). In the latter cells, treatment with trypsin resulted in dramatic changes in intracellular structure that were associated with 3-fold decrease in apparent Km for ADP in regulation of respiration. In contrast to permeabilized cardiomyocytes, in NB HL-1 cells creatine kinase activity was low and the endogenous ADP fluxes from MgATPases recorded spectrophotometrically by the coupled enzyme assay were not reduced after activation of mitochondrial oxidative phosphorylation by the addition of mitochondrial substrates, showing the absence of ADP channelling in the NB HL-1 cells. While in the permeabilized cardiomyocytes creatine strongly activated mitochondrial respiration even in the presence of powerful competing pyruvate kinase-phosphoenolpyruvate system, in the NB HL-1 cells the stimulatory effect of creatine was not significant. The results of this study show that in normal adult cardiomyocytes and HL-1 cells intracellular local restrictions of diffusion of adenine nucleotides and metabolic feedback regulation of respiration via phosphotransfer networks are different, most probably related to differences in structural organization of these cells.     

A C-NMR study on the influxes into the tricarboxylic acid cycle of a renal epithelial cell line, LLC-PK1/Cl4: the metabolism of [2-C]glycine, l-[3-C]alanine and l-[3-C]aspartic acid in renal epithelial cells

Perchloric acid extracts of LLC-PK₁/Cl₄ cells, a renal epithelial cell line, incubated with either [2-¹³C]glycine l-[3-¹³C]alanine, or d,l-[3-¹³C]aspartic acid were investigated by ¹³C-NMR spectroscopy. All amino acids, except labelled glycine, gave rise to glycolytic products and tricarboxylic acid cycle (TCA) intermediates. For the first time we also observed activity of γ-glutamyltransferase activity and glutathione synthetase activity in LLC-PK₁ cells, as is evident from enrichment of reduced glutathione. Time courseS showed that only 6% of the labelled glycine was utilized in 30 min, whereas 31% of l-alanine and 60% of l-aspartic acid was utilized during the same period. ¹³C-NMR was also shown to be a useful tool for the determination of amino acid uptake in LLC-PK₁ cells. These uptake experiments indicated that glycine alanine and aspartic acid are transported into Cl₄ cells via a sodium-dependent process. From the relative enrichment of the glutamate carbons, we calculated the activity of pyruvate dehydrogenase to be about 61% of when labelled l-alanine was the only carbon source for LLC-PK₁/Cl₄ cells. Experiments with labelled d,l-aspartic, however, showed that about 40% of C-3-enriched oxaloacetate (arising from a de-amination of aspartic acid) reached the pyruvate pool.