Recombinant Gaussia luciferase with a reactive cysteine residue for chemical conjugation: Expression, purification and its application for bioluminescent immunoassays

The mutated recombinant Gaussia luciferase (hgGLase) having the hinge sequence with a reactive cysteine residue at the carboxyl terminal region was purified from Escherichia coli cells by nickel-chelate affinity chromatography and hydrophobic chromatography. The biotinylated hgGLase (Biotin-hgGLase) was prepared by chemical conjugation with a maleimide activated biotin and apply to bioluminescent immunoassay. In the streptavidin and biotin complex system using Biotin-hgGLase, the measurable range of α-fetoprotein as a model analyte was 0.02–100 ng/ml with the coefficient of variation between 2.5% and 5.2%. The sensitivity of Biotin-hgGLase was similar to that by using the detection system of aequorin, alkaline phosophatase and horseradish peroxidase as a label enzyme.     

NaCl胁迫对嫁接黄瓜膜脂过氧化、渗透调节物质含量及光合特性的影响

以日本引进的耐盐品种‘帝王新土佐’南瓜为砧木,以‘津研4号’黄瓜品种为接穗,研究了NaCl胁迫对黄瓜嫁接植株和自根植株的膜脂过氧化、渗透调节物质含量及光合特性的影响。结果表明:在NaCl胁迫下,黄瓜嫁接植株叶片超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)活性均显著高于自根植株,O·2-产生速率及丙二醛(MDA)含量显著低于自根植株,嫁接植株膜脂过氧化轻于自根植株;嫁接植株叶片游离脯氨酸、可溶性糖、可溶性蛋白质等渗透物质含量均显著高于自根植株;嫁接植株的净光合速率(Pn)、气孔导度(Gs)、胞间CO2浓度(Ci)、叶绿素含量均显著高于自根植株;虽然NaCl胁迫抑制光合作用,但嫁接植株仍表现出优势。以上结果证明,嫁接植株耐盐性优于自根植株。     

Behavioral and neural responses of toads to salt solutions correlate with basolateral membrane potential of epidermal cells of the skin

Dehydrated toads initiated water absorption response (WR) behavior and absorbed water from dilute NaCl solutions. With 200-250 mM NaCl, WR behavior and water absorption were both suppressed. With 200-250 mM Na-gluconate, WR initiation was significantly greater than with NaCl but water loss was greater. Neural recordings from spinal nerve #6 showed a greater integrated response to 250 mM NaCl than to 250 mM Na-gluconate, whereas a larger rinse response was seen with Na-gluconate. Studies with isolated epithelium showed a large increase in conductance (G(t)) when 250 mM NaCl replaced NaCl Ringer's as the apical bathing solution that was accompanied by depolarization of the transepithelial potential (V(t)) and basolateral membrane potential (V(b)). Depolarization of V(b) corresponded with the neural response to 250 mM NaCl. When 250 mM Na-gluconate replaced Ringer's as the apical solution G(t) remained low, V(b) transiently hyperpolarized to values near the equilibrium potential for K(+) and corresponded with the reduced neural response. These results support the hypothesis that chemosensory function of the skin is analogous to that of mammalian taste cells but utilizes paracellular ion transport to a greater degree.     

外源ALA、SNP和Spd对NaCl胁迫下桔梗种子萌发特性的影响

以药用植物桔梗为研究对象,通过测定不同浓度的ALA、SNP和Spd对NaCl胁迫下桔梗种子发芽势、发芽率、萌发指数和平均根长等萌发指标的影响,寻找提高桔梗种子及幼苗在盐胁迫条件下抗性能力的途径.实验结果表明,75 mmol·L-1 NaCl胁迫下的桔梗种子萌发受到显著抑制,但是用不同浓度的ALA、SNP和Spd对桔梗种...     

Purification and properties of ATPase inhibitor from rat liver mitochondria

(1) The ATPase inhibitor protein has been isolated from rat liver mitochondria in purified form. The molecular weight determined by sodium dodecyl sulfate gel electrophoresis is approximately 9500, and the isoelectric point is 8.9.

(2) The protein inhibits both the soluble ATPase and the particle-bound ATPase from rat liver mitochondria. It also inhibits ATPase activities of soluble F₁, and inhibitor-depleted submitochondrial particles derived from bovine heart mitochondria.

(3) On particle-bound ATPase the inhibitor has its maximal effect if incubated in the presence of Mg²⁺. ATP at slightly acidic pH.

(4) The inhibitor has a minimal effect on P_i-ATP exchange activity in sonicated submitochondrial particles. However, unexpectedly the inhibitor greatly stimulates P_i-ATP exchange activity in whole mitochondria while the low ATPase activity of the mitochondria is not affected. The possible mechanism of action of the inhibitor on intact mitochondria is offered.     

The amphipathic membrane proteins associated with gangliosides: The Paul-Bunnell antigen is one of the gangliophilic proteins

Two amphipathic protein fractions soluble in organic solvents as well as in water have been isolated from the ganglioside fraction of bovine erythrocyte membranes by successive chromatography in chloroform-methanol mixture on DEAE-Sephadex, silicic acid, and α-hydroxypropylated Sephadex G50 (LH60) columns. These two fractions contained a similar low molecular weight protein but with distinctively different amino acid composition. One of these proteins has been characterized by having a strong Paul-Bunnell antigen activity and had a binding affinity to ganglioside. A similar protein without Paul-Bunnell antigen activity was isolated as the major ganglioside-associated protein.     

Effect of high external NaCl concentrations on ion transport within the shoot of Lupinus albus. I. Ions in xylem sap

Abstract. Xylem sap was collected from individual leaves of intact transpiring lupin plants exposed to increasing concentrations of NaCl by applying pneumatic pressure to the roots. Concentrations of Na⁺ and Cl⁻ in the xylem sap increased linearly with increases in the external NaCl concentration, averaging about 10% of the external concentration. Concentrations of K⁺ and NO₃⁻, the other major inorganic ions in the sap, were constant at about 2.5 and 1.5 mol m⁻³, respectively. There was no preferential direction of Na ⁺ or Cl⁻ to either young or old leaves: leaves of all ages received xylem sap having similar concentrations of Na⁺ and Cl⁻, and transpiration rates (per unit leaf area) were also similar for all leaves. Plants exposed to 120–160 mol m⁻³ NaCl rapidly developed injury of oldest leaves; when this occurred, the Na⁺ concentration in the leaflet midrib sap had increased to about 40 mol m⁻³ and the total solute concentration to 130 osmol m⁻³. This suggests that uptake of salts from the transpiration stream had fallen behind the rate of delivery to the leaf and that salts were building up in the apoplast.     

Identification and biochemical characterization of Leishmania strains isolated in Peru, Mexico, and Spain

Eight Leishmania promastigotes were isolated from different geographical areas: three (LP1, LP2, and LP3) from the provincial department La Libertad and the fourth (LP4) from the department of Cajamarca (northern Peru); another three (LM1, LM2, and LM3) in the province of Campeche (Mexico); and the last (LS1) from a clinical case of a dog in Madrid (Spain). The isolates were characterized by carbohydrate cell-surface residues using agglutinations with four purified lectins, by isoenzyme analysis using different isoenzymes, by analysis of kinetoplast DNA (kDNA) restriction fragment length polymorphism using four different restriction endonucleases and by the final metabolite patterns after in vitro culture. These isolates were compared with four reference strains and typified as: Leishmania (Leishmania) donovani, two strains of L. (L.) infantum, and one species of L. (Viania) peruviana. According to our results and the statistical study, the Peruvian isolates represent three different strains: one would be L. (V.) peruviana, another the strain isolated in Cajamarca (LP4) and the third would include the three strains from the department of La Libertad (LP1, LP2, and LP3), these latter three isolates being phylogenetically closer to the reference strain L. (L.) donovani. Meanwhile, the three isolates from Mexico form a group with close phylogenetic relationships to each other. The isolate from Spain belongs to the species L. (L.) infantum. Thus, a close correlation was drawn between the identity of each strain and its geographical origin.     

Osmotic and Atmospheric Dehydration Effects in the Lichens Hypogymnia Physodes,Lobaria Pulmonaria,and Peltigera Aphthosa: An in vivo Study of the Chlorophyll Fluorescence Induction

Inactivation of photosynthesis during atmospheric and osmotic (highly concentrated NaCl or sucrose solutions) dehydration was monitored by measurement of chlorophyll fluorescence induction (OIP-phase, Kautsky-curves) in three lichen species. The induction curves were changed in a very similar way by all three treatments. All dehydration effects were rapidly reversible after rehydration. At relatively mild water stress, the rise time to the transient peak F_p was prolonged, and the variable part of fluorescence was diminished. In addition, at severe water stress, a considerable decline of the F₀ value was observed. For NaCl treatment this effect started at water potentials <-8.5 MPa in P. aphthosa, <-12 MPa in H. physodes, and <-21 MPa in L. pulmonaria. Above these water potentials, our observations are in agreement with values from desiccation-tolerant algae, higher plants, and lichens, where an inactivation on the photosystem 2 (PS2) donor side has been postulated. At very low water potentials, the decrease in F₀ probably monitors changes in the organization of the antenna apparatus of PS2. This revised version was published online in June 2006 with corrections to the Cover Date.     

Cortical cell death, cell proliferation, macromolecular movements and rTip1 expression pattern in roots of rice (Oryza sativa L.) under NaCl stress

The mode of action of NaCl in terms of cell proliferation and cell death was examined in seminal roots of rice plants (Oryza sativa L.). Salt/sodium chloride was inhibitory to cell number increase and to cell death in cortical tissue, whereas final cortical cell size was the same as in control roots that were not exposed to NaCl. It seems that NaCl may stimulate the transition phase from cell division to cell elongation. Further analysis of the role of NaCl in the suppression of cortical cell death was confined to a delay in the early stage of cell collapse, which was caused by tonoplast disruption, and plasma-membrane destruction. Sodium chloride did not have any effect on the cell-to-cell movement of macromolecules in the root cortex. In-situ hybridization studies indicated that expression of the gene for tonoplast intrinsic protein (rTip1) was localized predominantly in the epidermal and exodermal cells as well as in metaxylem cells in seminal roots. Upon NaCl treatment, the intensity of rTip1 gene expression was raised in the cortical parenchyma, suggesting that salt plays a role in the rapid onset of cell elongation. Received: 2 April 1998 / Accepted: 18 September 1998