The discrimination between Rb+ and K+ by Escherichia coli is changed after bacteriophage T7 infection

Rb+ and K+ have similar chemical properties. They share the uptake systems in Escherichia coli and can replace each other inside the cell. These common features led to experiments in which the radioactive isotope 86Rb was used to trace intracellular K+ fluxes. However, the E. coli pumps discriminate between these two ions and one should thus be cautious using 86Rb+ as a tracer for K+. We now report that T7 infection alters the degree of discrimination in such a way that changes of intracellular Rb+ do not reflect changes of K+. It has been observed that shortly after infection the 86Rb+ level was strongly reduced (Ponta, H., Altendorf, K.-H. and Schweiger, M. (1976) Mol. Gen. Genet. 149, 145-150). In contrast, determination of the K+ content showed no change directly after infection (Kuhn, A., Jütte, H. and Kellenberger, E. (1983) J. Virol. 47, 540-552). The efflux of 86Rb was only evident when Rb+ was used in trace amounts. In media conditions under which intracellular K+ was mainly replaced by Rb+, 86Rb+ efflux was not observed.     

Drought resistance in pearl millet

The influence of wilting on the levels of free proline, soluble proteins, reducing sugars, starch and on the activities of nitrate reductase, invertase, amylase and pyrophosphatases have been studied in the leaf tissue of five cultivars of pearl millet at their vegetative stage under pot culture conditions. The metabolic changes could not be correlated with the yield behaviour of the cultivars under a drought condition in the field.     

Solution structure of [d(GGTATACC)]2: wrinkled D structure of the TATA moiety

Phase-sensitive two-dimensional nuclear Overhauser effect spectra of [d(GGTATACC)]2 in aqueous deuterium oxide solution at four mixing times were quantified to give all nonoverlapping cross-peak intensities. A structural model for [d(GGTATACC)]2 was built in which the GG- and -CC moieties were in the B-DNA form, while the middle -TATA- moiety was in the wrinkled-D form (BDB model). This model was subjected to energy refinement by molecular mechanics calculations with the program AMBER. Counterions (Na+) were added to neutralize the charges, and water molecules were placed bridging across the minor groove. A complete relaxation matrix analysis was used to calculate two-dimensional nuclear Overhauser effect spectra of [d(GGTATACC)]2 from the above models (before and after energy refinement) and from four other [d(GGTATACC)]2 structural models: regular A, crystalline A, regular B, and energy-minimized B. Among them, the energy-minimized BDB model yielded a set of theoretical spectra that gave the best fit to the experimental spectra. It was also the energetically most stable. Therefore, it is a good representation of the ensemble- and time-averaged structure of the octamer in solution. This model has backbone torsion angles similar to those of B-form DNA in the GG- and -CC moieties and torsion angles similar to those of wrinkled D form DNA in the -TATA- moiety. The base stacking and base pairing are not interrupted at the junctions between the two structural moieties. Its minor groove is narrower than that of B DNA, and the solvent-accessible surface of the minor groove forms a closed hydration tunnel in the middle -TATA- segment.     

Dynamics of lipid-protein interactions. Interaction of apolipoprotein A-II from human plasma high density lipoproteins with dimyristoylphosphatidylcholine

ApoA-II and dimyristoylphosphatidylcholine (DMPC) spontaneously associate to give three different complexes whose structures are determined by the initial reactant concentration and by the reaction temperature with respect to Tc (23.9 degrees C), the gel to liquid crystalline transition temperature of DMPC. At an initial lipid to protein ratio of 45/1, a single complex (2.29 x 10(5) daltons) is quantitatively formed at all temperatures between Tc - 4 degrees C and Tc + 6 degrees C. When the 45/1 complex is mixed with DMPC liposomes there is lipid exchange but no net transfer of lipid, so that the structure of the complex remains unaltered. At an initial molar ratio of 100 to 300:1, the reaction scheme is more complex. At 24 degrees C a 240/1 complex (1.5 x 10(6) daltons) is formed from a precursor 75/1 complex (3.43 x 10(5) daltons) if excess (approximately 300 mol/mol) lipid is present. The 75/1 complex exhibits lipid exchange in the presence of added DMPC liposomes at 24 degrees C, and both the 75/1 and the 240/1 complex can be converted to smaller protein-rich complexes in the presence of added apoA-II. These results suggest that the initial lipid/protein ratio and the physical state of a lipid or lipid . protein complex determines the composition and structure of the resulting complex and support the view that lipid-protein interactions are stronger than protein-protein or lipid-lipid interactions.     

The effect of acetaldehyde on gluconeogenesis from xylitol, sorbitol, and fructose by isolated rat liver cells.

In isolated rat liver cells, ethanol inhibited gluconeogenesis from xylitol and sorbitol but not from fructose. Acetaldehyde, at initial concentrations of 0.2, 0.5, and 1.0 mm, stimulated gluconeogenesis from xylitol and sorbitol in the absence of pyrazole but inhibited in the presence of pyrazole. There was no effect with fructose. Acetate had no effect. Methylene blue and pyruvate (but not lactate) prevented the stimulatory as well as the inhibitory effects of acetaldehyde. Acetoacetate (but not β3-hydroxybutyrate) prevented, to a large extent, the inhibitory effects of low (but not high) concentrations of acetaldehyde. The inhibition by low concentrations of acetaldehyde appears to be mediated via acetaldehyde oxidation in the mitochondria, whereas the inhibition by high concentrations of acetaldehyde appears to reflect acetaldehyde oxidation in the cytosol. These data indicate that the inhibitory action of ethanol on glucose production from xylitol and sorbitol can be reproduced by physiological concentrations of acetaldehyde. Changes in the $\text{NAD}{}^{\mathrm{+}}\text{NADH}$ ratio produced during acetaldehyde metabolism appear to be responsible for these effects of acetaldehyde. These changes may contribute to the actions of ethanol on gluconeogenesis from these substrates.     

Purification and properties of cathepsin D from the mammary glands of lactating rabbits

Cathepsin D was purified from the lactating rabbit mammary gland by a rapid procedure, which included fractionation with (NH4)2SO4, acid precipitation, double affinity chromatography on pepstatin-Sepharose 4B and gel filtration on Sephadex G-100, resulting in approximately 360-fold purification of the enzyme over the homogenate and approximately 16% recovery. After isoelectric focusing, the enzyme dissociated into four (pI 5.8, 6.3, 6.5 and 7.2) multiple forms, but appeared homogeneous on polyacrylamide gel electrophoresis. Cathepsin D has a Mr of 45 kDa as determined by Sephadex G-100 column chromatography. On sodium dodecylsulfate/polyacrylamide gel electrophoresis the enzyme gave a single protein band, corresponding to Mr of 45 kDa. The amino acid composition of the enzyme is similar to that of cathepsins D from other tissues. A single N-terminal amino acid was glycine. Cathepsin D contains 6.4% carbohydrates consisting of mannose, galactose, fucose and glucosamine at a ratio of 3:9:2:2. Cathepsin D is inhibited by pepstatin with Ki of 2.5 X 10(-9) M and irreversibly by N-diazoacetyl-N'-2.4-dinitrophenyl-ethylene diamine. The enzyme hydrolyzes bovine hemoglobin with the maximal activity at pH 3.0 with Km = 10(-5) M and HLeu-Ser-Phe(NO2)-Nle-Ala-Leu-OMe with Km = 4 X 10(-5) M and Rcat = 0.95 s-1. The major cleavage sites were Leu15-Tyr16, Phe24-Phe25 and Phe25-Tyr26 during hydrolysis of the oxidized insulin B-chain by cathepsin D.     

Study of the mechanism of initiation of enzymatic NADPH-dependent lipid peroxidation in membranes of the endoplasmic reticulum

The localization and mechanism of generation of active oxygen species in the enzymatic NADPH-dependent lipid peroxidation system in liver microsomes were studied. Using the spin-trapping method, the key role of active oxygen species in the initiation of NADPH-dependent enzymatic lipid peroxidation was confirmed. It was shown that active oxygen species are generated via consecutive one-electron reduction of the oxygen molecule by NADPH-cytochrome P-450 reductase.