Salicylate-promoted permeation of cefoxitin, insulin and phenylalanine across red cell membrane. Possible mechanism

Uptake of sodium cefoxitin, D-phenylalanine and insulin into human red blood cells was significantly enhanced by the presence of salicylate and 5-methoxysalicylate in the medium. The mechanism of adjuvant action appeared to depend on an affinity between the adjuvant and the protein fraction in the erythrocyte membrane. The inhibitory effect of DIDS and phlorizin on the salicylate-enhanced uptake of these compounds strongly suggests that the ability of salicylate to permeate the membrane may be essential for it to act as an adjuvant.     

Interactions of poly-N6-methyladenylic Acid and N6-substituted-9-methyladenines with poly-5-bromouridylic Acid: A Novel Base-pairing

Abstract

The interaction of poly-N⁶-methyladenylic acid (poly(m⁶A)) with poly-5-bromouridylic acid (poly(BU)) was studied by the mixing curve method. A 1 m⁶A: 2 BU stoichiometry was clearly indicated over a wide range of ionic strengths at neutral pH, while the binding of poly(m⁶A) to poly(U) is known to occur with 1 m⁶A:1 U. Digestion by nuclease S1 confirmed this stoichiometry, indicating the absence of single strands in a 1:2 mixture. Heating profile analysis and hydroxyapatite column chromatography provided further confirmation of this finding. To determine whether 1:2 stoichiometry holds in a monomer-polymer system, the interaction of N⁶-methyl-9-methyladenine (m⁶m⁹A), a corresponding monomer of poly(m⁶A), with poly(BU) was investigated.

Equilibrium dialysis experiments showed the stoichiometry of the interaction to be 1 m⁶A: 2 BU. Thus, we would describe some structural studies of the above complexes using c.d. and i. r. spectroscopy. Poly (m⁶A)·2poly(BU) and m⁶m⁹A·2poly(BU) are helical and analogous to each other in structure, and the bases in the complexes are all bound by hydrogen-bonding. N⁶-(Δ²-isopentenyl)- and N⁶-allyl-9-methyladenine were also found to form complexes with poly(BU), giving similar c.d. spectra with that of m⁶m⁹A·2poly(BU). The melting experiments indicated the T_ms to be substantially decreased, compared to the parent unmodified complexes, even though the T_m dependence of the polymer complex on salt concentration conforms to the typical triple strand. In the following, the biological significance of this novel pairing will be discussed.     

Expression of soybean glycinin subunit precursor cDNAs in Escherichia coli

As the cDNAs encoding A1aB1b and A2B1a subunit precursors of the glycinin A2 subfamily contain a unique NcoI site sequence, (A)CCATGG, occurring at their translation initiation sites, plasmids were constructed to direct the synthesis of those precursor proteins by inserting NcoI/PstI fragments derived from those cDNA clones into the NcoI/PstI-pKK233-2 expression vector in Escherichia coli MV1190, respectively. The resultant plasmids directed the expression of 57-kDa protein components that have molecular masses in agreement with those of the in vitro translation products directed by glycinin A2 subfamily mRNAs, by the addition of isopropyl beta-D-thiogalactoside. These proteins, which comprised as much as 1% of the total bacterial protein, are immunoprecipitable with rabbit antibodies specific for glycinin subunits. This procedure makes glycinin subunits available as a model for studying structure-function relationships in seed proteins using site-directed mutagenesis. This is the first expression of glycinin-like storage protein in E. coli.     

Mitochondrial DNA of the extinct quagga: Relatedness and extent of postmortem change

Sequences are reported for portions of two mitochondrial genes from a domestic horse and a plains zebra and compared to those published for a quagga and a mountain zebra. The extinct quagga and plains zebra sequences are identical at all silent sites, whereas the horse sequence differs from both of them by 11 silent substitutions. Postmortem changes in quagga DNA may account for the two coding substitutions between the quagga and plains zebra sequences. The hypothesis that the closest relative of the quagga is the domestic horse receives no support from these data. From the extent of sequence divergence between horse and zebra mitochondrial DNAs (mtDNAs), as well as from information about the fossil record, we estimate that the mean rate of mtDNA divergence in Equus is similar to that in other mammals, i.e., roughly 2% per million years.     

Lattice swelling with the selective digestion of elastic components in single-skinned fibers of frog muscle.

Changes in the 1.0 lattice spacing during trypsin (0.25 micrograms/ml) treatment in mechanically skinned single fibers of frog muscle was examined by an x-ray diffraction method at various sarcomere lengths. The resting tension of a relaxed fiber was decreased by trypsin treatment but the stiffness of a rigor fiber was not, suggesting that elastic components were selectively digested. With progression of the digestion, the lattice spacing increased remarkably at longer sarcomere lengths and finally became independent of the sarcomere length. The increase in the lattice spacing was proportional to the decrease in the resting tension. These results support our previous suggestion (Higuchi, H., and Y. Umazume, 1986, Biophys. J., 50:385-389) that the lattice spacing decreases at long lengths due to compressive force exerted by a lateral elastic component that connects thick filaments to an axial elastic component. Consequently, it is unlikely that the decrease in the lattice spacing is determined by a decrease in the repulsive force between thick and thin filaments with stretching a fiber.     

Cardiovascular changes associated with decreased aerobic capacity and aging in long-distance runners

Fifty-five male runners aged between 30 to 80 years were examined to determine the relative roles of various cardiovascular parameters which may account for the decrease in maximal oxygen uptake (VO2max) with aging. All subjects had similar body fat composition and trained for a similar mileage each week. The parameters tested were VO2max, maximal heart rate (HRmax), cardiac output (Q), and arteriovenous difference in oxygen concentration (Ca-Cv)O2 during graded, maximal treadmill running. Average body fat and training mileage were roughly 12% and 50 km.week-1, respectively. The average 10-km run-time slowed significantly by 6.0%.decade-1 [( 10-km run-time (min) = 0.323 x age (years) + 24.4] (n = 49, r = 0.692, p less than 0.001]. A strong correlation was found between age and VO2max [( VO2max (ml.kg-1.min-1) = -0.439 x age + 76.5] (n = 55, r = -0.768, p less than 0.001]. Thus, VO2max decreased by 6.9%.decade-1 along with reductions of HRmax (3.2%.decade-1, p less than 0.001) and Q (5.8%.decade-1, p less than 0.001), while no significant change with age was observed in estimated (Ca-Cv)O2. It was concluded that the decline of VO2max with aging in runners was mainly explained by the central factors (represented by the decline of HR and Q in this study), rather than by the peripheral factor (represented by (Ca-Cv)O2).     

Evidence for age-related change in plasma 19-hydroxyandrostenedione

The steroid, 19-hydroxyandrost-4-ene-3, 17-dione (19-hydroxyandrostene-dione, 19-OH-A-dione) has been known to enhance the mineralocorticoid action of aldosterone. To investigate the age-related change in the plasma 19-OH-A-dione concentration, plasma 19-OH-A-dione, androst-4-ene-3, 17-dione (A-dione), aldosterone and cortisol of 38 non-hypertensive healthy subjects (18 young men and 20 aged men) measured by specific radioimmunoassays. The basal plasma 19-OH-A-dione and A-dione concentration in aged men was significantly lower than in young men (P less than 0.01). Moreover, there was found to be a positive correlation between plasma 19-OH-A-dione and A-dione (P less than 0.01). On the other hand, plasma aldosterone and cortisol in aged men showed a tendency to decrease, but no statistical significance compared to young men was observed. This study demonstrated that there was an apparent age-related decrease not only in plasma A-dione, but also in plasma 19-OH-A-dione, an amplifier or aldosterone action.     

12S,19- and 12S,20-dihydroxyeicosanoids: novel 12S-hydroxy-5,8-cis-10-trans-14-cis-eicosatetraenoic acid metabolites formed by hydroxylation and reduction in murine lymphocytes

Murine spleen cells and purified B lymphocytes oxidized arachidonic acid via the lipoxygenase pathway. The major metabolite of both the whole spleen and enriched B lymphocytes was 12S-hydroxy-5,8-cis-10-trans-14-cis-eicosatetraenoic acid. A novel metabolite was observed that did not have an absorbance from 210 to 400 nm, indicating the absence of a conjugated double bond system. The new metabolite was converted to the methyl ester, reduced by platinum oxide, derivatized to the trimethylsilyl ether, and analyzed by gas chromatography-mass spectrometry. A major and a minor component were observed in the analysis of the new compound. The major component had major diagnostic ions indicating the presence of hydroxyl groups at C-12 and C-19. The minor component had major diagnostic ions indicating the presence of hydroxyl groups at C-12 and C-20. The new metabolites are characterized as a mixture of 12S,19- and 12S,20-dihydroxyeicosanoids presumably formed by hydroxylation and reduction of one or more double bonds of 12S-hydroxy-5,8-cis-10-trans-14-cis-eicosatetraenoic acid. These metabolites were formed predominantly with whole spleen lymphocytes but could be detected at longer incubation times or by using 12S-hydroxy-5,8-cis-10-trans-14-cis-eicosatetraenoic acid as the starting substrate with highly enriched B lymphocytes.