Metabolic Syndrome Is Associated with Increased Oxo-Nitrative Stress and Asthma-Like Changes in Lungs

Epidemiological studies have shown an increased obesity-related risk of asthma. In support, obese mice develop airway hyperresponsiveness (AHR). However, it remains unclear whether the increased risk is a consequence of obesity, adipogenic diet, or the metabolic syndrome (MetS). Altered L-arginine and nitric oxide (NO) metabolism is a common feature between asthma and metabolic syndrome that appears independent of body mass. Increased asthma risk resulting from such metabolic changes would have important consequences in global health. Since high-sugar diets can induce MetS, without necessarily causing obesity, studies of their effect on arginine/NO metabolism and airway function could clarify this aspect. We investigated whether normal-weight mice with MetS, due to high-fructose diet, had dysfunctional arginine/NO metabolism and features of asthma. Mice were fed chow-diet, high-fat-diet, or high-fructose-diet for 18 weeks. Only the high-fat-diet group developed obesity or adiposity. Hyperinsulinemia, hyperglycaemia, and hyperlipidaemia were common to both high-fat-diet and high-fructose-diet groups and the high-fructose-diet group additionally developed hypertension. At 18 weeks, airway hyperresponsiveness (AHR) could be seen in obese high-fat-diet mice as well as non-obese high-fructose-diet mice, when compared to standard chow-diet mice. No inflammatory cell infiltrate or goblet cell metaplasia was seen in either high-fat-diet or high-fructose-diet mice. Exhaled NO was reduced in both these groups. This reduction in exhaled NO correlated with reduced arginine bioavailability in lungs. In summary, mice with normal weight but metabolic obesity show reduced arginine bioavailability, reduced NO production, and asthma-like features. Reduced NO related bronchodilation and increased oxo-nitrosative stress may contribute to the pathogenesis.     

Direct Evidence for Changes in the Resistance of Legume Root Nodules to O2 Diffusion

Indirect evidence suggests that legumes can adjust rapidly theresistance of their root nodules to O₂ diffusion. Here we describeexperiments using O₂ specific micro-electrodes and dark fieldmicroscopy to study directly the operation of this diffusionbarrier. The O₂ concentration sensed by the electrode decreasedsharply in the region of the inner cortex and was less than1.0 mmol m^–3 throughout the infected tissue in nodulesof both pea (Pisum sativum) and french bean (Phaseolus vulgaris).In a number of experiments the ambient O₂ concentration wasincreased to 40% while the electrode tip was just inside theinner cortex. In 13 out of 21 cases the O₂ concentration atthis position either remained low and unchanged or increasedirreversibly to near ambient values. In the remaining casesthe O₂ concentration increased after 1 to 2.5 min and then decreasedto its former value. These results are ascribed to an increasein resistance of the barrier in response to increased O₂ fluxinto the nodule. It was shown microscopically that air spacesboth at the boundary between the infected zone and the innercortex, and within the infected zone started to disappear 3min after nodules were exposed to high ambient O₂ concentrationsand had disappeared completely after 8 min. These spaces werenot changed by exposure of the nodule for 10 min to either N₂or air. Key words: Oxygen, root nodules, air spaces     

Evolution of the 28S ribosomal RNA gene in anurans: regions of variability and their phylogenetic implications

Fifteen restriction sites were mapped to the 28S ribosomal RNA gene of individuals representing 54 species of frogs, two species of salamanders, a caecilian, and a lungfish. Eight of these sites were present in all species examined, and two were found in all but one species. Alignment of these conserved restriction sites revealed, among anuran 28S rRNA genes, five regions of major length variation that correspond to four of 12 previously identified divergent domains of this gene. One of the divergent domains (DD8) consists of two regions of length variation separated by a short segment that is conserved at least throughout tetrapods. Most of the insertions, deletions, and restriction-site variations identified in the 28S gene will require sequence-level analysis for a detailed reconstruction of their history. However, an insertion in DD9 that is coextensive with frogs in the suborder Neobatrachia, a BstEII site that is limited to representatives of two leptodactylid subfamilies, and a deletion in DD10 that is found only in three ranoid genera are probably synapomorphies.      

Conformations of peptides containing Z-alpha,beta-dehydroleucine (delta ZLeu). A comparison of Boc-Pro-delta ZLeu-Gly-NHEt and Boc-Pro-delta ZPhe-Gly-NHEt

Two tripeptides of the type Boc-Pro-delta ZX-Gly-NHEt (where X = Leu, Phe) have been synthesized and their solution conformations investigated by 270 MHz 1H n.m.r. and i.r. spectroscopy. These conformational studies indicated that delta ZLeu, similar to delta ZPhe, has a strong tendency to stabilize folded Type II beta-turn conformations when present at i + 2 position.     

Helix packing of leucine-rich peptides: a parallel leucine ladder in the structure of Boc-Aib-Leu-Aib-Aib-Leu-Leu-Leu-Aib-Leu-Aib-OMe.

The packing of peptide helices in crystals of the leucine-rich decapeptide Boc-Aib-Leu-Aib-Aib-Leu-Leu-Leu-Aib-Leu-Aib-OMe provides an example of ladder-like leucylleucyl interactions between neighboring molecules. The peptide molecule forms a helix with five 5----1 hydrogen bonds and two 4----1 hydrogen bonds near the C terminus. Three head-to-tail NH ... O = C hydrogen bonds between helices form continuous columns of helices in the crystal. The helicial columns associate in an antiparallel fashion, except for the association of Leu ... Leu side chains, which occurs along the diagonal of the cell where the peptide helices are parallel. The peptide, with formula C56H102N10O13, crystallizes in space group P2(1)2(1)2(1) with Z = 4 and cell parameters a = 16.774(3) A, b = 20.032(3) A and c = 20.117(3) A; overall agreement factor R = 10.7% for 2014 data with magnitude of F(obs) greater than 3 sigma (F); resolution 1.0 A.     

Membrane channel forming polypeptides. Molecular conformation and mitochondrial uncoupling activity of antiamoebin, an alpha-aminoisobutyric acid containing peptide

The conformations of the 16-residue fungal peptide antiamoebin I (Ac-Phe-Aib-Aib-Aib-D-Iva-Gly-Leu-Aib-Aib-Hyp-Gln-D-Iva-Hyp-Aib-Pro-P hol) have been investigated in dimethyl sulfoxide solution by one- and two-dimensional NMR techniques. A substantial number of resonances in the 270-MHz 1H NMR spectrum have been assigned. Intramolecularly hydrogen-bonded (solvent inaccessible) NH groups have been identified by determining solvent and temperature dependence of NH chemical shifts and rates of hydrogen-deuterium exchange. Ten backbone NH groups are inaccessible to solvent, while three NH groups assigned to the Phe(1), Aib(2), and Aib(8) residues are exposed to solvent. Interresidue nuclear Overhauser effects are consistent with psi values of approximately 120 +/- 30 degrees for Phe(1) and Leu(7). The NMR results, together with the stereochemical constraints imposed by the presence of alpha-aminoisobutyryl, isovalyl, prolyl, and 4-hydroxyprolyl residues, favor a highly ordered structure. Two backbone conformations consistent with the data are considered. Antiamoebin is shown to be an effective uncoupler of oxidative phosphorylation in rat liver mitochondria, providing evidence for its membrane-modifying activity.     

Proton translocation during denitrification in Rhodopseudomonas sphaeroides f. denitrificans

Proton translocation during the reduction of NO ₃ ^- , NO ₂ ^- , N₂O and O₂, with endogenous substrates, in washed cells of Rhodopseudomonas sphaeroides f. denitrificans was investigated by an oxidant pulse method. On adding NO ₂ ^- to washed cells, anaerobically in the dark, an alkalinization occurred in the reaction mixture followed by acidification. When NO ₃ ^- , N₂O or O₂ was added to cells in the dark or with these compounds and NO ₂ ^- in light an acidification only was observed. Proton translocation was inhibited by carbonyl cyanide-m-chlorophenyl hydrazone.Valinomycin treated cells produced acid in response to the addition of either NO ₃ ^- , NO ₂ ^- , N₂O or O₂. The proton extrusion stoichiometry ( ratios) in illuminated cells were as follows: NO ₃ ^- 0.5N₂, 4.82; NO ₂ ^- 0.5N₂, 5.43; N₂ON₂, 6.20; and O₂H₂O, 6.43. In the dark the comparable values were 3.99, 4.10, 4.17 and 3.95. Thus, illuminated cells produced higher values than those in the dark, indicating a close link between photosynthesis and denitrification in the generation of proton gradients across the bacterial cell membranes.When reduced benzyl viologen was the electron donor in the presence of 1 mM N-ethylmaleimide and 0.5 mM 2-n-heptyl-4-hydroxyquinoline-N-oxide in the dark, the addition of either NO ₃ ^- , NO ₂ ^- or N₂O to washed cells resulted in a rapid alkalinization of the reaction mixture. The stoichiometries for proton consumption, ratios without a permeant ion were NO ₃ ^- NO ₂ ^- ,-1.95; NO ₂ ^- 0.5 N₂O,-3.03 and N₂ON₂,-2.02. The data indicate that these reductions occur on the periplasmic side of the cytoplasmic membrane.Abbreviations BVH reduced benzyl viologen - CCCP carbonyl cyanide m-chlorophenyl hydrazone - DIECA N, N-diethyl-dithiocarbamate - HOQNO 2-n-heptyl-4-hydroxyquinoline-N-oxide - NEM N-ethylmaleimide     

Crystal structure of Boc-Ala-Aib-Ala-Aib-Aib-methyl ester,a pentapeptide fragment of the channel-forming ionophore suzukacillin

t-Buthyoxycarbonyl-L -alanyl-α-aminiosobutyryl-L -alanyl-α-aminoisobutyryl-α-aminoisobutyric acid methyl ester (t-Boc-L -Ala-Aib-L -Ala-Aib-Aib-OMe), C₂₄H₄₃N₅O₈, an end-protected pentapeptide with a sequence corresponding to the 6th through the 10th residues in suzukacillin, crystallizes in the orthorhombic space group P2₁2₁2₁ with a = 11.671, b = 14.534, c = 17.906 Å and z = 4. The molecule exists as a right-handed 3₁₀-helix with a pitch of 6.026 Å. The helix is stabilized by three 4 → 1 hydrogen bonds with the NH groups of Ala(3), Aib(4), and Aib(5) hydrogen bonding to the carbonyl oxygens of t-Boc, Ala(1), and Aib(2), respectively. The helical molecules arrange themselves in a head-to-tail fashion along the a direction in such a way that the NH groups of Ala(1) and Aib(2) hydrogen bond to the carbonyl oxygens of Aib(4) and Aib(5), respectively, of a translationally related molecule. The helical columns thus formed close-pack nearly hexagonally to form the crystal.     

Fluorescence polarization as a tool to study lectin-sugar interaction

The binding ofRicinus communis agglutinin andAbrus agglutinin to 4-methylumbelliferyl β-D-galactopyranoside was studied by equilibrium dialysis, fluo-rescence quenching and fluorescence polarization. The number of binding sites and the association constant value obtained by fluorescence polarization for bothRicinus communis agglutinin andAbrus agglutinin are in close agreement with those obtained by the other methods. This indicates the potential of ligand-fluorescence polarization measurements in the investigation of lectin-sugar interactions.