Salinity-induced changes in the structure and ultrastructure of bean root cells

The effect of 80 mM NaCl on the structure and ultrastructure of root cells ofPhaseolus vulgaris plants has been investigated. Roots of plants treated with NaCl were shorter and had less secondary roots than control plants. In control plants, epidermal cells were isodiametric and uniformly placed forming a thin layer, whereas in stressed plants, the shape and disposition of epidermal cells was less regular. The cortical cells of control plant were round-shaped and distributed allowing large and well defined intercellular spaces, whereas stressed plants presented irregular cells, which were interdigitated, thus decreasing the volume of the intercellular spaces. Presence of 80 mM NaCl did not result in significant differences in the number, shape or distribution of the cell organelles. Membrane vesiculation was often observed in cells of NaCl treated plants. Addition of 80 mM NaCl to the growth medium considerably increased the leakage of solutes from intact plant roots back to the solution especially K⁺ and Ca²⁺.     

Isoenzymatic characterization of potentially useful forage shrubs belonging to the genus Cytisus and allies (Genistea: Leguminosae)

Twenty-five accessions belonging to the genus Cytisus and allies (Genisteae: Leguminosae), potentially useful for agrosylvopastoral systems in the Mediterranean basin, were analyzed using isoenzymes. The aim was to assess the taxonomic usefulness of these characters at the specific and generic levels, and its value as an aid for the taxonomy. A high level of infrageneric and infraspecific variability was detected. Numerical analysis showed that isoenzymes are useful for the characterization both at the specific and generic levels. The obtained dendrogram supports the generic arrangement proposed by Bisby [(1981) Advances in Legume Systematics, Vol. 1, pp. 409–425], therefore helping to elucidate the difficult taxonomy of the studied species.     

A kinetic study of the interaction of vanadate with the Ca2+ + Mg2+-dependent ATPase from sarcoplasmic reticulum.

Ca2+ + Mg2+-dependent ATPase from sarcoplasmic reticulum was inhibited by preincubation with vanadate. When the inhibited enzyme was preincubated in the presence of vanadate and assayed in its absence, a slow reactivation process was observed. This slow, hysteretic, process was exploited to study the influence of Ca2+ and ATP on the dissociation of vanadate. Ca2+ alone slowly displaced vanadate from the inhibited enzyme, and a rate constant of 0.1 min-1, at 25 degrees C, was calculated for this re-activation process. However, ATP re-activated with an apparent constant that hyperbolically depended on ATP concentration, and from it a rate constant for vanadate dissociation induced by ATP of 0.5 min-1 was calculated. It is deduced from the kinetic studies that ATP binds to the enzyme-vanadate complex, forming a ternary complex, with a dissociation constant of 4 microM, and that this binding accelerates vanadate dissociation. Binding experiments with [14C]ATP showed that ATP binds to the enzyme-vanadate complex with a dissociation constant of 12 microM, i.e. the affinities calculated with the isotope technique and the kinetic procedure are of the same order of magnitude.     

Naltrexone effects on pituitary neurointermediate lobe and median eminence

The long-acting opiate antagonist naltrexone hydrochloride was administered by intraperitoneal injection, in a dose response protocol, to adult rats. The drug was used to observe effects of opiate receptor blockade on cells of the pituitary gland and adjacent hypothalamus. At higher drug doses (5mg/kg or 10mg/kg), neurites directly innervating pars intermedia cells contained swollen vesicles and disrupted membranous elements. Fibers within the median eminence of the hypothalamus appeared swollen, and contained myelin figures. Despite the consistent degenerative changes appearing in neurites, measurements of levels of dopamine, norepinephrine, and epinephrine in striatum, and hypothalamus did not differ significantly between naltrexone-treated or control animals, although there was a significant elevation of norepinephrine in the pituitary after drug treatment. At all drug dose levels administered, supraependymal neuron-like cells appeared atop the ependyma of the third ventricle above the median eminence. These observations suggest that naltrexone produces specific neurotoxic effects on neurites of the tuberoinfundibular system, and may induce changes in the ventricular environment which stimulate the appearance of supraependymal neurons.     

Efficient transformation of a Yersinia enterocolitica strain (serotype O:3) by pBR322 DNA

Abstract The Yersinia enterocolitica strain MS201 (serotype O:3) was transformed by pBR322 DNA extracted from an Escherichia coli strain. The pBR322 DNA extracted from an Escherichia coli strain. The pBR322 DNA extracted from the transformed Y. enterocolitica is able to transform plasmid-free MS201 at a significantly higher frequency, suggesting the existence of a restriction-modification system in MS201.     

Substrate selectivity of squalene synthetase.

Six 1-3H-labeled analogues of farnesyl pyrophosphate have been studied as potential substrates for yeast and rat liver squalene synthetases: 2-methylfarnesyl pyrophosphate (4), 3-demethylfarnesyl pyrophosphate (5), 7,11-dimethyl-3-ethyl-2,6,10-dodecatrienyl pyrophosphate (6), 6,7,10,11-tetrahydrofarnesyl pyrophosphate (7), 4-methylthiofarnesyl pyrophosphate (8), and 4-fluorofarnesyl pyrophosphate (9). Analogues 4 and 5 are enzymatically incorporated into 11-methylsqualene (10) and 10-demethylsqualene (11), respectively, even if no farnesyl pyrophosphate is added to the incubations. None of the other analogues gives nonpolar products with either the yeast or liver enzymes. No tritium is enzymatically released to the medium from any of the analogues, indicating that they are not accepted at the first (proton exchanging) site. The data rule out formation of dead-end presqualene pyrophosphate products with analogues as first, but not as second, substrates. Implications of these results for the enzyme active-site topology and mechanism are discussed.     

Glucosylenamines as glycosyl acceptors: synthesis of gentiobiosylenamines.

The preparation of 2,3,4-tri-O-benzyl- (3), 2,3,4-tri-O-acetyl- (4), and 2,3,4-tri-O-benzoyl-N-(2,2-diethoxycarbonylvinyl)-6-O-trityl-beta- D-glucopyranosylamine (5) is described. The reaction of 3-5 with 2,3,4,6-tetra-O-acetyl-alpha-D-glucopyranosyl bromide yields 2,3,4-tri-O-benzyl- (9), 2,3,4-tri-O-acetyl- (10), and 2,3,4-tri-O-benzoyl-N-(2,2-diethoxycarbonylvinyl)-6-O-(2,3,4,6-tet ra-O- acetyl-beta-D-glucopyranosyl)-beta-D-glucopyranosylamine (11), respectively. 2,3,4-Tri-O-benzyl- (6), 2,3,4-tri-O-acetyl- (7), and 2,3,4-tri-O- benzoyl-N-(2,2-diethoxycarbonylvinyl)-beta-D-glucopyranosylamine (8) are also described.     

Monooxygenase activity of cytochrome c peroxidase.

Recombinant cytochrome c peroxidase (CcP) and a W51A mutant of CcP, in contrast to other classical peroxidases, react with phenylhydrazine to give sigma-bonded phenyl-iron complexes. The conclusion that the heme iron is accessible to substrates is supported by the observation that CcP and W51A CcP oxidize thioanisole to the racemic sulfoxide with quantitative incorporation of oxygen from H2O2. Definitive evidence for an open active site is provided by stereoselective epoxidation by both enzymes of styrene, cis-beta-methylstyrene, and trans-beta-methylstyrene. trans-beta-methylstyrene yields exclusively the trans-epoxide, but styrene yields the epoxide and phenylacetaldehyde, and cis-beta-methylstyrene yields both the cis- and trans-epoxides and 1-phenyl-2-propanone. The sulfoxide, stereoretentive epoxides, and 1-phenyl-2-propanone are formed by ferryl oxygen transfer mechanisms because their oxygen atom derives from H2O2. In contrast, the oxygen in the trans-epoxide from the cis-olefin derives primarily from molecular oxygen and is probably introduced by a protein cooxidation mechanism. cis-[1,2-2H]-1-Phenyl-1-propene is oxidized to [1,1-2H]-1-phenyl-2-propanone without a detectable isotope effect on the epoxide:ketone product ratio. The phenyl-iron complex is not formed and substrate oxidation is not observed when the prosthetic group is replaced by delta-meso-ethylheme. CcP thus has a sufficiently open active site to form a phenyl-iron complex, to oxidize thioanisole to the sulfoxide, and to epoxidize styrene and beta-methylstyrene. The results indicate that a ferryl (Fe(IV) = O)/protein radical pair can be coupled to achieve two-electron oxidations. The unique ability of CcP to catalyze monooxygenation reactions does not conflict with its peroxidase function because cytochrome c is oxidized at a distinct surface site (DePillis, G. D., Sishta, B. P., Mauk, A. G., and Ortiz de Montellano, P. R. (1991) J. Biol. Chem. 266, 19334-19341).