Evidence for two possible glnB-type genes in Herbaspirillum seropedicae.

Two glnB-like genes have been isolated from Herbaspirillum seropedicae by complementation of the Klebsiella pneumoniae glnB502 mutant for growth on nitrate. One of these glnB-like genes has been sequenced and shows strong identity with GlnB proteins derived from other organisms. A Tn5-20 mutation of this glnB was Nif negative.     

Comparative Physiology of Dimethyl Sulfide Production by Dimethylsulfoniopropionate Lyase in Pseudomonas doudoroffii and Alcaligenes sp. Strain M3A

Dimethylsulfoniopropionate (DMSP) lyase enzymatically cleaves DMSP, an algal metabolite, to produce acrylate, a proton, and dimethyl sulfide (DMS), the most abundant volatile sulfur compound emitted from oceans. The physiology of DMS production by DMSP lyase was studied in vivo in an Alcaligenes-like organism, strain M3A, a salt marsh bacterial isolate, and in a marine strain, Pseudomonas doudoroffii. Enzymes from both strains were induced at optimum rates by 1 mM DMSP and vigorous aeration. P. doudoroffii was very sensitive to continued aeration and lost activity rapidly; the enzyme was more stable when aeration ceased. In addition to DMSP, acrylate and several of its analogs acted as inducers of DMSP lyase in Alcaligenes sp. strain M3A but not in P. doudoroffii. Turnover of DMSP by P. doudoroffii was enhanced by 3.5% NaCl or seawater, whereas the Alcaligenes sp. strain M3A enzyme was not salt dependent and salt did not greatly affect its activity. The pH profile showed two peaks of DMSP lyase activity (6.5 and 8.8) for Alcaligenes sp. strain M3A and a single peak at pH 8 for P. doudoroffii. Enzyme activity in both organisms was inhibited by methyl-3-mercaptopropionate and homocysteine. Cyanide, azide and p-chloromercuribenzoate inhibited only the P. doudoroffii DMSP lyase. The apparent K(infm) values for DMSP for cell cultures of Alcaligenes sp. strain M3A and P. doudoroffii were ca. 2 mM and <20 (mu)M, respectively. The differences in the physiology of DMSP metabolism in these two bacterial isolates may enable them to exist in diverse ecological niches.     

Neurochemical effects ofl-pyroglutamic acid

The effect ofl-pyroglutamic acid, a metabolite that accumulates in pyroglutamic aciduria, on different neurochemical parameters was investigated in adult male Wistar rats. Glutamate binding, adenylate cyclase activity and G protein coupling to adenylate cyclase were assayed in the presence of the acid.l-pyroglutamic acid decreased Na⁺-dependent and Na⁺-independent glutamate binding Basal and GMP-PNP stimulated adenylate cyclase activity were not affected by the acid. Furthermore, rats received unilateral intrastriatal injections of 10–300 nmol of bufferedl-pyroglutamic acid. Vehicle (0.25 M Tris-Cl, pH 7.35–7.4) was injected into the contralateral striatum. Neurotoxic damage was assessed seven days after the injection by histological examination and by weighing both cerebral hemispheres. No difference in histology or weight could be identified between hemispheres. These results suggest that, although capable of interfering with glutamate binding, pyroglutamate did not cause a major lesion in the present model of neurotoxicity.     

Immunological and genetic characterization of 2-deoxygalactose-resistant, galactokinase-deficient mutants of Chinese hamster cells: evidence for structural mutations at the galK locus.

Ten independent mutants resistant to 2-deoxygalactose and without any detectable galactokinase activity (null-galactokinase mutations) were isolated from mutagenized Chinese hamster somatic cells. They were analyzed for the presence of serologically cross-reacting material (CRM) with antiserum generated against highly purified Chinese hamster galactokinase. All 10 mutants contain cross-reacting material (i.e., were CRM+), indicating that all the mutations affect the correct expression of a product of the galactokinase structural gene. Complementation analysis among them shows that the 10 mutations fall in one functional genetic unit.     

Denaturation of mouse satellite DNA upon melting of chromatin in solution

The denaturation of mouse satellite DNA upon melting of chromatin in solution of low ionic strength has been studied. A procedure for preparation of partially denaturated chromatin was developed which enabled the isolation of double-stranded (non-denatured) DNA sequences according to their thermal stability in chromatin. The content of mouse satellite DNA in these DNA sequences was determined by hybridization with RNA, complementary to satellite DNA in order to find the temperature interval of denaturation of satellite DNA. It was found that the melting temperature of satellite DNA in chromatin was lower than that of the total DNA. The results are discussed in relation to previously reported anomalous behaviour of satellite DNA upon melting of chromatin on hydroxyapatite.     

Temperature-dependent morphological changes in membranes of bacillus stearothermophilus

Bacillus stearothermophilus cells vary the lipid fatty acid composition of cytoplasmic membranes with growth temperature. Spin label studies of such membranes have been interpreted to indicate lateral lipid phase separations at the growth temperature. We have now used freeze-fracture electron microscopy to confirm the spin label studies. Freeze-fracture faces of protoplasts indicate slight but distinct protein aggregation at the growth temperature. Aggregation increases rapidly with decreasing quench temperature in wild-type cells. In contrast we were unable to demonstrate extended protein segregation in membranes of a temperature-sensitive mutant that contains more than 58% branched fatty acids. Storage of protoplasts for prolonged times below the lipid phase transition results in the appearance of corrugated fracture faces with 300- to 500-Å repeat patterns, although this organism does not synthesize lecithins.     

Isolation and properties of flagella of trypanosomatids.

A procedufe is described for the isolation of flagella of Crithidia fasciculata, Herpetomonas samuelpessoai and Leishmania tarentolae in a highly purified state and giving reasonably good yield. The 3 types of flagella give a similar electrophoretic pattern of proteins. It is shown that H. samuelpessoai and, to a lesser extent, C. fasciculata flagella confer protection against Trypanosoma cruzi infection.     

Immobilization of alcohol dehydrogenase by gel entrapment of cells of Saccharomyces cerevisiae

A stable immobilized preparation of alcohol dehydrogenase (ADH) (EC 1.1.1.1) was obtained by entrapment of ADH-containing Saccharomyces cerevisiae cells in polyacrylamide, polymerized by gamma-rays (100 kR). The permeability barrier for the substrate through the cell membrane was found to be eliminated on entrapment. The stability characteristics, pH-activity profile and other properties of the entrapped ADH are presented. A four-fold enhancement in K_m for NAD⁺ was observed on entrapment, whereas K_m for ethanol was not altered.     

Effects of β-endorphin on brain serotonin metabolism

Human β-endorphin (15 μg) administered intracisternally increased concentrations of serotonin (5HT) and its metabolite, 5-hydroxyindoleacetic. acid (5-HIAA), in brain stem and hypothalamus and decreased 5-HIAA concentrations in hippocampus. These data are compatible with the hypothesis that β-endorphin increases 5HT turnover in brain stem and hypothalamus and decreases 5HT turnover in hippocampus. β-endorphin increased in brain stem and hypothalamus and decreased in hippocampus the rate of pargyline-induced decline of 5-HIAA. β-endorphin decreased the rate of pargyline-induced accumulation of 5HT in all these brain regions. The probenecid-induced accumulation of 5-HIAA in brain stem was decreased by β-endorphin. These data are compatible with the hypothesis that β-endorphin increases release of 5HT from neurons in brain stem and hypothalamus and decreases release of 5HT from neurons in hippocampus. The data require further a hypothesis that β-endorphin either decreases 5HT reuptake in these three brain regions or increases 5-HIAA egress from brain.     

Secondary flow in the human common carotid artery imaged by MR angiography.

The blood flow in arteries affects both the biology of the vessels and the development of atherosclerosis. The flow is three-dimensional, unsteady, and difficult to measure or to model computationally. We have used phase-shift-based magnetic resonance angiography to image and measure the flow in the common carotid arteries of a healthy human subject. There was curvature of the vessels and thin-slice dynamic flow imaging showed evidence of the presence of secondary motions. Flexing the cervical spine straightened the vessels and reduced the asymmetry of the flow.