γ-Glutamyl conjugation of 5-hydroxytryptamine (serotonin) in the earthworm (Lumbricus terrestris)

The conversion of 5-hydroxytryptamine to several potential metabolites was examined in the annelid earthworm (Lumbricus terrestris). 5-hydroxytryptamine and some related amines were found to be present in several tissues of the earthworm. Injection of 5-hydroxytryptamine into the body cavity of the earthworm resulted in the production of a -glutamyl conjugate of 5-hydroxytryptamine. Incubations of the anterior nerve cord of the earthworm resulted in the accumlation of considerable amounts of 5-hydroxytryptamine and -glutamyl 5-hydroxytryptamine in the incubation medium. The earthworm did not produce any N-acetyl 5-hydroxytryptamine and only very little 5-hydroxyindoleacetic acid. Experiments involving the injection of radiolabeled 5-hydroxytryptamine or coninjection of radiolabeled glutamic acid with unlabeled 5-hydroxytryptamine into the earthworm resulted in the production of radiolabeled -glutamyl 5-hydroxytryptamine. This work demonstrates that the enzymatic conversion of 5-HT in the earthworm is markedly different from that of vertebrates and insects.     

The evolution of pathways for aromatic hydrocarbon oxidation inPseudomonas

The organisation and nucleotide sequences coding for the catabolism of benzene, toluene (and xylenes), naphthalene and biphenylvia catechol and the extradiol (meta) cleavage pathway inPseudomonas are reviewed and the various factors which may have played a part in their evolution are considered. The data suggests that the complete pathways have evolved in a modular way probably from at least three elements. The commonmeta pathway operons, downstream from the ferredoxin-like protein adjacent to the gene for catechol 2,3-dioxygenase, are highly homologous and clearly share a common ancestry. This common module may have become fused to a gene or genes the product(s) of which could convert a stable chemical (benzoate, salicylate, toluene, benzene, phenol) to catechol, thus forming the lower pathway operons found in modern strains. The upper pathway operons might then have been acquired as a third module at a later stage thus increasing the catabolic versatility of the host strains.     

Guanosine triphosphate catabolism in purine nucleoside phosphorylase deficient human B lymphoblastoid cells

GTP catabolism induced by sodium azide or deoxyglucose was studied in purine nucleoside phosphorylase (PNP) deficient human B lymphoblastoid cells. In PNP deficient cells, as in control cells, guanylate was both dephosphorylated and deaminated but dephosphorylation was the major pathway. Only nucleosides were excreted during GTP catabolism by PNP deficient cells and the main product was guanosine. The level of nucleoside excretion was largely affected by intracellular orthophosphate (P_i) level. In contrast, normal cells excreted nucleosides only at low P_i level while at high P_i levels, purine bases (guanine and hypoxanthine) were exclusively excreted. PNP deficiency had no effect on the extent of GMP deamination.     

Stimulation of the anaerobic growth ofSalmonella typhimurium by reduction ofl-carnitine,carnitine derivatives and structure-related trimethylammonium compounds

In view of the development of al-carnitine deficiency, the metabolism ofl-carnitine and structure-related trimethylammonium compounds was studied inSalmonella typhimurium LT₂ by means of thin-layer chromatography (TLC).l-Carnitine, crotonobetaine and acetyl-l-carnitine stimulated the anaerobic growth in a complex medium significantly. The stimulation depended on the formation of -butyrobetaine. The reduction ofl-carnitine proceeded in two steps: (1) Dehydration of thel-carnitine to crotonobetaine, (2) hydrogenation of crotonobetaine to -butyrobetaine. The reduction of crotonobetaine was responsible for the growth stimulation. Terminal electron acceptors of the anaerobic respiration such as nitrate and trimethylamine N-oxide, but not fumarate, suppressed the catabolism ofl-carnitine completely. Glucose fermentation, too, inhibited the reduction ofl-carnitine but optimal growth with a high carnitine catabolism was achieved byd-ribose. The esters of carnitine with medium- and long-chain fatty acids inhibited the growth considerably because of their detergent properties.Abbreviations TLC thin-layer chromatography     

Brain Cytochrome Oxidase in Alzheimer''s Disease

A recent demonstration of markedly reduced (-50%) activity of cytochrome oxidase (CO; complex 4), the terminal enzyme of the mitochondrial enzyme transport chain, in platelets of patients with Alzheimer's disease (AD) suggested the possibility of a systemic and etiologically fundamental CO defect in AD. To determine whether a CO deficiency occurs in AD brain, we measured the activity of CO in homogenates of autopsied brain regions of 19 patients with AD and 30 controls matched with respect to age, postmortem time, sex, and, as indices of agonal status, brain pH and lactic acid concentration. Mean CO activity in AD brain was reduced in frontal (-26%: p less than 0.01), temporal (-17%; p less than 0.05), and parietal (-16%; not significant, p = 0.055) cortices. In occipital cortex and putamen, mean CO levels were normal, whereas in hippocampus, CO activity, on average, was nonsignificantly elevated (20%). The reduction of CO activity, which is tightly coupled to neuronal metabolic activity, could be explained by hypofunction of neurons, neuronal or mitochondrial loss, or possibly by a more primary, but region-specific, defect in the enzyme itself. The absence of a CO activity reduction in all of the examined brain areas does not support the notion of a generalized brain CO abnormality. Although the functional significance of a 16-26% cerebral cortical CO deficit in human brain is not known, a deficiency of this key energy-metabolizing enzyme could reduce energy stores and thereby contribute to the brain dysfunction and neurodegenerative processes in AD.     

Long noncoding RNA OIP5-AS1 acts as a competing endogenous RNA to promote glutamine catabolism and malignant melanoma growth by sponging miR-217

The long noncoding RNA (lncRNA) OIP5-AS1 has been considered to promote the growth and metastasis of many human tumors. However, the role of OIP5-AS1 in melanoma has not been reported. In this study, we found that OIP5-AS1 levels were significantly elevated in melanoma tissue and that high OIP5-AS1 expression was an independent risk factor for the poor survival of patients with melanoma. miR-217 suppressed glutamine catabolism in melanoma cells by targeting glutaminase (GLS), the rate-limiting enzyme of glutamine catabolism. We also demonstrated that OIP5-AS1 acted as a sponge of miR-217 to upregulate GLS expression, thus promoting glutamine catabolism and melanoma growth. Overall, this result elucidates a new mechanism for OIP5-AS1 in metabolism in melanoma and provides a potential therapeutic target for patients with melanoma.     

Nicotine Effects on Dopamine Clearance in Rat Nucleus Accumbens

Abstract: In vivo voltammetry was used to measure the clearance of exogenously applied dopamine (DA) in the nucleus accumbens following acute systemic nicotine administration in urethane-anesthetized rats. The IVEC-5 system was used for continuous in vivo electrochemical measurements. A finite amount of DA was pressure-ejected (25–100 nl, 200 µ M barrel concentration) at 5-min intervals from micropipettes (tip diameter, 10–15 µm) positioned 250 ± 50 µm from the recording electrode. The peak DA concentration after each DA ejection was significantly decreased in rats following nicotine, but not in rats given saline. In addition, when mecamylamine was administered 20 min before nicotine it clearly antagonized nicotine effects. These results suggest that nicotine may actually facilitate DA transporter systems within the nucleus accumbens.     

Synthesis of 4-methylumbelliferyl α-d-mannopyranosyl-(1→6)-β-d-mannopyranoside and development of a coupled fluorescent assay for GH125 exo-α-1,6-mannosidases

Certain bacterial pathogens possess a repertoire of carbohydrate processing enzymes that process host N-linked glycans and many of these enzymes are required for full virulence of harmful human pathogens such as Clostridium perfringens and Streptococcus pneumoniae. One bacterial carbohydrate processing enzyme that has been studied is the pneumococcal virulence factor SpGH125 from S. pneumoniae and its homologue, CpGH125, from C. perfringens. These exo-α-1,6-mannosidases from glycoside hydrolase family 125 show poor activity toward aryl α-mannopyranosides. To circumvent this problem, we describe a convenient synthesis of the fluorogenic disaccharide substrate 4-methylumbelliferone α-d-mannopyranosyl-(1→6)-β-d-mannopyranoside. We show this substrate can be used in a coupled fluorescent assay by using β-mannosidases from either Cellulomonas fimi or Helix pomatia as the coupling enzyme. We find that this disaccharide substrate is processed much more efficiently than aryl α-mannopyranosides by CpGH125, most likely because inclusion of the second mannose residue makes this substrate more like the natural host glycan substrates of this enzyme, which enables it to bind better. Using this sensitive coupled assay, the detailed characterization of these metal-independent exo-α-mannosidases GH125 enzymes should be possible, as should screening chemical libraries for inhibitors of these virulence factors.     

Nicotine-induced vascular endothelial growth factor release via the EGFR-ERK pathway in rat vascular smooth muscle cells

Cigarette smoke has been firmly established as an independent risk factor for atherosclerosis and other vascular diseases. The proliferation and migration of vascular smooth muscle cells (VSMC) induced by growth factors have been proposed to play an important role in the progression of atherosclerosis. In the present study, we investigated the effects of nicotine, which is one of the important constituents of cigarette smoke, on vascular endothelial growth factor (VEGF) release, in rat VSMC. The stimulation of cells with nicotine resulted in a time- and concentration-dependent release of VEGF. Hexamethonium, an antagonist of nicotinic acetylcholine receptor (nAChR), inhibited nicotine-induced VEGF release. We next investigated the mechanisms by which nicotine induces VEGF release in the cells. The nicotine-induced VEGF release was inhibited by treatment with U0126, a selective inhibitor of MEK, which attenuated the nicotine-induced ERK phosphorylation. Nicotine induced a transient phosphorylation of ERK. Furthermore, AG1478, a selective inhibitor of epidermal growth factor receptor (EGFR) kinase, inhibited nicotine-induced ERK phosphorylation and VEGF release. These data suggest that nicotine releases VEGF through nAChR in VSMC. Moreover, VEGF release induced by nicotine is mediated by an EGFR-ERK pathway in VSMC. VEGF may contribute to the risk of cardiovascular diseases in cigarette smokers.     

Selective enrichment and isolation of Rhodopseudomonas palustris using trans-cinnamic acid as sole carbon source

Abstract Enrichment cultures for anoxygenic phototrophs capable of using cinnamic acid as sole organic carbon source consistently yielded the nonsulfur purple bacterium Rhodopseudomonas palustris . Pure cultures of R. palustris obtained from the enrichments grew photoheterotrophically on cinnamate and benzoate as well as on derivatives of these compounds. Photosynthetic growth on cinnamate was greatly stimulated by addition of exogenous CO₂, and resulted in breakage of the aromatic nucleus. Growth yield studies suggested that cinnamate was converted by R. palustris to intermediates that can be quantitatively assimilated into cell material.