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Molecular identification of human glutamine- and ammonia-dependent NAD synthetases. Carbon-nitrogen hydrolase domain confers glutamine dependency.

NAD synthetase catalyzes the final step in the biosynthesis of NAD. In the present study, we obtained cDNAs for two types of human NAD synthetase (referred as NADsyn1 and NADsyn2). Structural analysis revealed in both NADsyn1 and NADsyn2 a domain required for NAD synthesis from ammonia and in only NADsyn1 an additional carbon-nitrogen hydrolase domain shared with enzymes of the nitrilase family that cleave nitriles as well as amides to produce the corresponding acids and ammonia. Consistent with the domain structures, biochemical assays indicated (i) that both NADsyn1 and NADsyn2 have NAD synthetase activity, (ii) that NADsyn1 uses glutamine as well as ammonia as an amide donor, whereas NADsyn2 catalyzes only ammonia-dependent NAD synthesis, and (iii) that mutant NADsyn1 in which Cys-175 corresponding to the catalytic cysteine residue in nitrilases was replaced with Ser does not use glutamine. Kinetic studies suggested that glutamine and ammonia serve as physiological amide donors for NADsyn1 and NADsyn2, respectively. Both synthetases exerted catalytic activity in a multimeric form. In the mouse, NADsyn1 was seen to be abundantly expressed in the small intestine, liver, kidney, and testis but very weakly in the skeletal muscle and heart. In contrast, expression of NADsyn2 was observed in all tissues tested. Therefore, we conclude that humans have two types of NAD synthetase exhibiting different amide donor specificity and tissue distributions. The ammonia-dependent synthetase has not been found in eucaryotes until this study. Our results also indicate that the carbon-nitrogen hydrolase domain is the functional domain of NAD synthetase to make use of glutamine as an amide donor in NAD synthesis. Thus, glutamine-dependent NAD synthetase may be classified as a possible glutamine amidase in the nitrilase family. Our molecular identification of NAD synthetases may prove useful to learn more of mechanisms regulating cellular NAD metabolism.

Amide Synthases↗

Tor1/2 regulation of retrograde gene expression in Saccharomyces cerevisiae derives indirectly as a consequence of alterations in ammonia metabolism.

Retrograde genes of Saccharomyces cerevisiae encode the enzymes needed to synthesize alpha-ketoglutarate, required for ammonia assimilation, when mitochondria are damaged or non-functional because of glucose fermentation. Therefore, it is not surprising that a close association exists between control of the retrograde regulon and expression of nitrogen catabolic genes. Expression of these latter genes is nitrogen catabolite repression (NCR)-sensitive, i.e. expression is low with good nitrogen sources (e.g. glutamine) and high when only poor (e.g. proline) or limiting nitrogen sources are available. It has been reported recently that both NCR-sensitive and retrograde gene expression is negatively regulated by glutamine and induced by treating cells with the Tor1/2 inhibitor, rapamycin. These conclusions predict that NCR-sensitive and retrograde gene expression should respond in parallel to nitrogen sources, ranging from those that highly repress NCR-sensitive transcription to those that elicit minimal NCR. Because this prediction did not accommodate earlier observations that CIT2 (a retrograde gene) expression is higher in glutamine than proline containing medium, we investigated retrograde regulation further. We show that (i) retrograde gene expression correlates with intracellular ammonia and alpha-ketoglutarate generated by a nitrogen source rather than the severity of NCR it elicits, and (ii) in addition to its known regulation by NCR, NAD-glutamate dehydrogenase (GDH2) gene expression is down-regulated by ammonia under conditions where NCR is minimal. Therefore, intracellular ammonia plays a pivotal dual role, regulating the interface of nitrogen and carbon metabolism at the level of ammonia assimilation and production. Our results also indicate the effects of rapamycin treatment on CIT2 transcription, and hence Tor1/2 regulation of retrograde gene expression occur indirectly as a consequence of alterations in ammonia and glutamate metabolism.

Ammonia↗

Incorporation of nitrogen into rumen bacterial fractions of steers given protein- and urea-containing diets. Ammonia assimilation into intracellular bacterial amino acids.

Experiments were carried out in vivo to investigate the pathways of ammonia incorporation into rumen bacteria, bacterial fractions and free amino acids within the bacteria. Steers were alternately given two isoenergetic, isonitrogenous diets containing the nitrogen mainly as either urea or decorticated groundnut meal (DCGM). At the end of each period on a given diet, a solution of 15NH4Cl was infused into the rumen and samples of rumen contents were removed at 2, 10, 20 and 90 min and 5, 10 and 24 h afterwards. Concentrations of ammonia and its 15N enrichment were determined and samples of mixed rumen bacteria were prepared. Bacteria were disrupted ultrasonically and separated into bacterial protein, cell wall and protein-free cell supernatant fractions. Amino acids were separated after hydrolysis and their 15N contents determined. A rumen fluid circulation pump was developed so that representative samples could be taken at very short time intervals after the introduction of the 15N label. Rumen pH changes, rumen fluid dilution rates and patterns of rumen ammonia concentrations were consistent with normal rumen metabolism. Net bacterial synthesis (as calculated from the net outflow of bacteria from the rumen) was significantly (P less than 0.05) greater with the DCGM diet (12.4 g bacterial N/d) than with the urea diet (9.24 g bacterial N/d). With both diets the 15N label rapidly left the rumen ammonia pool and entered the rumen bacteria. Analysis of the bacterial fractions indicated that the label appeared rapidly in the protein-free cell supernatant fraction and more slowly in the bacterial protein and cell wall fractions. With the DCGM diet bacteria apparently utilized intracellular label less efficiently than with the urea diet. The proportion of N in the protein-free cell supernatant was higher with the DCGM diet, suggesting increased levels of intracellular amino acids and peptides, following extracellular protein degradation. Levels of enrichment of the amino acids alanine and glutamate in the protein-free cell supernatant fraction suggested that the enzymes alanine dehydrogenase (EC 1.4.1.1) and glutamate dehydrogenase (EC 1.4.1.2 and 1.4.1.4) may be the major enzymes for assimilating ammonia when concentrations of soluble carbohydrate and rumen ammonia are high in the rumen. The high levels of intracellular alanine are discussed with reference to published work on the excretion of alanine by rumen bacteria.

Amino Acids↗

Breath ammonia depletion and its relevance to acidic aerosol exposure studies.

It is thought that gaseous ammonia in breath neutralizes acidic air pollution and thereby potentially mitigates the pulmonary effects of pollution. The efficacy of breath ammonia depletion methods reported in recent acid aerosol exposure-health response studies was investigated. Fourteen subjects (21 to 54 y of age) performed one or more of the following hygiene maneuvers: (a) acidic oral rinse (pH 2.5); (b) tooth brushing, followed by acidic oral rinse; (c) tooth brushing, followed by distilled water rinse; and (d) distilled water rinse. Initial ammonia levels ranged from 120 to 1,280 ppb (147-1,570 micrograms/m3). Acidic rinsing resulted in an immediate 90% reduction in exhaled ammonia in all subjects, and a return to 50% of baseline levels occurred within 1 h. Depletion that resulted from tooth brushing or distilled water alone was not significant. It was concluded that acidic oral rinsing is an effective method of reducing airway ammonia, but repeated oral rinsing may be required to maintain consistent, low-breath-ammonia conditions during acid aerosol exposure studies.

Adult↗

Effect of type of aviary, manure and litter handling on the emission kinetics of ammonia from layer houses.

1. An experiment with laying hens of 16 to 36 weeks of age was carried out to investigate differences in ammonia emission between 3 commercially available aviary housing systems and the additional effect of manure and litter handling. 2. Ammonia emission from Tiered Wire Floor (TWF), Natura and Boleg aviary systems increased rapidly from placement of the hens to a peak at 20 weeks of age and showed respective equilibrium level at 11.55, 11.24 (N.S. compared to TWF) and 14.55 (P < 0.001 compared to TWF) mg ammonia per h per hen. 3. Emission increased by 5.6% on the first day and 11% on subsequent days after removal of the manure on the belts. The litter layer increased up to about 7 cm; removal of 6.5 cm reduced emission by 20%. 4. Ammonia concentrations varied between 1 and 16 ppm, while ventilation rates were between 1 and 4 m3/h per hen to maintain inside temperature at about 22 degrees C. 5. About 82% of the droppings produced by the hens was found on the belts, either directly deposited there, or put there as litter material. The composition of the manure on the belts and the litter, a sand-droppings mixture, changed significantly during the first part of the laying cycle and differences were found between the 3 aviary systems with respect to the DM, pH, ash, Nkjeldahl and Total Ammoniacal Nitrogen concentration. 6. The changes in time and differences between the 3 systems in ammonia emissions and manure and litter composition were related to design of the systems, behaviour of the hens and degradation (of nitrogenous components) and volatilisation processes (water and ammonia).

Aging↗

Ammonia and endogenous benzodiazepine-like compounds in the pathogenesis of hepatic encephalopathy.

BACKGROUND: Ammonia and endogenous benzodiazepines (BDZs) are two of the most important agents among those taken into consideration in the pathogenesis of hepatic encephalopathy (HE). METHODS: Venous ammonia and endogenous BDZs sera levels were assayed in 58 liver cirrhosis patients (34 male, 24 female) free of commercial BDZs. Endogenous BDZs were measured by binding assay after high-performance liquid chromatography purification. Ammonia was assessed by colorimetric test. RESULTS: Endogenous BDZs and ammonia were significantly higher in Child-Pugh class C than in class B and class A (P < 0.05), correlating to the severity of the liver dysfunction but not with the degree of HE. A significant difference, in fact, was noted between degree 0 (no HE) versus III-IV of HE (P < 0.05), but not between degrees I-II versus III-IV. Regression analysis performed to find a correlation between the ammonia and BDZ levels in HE resulted negative. CONCLUSION: Clinical evidence is provided in cirrhotic patients that ammonia and endogenous BDZ levels do not correlate with each other in the outcome of HE.

Ammonia↗

Blood ammonia levels after intravenous infusion of glycine solution with and without ethanol.

OBJECTIVE: Absorption of glycine 1.5% during transurethral resection of the prostate may increase blood ammonia levels, but hyperammonaemia has not been described when the fluid also contained ethanol 1%. The aim of this experimental study was to evaluate whether ethanol 1% reduces glycine-induced hyperammonaemia. MATERIAL AND METHODS: Two intravenous infusions of glycine solution with and without ethanol 1% added were given on different occasions to 20 male volunteers (mean age 30 years). Half of them received 22 g of glycine over 50 min and the others approximately 18 g over 30 min. Blood ammonia was measured before and 30 min after the infusion. The serum levels of free amino acids were measured on 7 occasions during 10 of the experiments. RESULTS: The glycine infusions increased blood ammonia levels from 37 micromol/l (median, 10th and 90th percentile limits 34-53) to 57 micromol/l (27-110; p < 0.001). The change was greater after the larger glycine dose, regardless of whether the fluid contained ethanol (p < 0.05). The only amino acid concentration correlating with blood ammonia was glycine, which showed higher levels in those who had a rise in blood ammonia of 50% or more. CONCLUSIONS: Ethanol 1% did not reduce the increase in blood ammonia concentration after the administration of glycine solution.

Adult↗

Recovery of ammonia as struvite from anaerobic digester effluents.

The effects of environmental conditions on ammonia removal as struvite (Magnesium ammonium phosphate, MAP) were studied in a laboratory scale batch reactor. MAP precipitation was carried out by adding phosphoric acid and magnesium source either as MgCl, or MgO. The effect of temperature, pH, MgN:P ratios were studied. Temperature did not significantly affect ammonia removal between 25-40 degrees C and over 90% removal was obtained. The effect of pH, however,was significant and highest removal was reached at pH 8.5-9.0. The various stoichiometric ratios of ammonium to Mg and P have been tested and slight excess of Mg and P found to be beneficial for higher recovery of ammonia as struvite. However further increase in Mg and P ratios did not result in further ammonia removal which is also costly for the practical application of the process. When MgO was used as M source,the ammonia recovery was 60-70% whereas the useMgCl2 has increased this figure up to 95%. In addition a two step purification process was developed to recover MAP crystals from impurities of the anaerobic digester. Firstly, precipitates were dissolved in acid and impurities were removed by centrifugation. The clarified supernatant was re-precipitated by adjusting its pH with caustic. It was shown that in the two steps process white MAP crystals could be obtained with over 85% recovery to be used for another applications. The economical analysis of the process has shown that ammonia in the digester effluents can be recovered at the cost of $7.5-8.0 kg(-1) NH4+-N. The rate of reaction is very fast and is completed almost in minutes. This simplifies the process design resulting in a smaller reaction vessel.

Ammonia↗

Biofiltration of ammonia gas with sponge cubes coated with mixtures of activated carbon and zeolite.

Removal of ammonia gas was investigated using a biofilter system packed with small cubes of polyurethane sponge that were coated with a powder mixture of activated carbon and natural zeolite. Experimental tests and measurements include removal efficiency, pH, metabolic products of ammonia and kinetic analysis. A removal efficiency over 90% can be obtained with ammonia concentrations below 150 ppm and at contact times above 23 sec.The ammonia adsorbing power of the present biocarrier can protect the biofilter system from a high ammonia shock loading in the feed. The maximum removal rate, Vm, obtained from the kinetic analysis is 8.47 g N (kg carrier)(-1) day(-1) and the saturation constant Ks is 50.36 ppm. Nitrite is produced dominantly during the entire experiments. The cell number of nitrifying bacteria is 1.58 x 10( cell (g carrier)(-1). The present synthetic bio-carrier is considered to be one of the best among bio-carriers that have been used for the biofiltration of ammonia.

Ammonia↗

Measuring ammonia concentrations and emissions from agricultural land and liquid surfaces: a review.

Aerial ammonia concentrations (Cg) are measured using acid scrubbers, filter packs, denuders, or optical methods. Using Cg and wind speed or airflow rate, ammonia emission rate or flux can be directly estimated using enclosures or micrometeorological methods. Using nitrogen (N) recovery is not recommended, mainly because the different gaseous N components cannot be separated. Although low cost and replicable, chambers modify environmental conditions and are suitable only for comparing treatments. Wind tunnels do not modify environmental conditions as much as chambers, but they may not be appropriate for determining ammonia fluxes; however, they can be used to compare emissions and test models. Larger wind tunnels that also simulate natural wind profiles may be more useful for comparing treatments than micrometeorological methods because the latter require larger plots and are, thus, difficult to replicate. For determining absolute ammonia flux, the micrometeorological methods are the most suitable because they are nonintrusive. For use with micrometeorological methods, both the passive denuders and optical methods give comparable accuracies, although the latter give real-time Cg but at a higher cost. The passive denuder is wind weighted and also costs less than forced-air Cg measurement methods, but it requires calibration. When ammonia contamination during sample preparation and handling is a concern and separating the gas-phase ammonia and aerosol ammonium is not required, the scrubber is preferred over the passive denuder. The photothermal interferometer, because of its low detection limit and robustness, may hold potential for use in agriculture, but it requires evaluation. With its simpler theoretical basis and fewer restrictions, the integrated horizontal flux (IHF) method is preferable over other micrometeorological methods, particularly for lagoons, where berms and land-lagoon boundaries modify wind flow and flux gradients. With uniform wind flow, the ZINST method requiring measurement at one predetermined height may perform comparably to the IHF method but at a lower cost.

Agriculture↗

A placebo-controlled experimental study of steroid inhalation therapy in ammonia-induced lung injury.

BACKGROUND: The use of corticosteroids in toxic lung injury caused by exposure to an irritating gas such as ammonia has not been adequately studied. OBJECTIVE: To evaluate the effects of budesonide inhalation in a rabbit model of toxic lung injury induced by ammonia. DESIGN: Randomized, blind placebo-controlled laboratory investigation employing 16 New Zealand White rabbits. Lung injury was induced by inhalation of a defined amount of aerosolized ammonia. Thirty minutes later, the rabbits were randomized to receive either inhalation therapy with 0.5 mg budesonide or placebo. After another 2 hours, a second treatment inhalation, identical to the first one, was administered. RESULTS: Airway pressures, hemodynamics, and gas exchange were measured at baseline, 5, and 15 minutes after ammonia administration and every 30 minutes during a 6-hour period after the first blind inhalation of corticosteroids or placebo. The ammonia inhalation resulted in an acute severe lung injury, detected after 15 minutes as a decrease in Pao2 from 23.3 (+/- 3.6) to 11.0 (+/- 3.6) kPa (p < 0.005) and an increase in peak airway pressure from 13 (+/- 2) to 17 (+/- 2) cm H2O (p < 0.005). During the 6-hour observation period, the blood gas parameters improved gradually in all rabbits. In comparison with placebo, budesonide did not result in improved gas exchange or reduced airway pressure levels during the observation period. CONCLUSION: In this animal model corticosteroid inhalation therapy had no effect on ammonia-induced lung injury.

Administration, Inhalation↗

Ammonia removal from compost leachate using zeolite. II. A study using continuous flow packed columns.

Bench-scale packed zeolite columns were set up and operated to investigate the continuous removal of ammonium ions from compost leachate. The effects of hydraulic retention time (HRT), and particle size of the zeolite on the ammonia adsorption capacity were studied. For both the coarse particle and the powdered zeolite columns, higher ammonia removal efficiencies were achieved with longer HRT (i.e., lower influent flow rate) tests. At the same HRT, ammonia removal efficiencies from tests with powdered zeolite were generally 20% higher than tests with the coarse particle zeolite. A HRT of 6 hours was found appropriate for efficient ammonia removal, and an operating capacity of 1.31 mg N/g zeolite was obtained. Over 98% of the ammonia input from the influent was consistently removed for over 5 bed volumes (BV) of compost leachate flowing through the zeolite column. Zeolite proved to have a great potential as a medium for ammonia removal in treating composting leachate.

Adsorption↗

Microbial degradation of livestock-generated ammonia using biofilters at typical ambient temperatures.

The purpose of this research was to neutralize livestock-generated ammonia by using biofilters packed with inexpensive inorganic and organic packing material combined with multicultural microbial load at typical ambient temperatures. Peat and inorganic supporting materials were used as biofiltration matrix packed in a perfusion column through which gas was transfused. Results show the ammonia removal significantly fell in between 99 and 100% when ammonia concentration of 200 ppmv was used at different gas flow rates ranged from 0.030 to 0.060 m3 h(-1) at a fluctuating room temperature of 27.5 +/- 4.5 C (Mean +/- SD). Under these conditions, the emission concentration of ammonia that is liberated after biofiltration is less than 1 ppmv (0.707 mg m(-3)) over the period of our study, suggesting the usage of low-cost biofiltration systems for long-term function is effective at wider ranges of temperature fluctuations. The maximum (100%) ammonia removal efficiency was obtained in this biofilter was having an elimination capacity of 2.217 g m(-3) h(-1). This biofilter had high nitrification efficiencies and hence controlled ammonia levels with the reduced backpressure. The response of this biofilter to shut down and start up operation showed that the biofilm has a superior stability.

Air Pollutants↗

Light induces an increase in the pH of and a decrease in the ammonia concentration in the extrapallial fluid of the giant clam Tridacna squamosa.

The objective of this study was to examine whether 12 h of light exposure would lead to an increase in the pH of and a decrease in the concentration of total ammonia in the extrapallial fluid of the giant clam Tridacna squamosa. We also aimed to elucidate indirectly whether movements of ammonia and/or protons (H(+)) occurred between the extrapallial fluid and the outer mantle epithelium. The pH of the extrapallial fluid of T. squamosa exposed to 12 h of light was significantly higher than that of clams exposed to 12 h of darkness. Conversely, the total ammonia concentration in the extrapallial fluid of the former was significantly lower than that of the latter. In addition, the glutamine content in the mantle adjacent to the extrapallial fluid of clams exposed to 12 h of light was significantly greater than that of clams exposed to 12 h of darkness. These results suggest that in the extrapallial fluid of T. squamosa exposed to light, NH(3) combined with H(+) as NH(+)(4) and that NH(+)(4) was transported into the mantle and used as a substrate for glutamine formation. Injection of NH(4)Cl into the extrapallial fluid led to an instantaneous increase in the total ammonia concentration therein, but the total ammonia concentration decreased subsequently and returned to the control value within 1 h. This is in support of the proposition that NH(+)(4) could be transported from the extrapallial fluid to the mantle. Injection of HCl into the extrapallial fluid led to an instantaneous decrease in the pH of the extrapallial fluid. However, there was a significant increase in pH within 1 h in light or darkness, achieving a partial recovery toward the control pH value. The increase in pH within this 1-h period in light or darkness was accompanied by a significant decrease in the total ammonia concentration in the extrapallial fluid, which supports the proposition that H(+) could be transported in combination with NH(3) as NH(+)(4). Therefore, our results prompt a reexamination of the previous proposition that the removal of H(+) by NH(3) can facilitate calcification in molluscs in general and an investigation of the relationship between H(+) removal through NH(+)(4) transport and light-enhanced calcification in T. squamosa.

Ammonia↗

Low levels of environmental ammonia increase susceptibility to disease in Chinook salmon smolts.

Ammonia criteria are established using data from standardized toxicity tests involving healthy animals. Both intrinsic and extrinsic environmental changes affect the immune system, but few toxicity studies consider the overall impact on this system and potential changes in resistance to infection. To investigate the effects of subacute levels of ammonia in coastal waters on physiological and immunological systems of fish, juvenile Chinook salmon were maintained in seawater (10 degrees C, pH 7.8) and exposed to two concentrations of ammonia, 2.5 and 10 mg/L total nitrogen. Both test levels resulted in increased internal levels of ammonia in the fish. Neither treatment level affected feeding rates. Over a time course of 10 d, numerous significant effects were observed. White blood cell counts changed significantly, as did respiratory burst activity, plasma lysozyme activity, and plasma glucose concentration in both treatments compared to controls. In an experimental infection with Vibrio anguillarum, fish previously exposed to subacute levels of ammonia were more susceptible to pathogenic challenge. The findings of this study indicate that a more thorough investigation into the effects of environmental ammonia on fish populations in coastal waters should be undertaken and the current environmental standards reassessed.

Ammonia↗

Oxidation of ammonia and hydroxylamine to nitrate in the rat and in vitro.

We have demonstrated that ammonia is oxidized to nitrate in the rat. Male Sprague-Dawley rats gavaged with 1000 mumol N-15-ammonium chloride each day for 5 days were found to excrete low, but significant amounts of excess N-15-nitrate in their urines on the five days of treatment and on the five subsequent days. We recovered a total of 0.28 +/- 0.03 mumol excess N-15-nitrate (mean +/- SE) per rat, which indicates that ammonia is converted to nitrate in a yield of approximately 0.0080%. The oxidation of N-15-labeled glycine and L-glutamic acid to N-15-nitrate could not be detected. N-15-Hydroxylamine was oxidized in the rat to N-15-nitrate in a yield of 4.7%, which shows that hydroxylamine is a possible intermediate in the ammonia oxidation process. Injection of rats with Arochlor 1254, an inducer of several isozymes of cytochrome P-450, did not significantly affect the rate of endogeneous nitrate synthesis. Carbon tetrachloride, which causes hepatic lipid peroxidation, produced a small but significant increase in nitrate synthesis. We confirmed the observation that a bacterial endotoxin can greatly stimulate nitrate synthesis, and we showed that concurrent treatment with superoxide dismutase does not modify the effect of the endotoxin. An in vitro chemical model system was used to demonstrate that oxidation of ammonia to nitrate by the hydroxyl radical at physiological pH is chemically feasible. Our results are consistent with the hypothesis that ammonia is oxidized to nitrate in vivo by a non-enzymatic process which involves active oxygen species such as the hydroxyl radical. We estimate that a 215 g rat produces 3.0 mumol of nitrate per day via ammonia oxidation.

Ammonia↗

Features of myocardial metabolism of some amino acids and ammonia in patients with coronary artery disease.

Fractional myocardial extraction/release of glutamate, glutamine, alanine, ammonia, asparagine, glucose and lactate was studied in 12 subjects with normal coronary anatomy (controls) and 28 patients with coronary artery disease (CAD) during rest and atrial pacing. At rest patients with CAD showed an increased myocardial extraction of glutamate, glucose and lactate and an augmented glutamine and alanine release compared with controls. In all CAD patients myocardial ammonia and asparagine release was found at rest, while all controls showed myocardial extraction of these compounds. Myocardial glutamate extraction correlated positively with glucose and lactate extraction, glutamine and alanine release and inversely with ammonia release in CAD patients at rest. In patients with two- and three-vessel disease pacing-induced ischaemia resulted in a pronounced decrease in myocardial glutamate extraction and glutamine release, augmented myocardial production of ammonia and asparagine and a conversion of lactate extraction into lactate release. During pacing myocardial glutamate extraction was related to alanine and glutamine release and correlated inversely with ammonia and lactate release in these patients. The results indicate that glutamate extraction is closely connected with glucose and lactate extraction and ammonia binding via glutamine formation in the hearts of CAD patients and, thus, with the energy supply of ischaemic myocardium. An assessment of myocardial exchange of the nitrogenous compounds we have studied, complimentary to lactate, is a promising biochemical test for the identification of ischaemic heart disease in man.

Adult↗

Ammonia secretion from fish gill depends on a set of Rh glycoproteins.

Ammonia excretion from the gill in teleost fish is essential for nitrogen elimination. Although numerous physiological studies have measured ammonia excretion, the mechanism of ammonia movement through the membranes of gill epithelial cells is still unknown. Mammalian Rh glycoproteins are members of a family of proteins that mediate ammonia transport in bacteria, yeast, and plants. We identified the Rh glycoprotein homologs, fRhag, fRhbg, fRhcg1, and fRhcg2, of the pufferfish, Takifugu rubripes. Northern blot, in situ hybridization, and immunohistochemistry revealed that the pufferfish erythroid Rh glycoprotein homologue fRhag was present in red blood cells and the hematological organs (spleen and kidney) in fish. All four pufferfish Rh glycoproteins are specifically localized in the gill and line the pillar cells, pavement cells, and the mitochondrion-rich cells. Heterologous expression in Xenopus oocytes showed that they mediate methylammonium (an analog of ammonium) transport. These results suggest that pufferfish Rh glycoproteins are involved in ammonia excretion from the gill. These findings challenge the classic view that ammonia excretion in the fish gill occurs by passive diffusion.

Amino Acid Sequence↗