共查询到20条相似文献,搜索用时 31 毫秒
1.
Carol S. Surowy Nathan A. Berger 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》1985,847(3)
A series of proteins are covalently labeled when human lymphocytes are incubated with [32P]NAD+. The majority of this labeling is effectively inhibited when the lymphocytes are coincubated with 3-aminobenzamide, a potent inhibitor of poly(ADP-ribose) polymerase. However, labeling of a 72 000 molecular weight protein was resistant to the inhibitory effect of 3-aminobenzamide. Labeling of this protein from [32P]NAD+ was shown to be Mg2+-dependent. The 72 000 molecular weight protein could also be labeled on incubation with [α-32P]ATP, [γ-32P]ATP and [32P]orthophosphate, but not from [3H]NAD+ or [14C]NAD+. In the present study, we show that the 72 000 molecular weight protein is not ADP-ribosylated but rather, phosphorylated on incubation with [32P]NAD+. This phosphorylation appears to occur via an Mg2+-dependent conversion of NAD+ to AMP with the eventual utilization of the α-phosphate for phosphorylation of the 72 000 molecular weight protein. 相似文献
2.
《Journal of steroid biochemistry》1982,16(5):529-540
Male hamster kidney cytosol exhibited strong 5β-reductase activity. Incubation of cytosol with [4-14C]-testosterone at pH 6.7 yielded 5β-DHT with minor quantities of 5β-androstane-3α,17β-diol and 5β-androstane-3β,17β-diol. Incubation with [4-14C]-androstendione yielded 5β-androstanedione and smaller quantities of testosterone, 5β-DHT, 3α-hydroxy-5β-androstan-17-one, 3β-hydroxy-5β-androstan-17-one and 5β-androstane-3α,17β-diol. The two major metabolites were progressively increased with increase in the concentration of the respective substrates but the other metabolites showed very little change. The metabolism of the respective substrates was progressively decreased with changes in pH of the incubation mixture from 6.0–7.5 accompanied by a parallel decrease in the formation of the respective major metabolites. NADPH was much more effective than NADH as coenzyme. The microsomes exhibited a trace of 5β-reductase activity only with NADPH and androstenedione.The kidney homogenate at pH 10.1 effectively converted [4-14C]-testosterone to [4-14C]-androstenedione. The dehydrogenase activity was present in the cytosol and microsomes. NAD+ was more effective than NADP+ in the cytosol and the reverse was indicated for the microsomes. Spectrophotometric assay revealed not only NADP+-linked Hβ-dehydrogenase activity but also a lower 3α-dehydrogenase activity but no detectable 3β- or 17α-dehydrogenase activity. NAD+-linked activity was not explored because of the interference by the very high endogenous NAD+-reduetase activity. 相似文献
3.
Gerda Venter Frank T. J. J. Oerlemans Marieke Willemse Mietske Wijers Jack A. M. Fransen Bé Wieringa 《PloS one》2014,9(5)
Functional morphodynamic behavior of differentiated macrophages is strongly controlled by actin cytoskeleton rearrangements, a process in which also metabolic cofactors ATP and NAD(H) (i.e. NAD+ and NADH) and NADP(H) (i.e. NADP+ and NADPH) play an essential role. Whereas the link to intracellular ATP availability has been studied extensively, much less is known about the relationship between actin cytoskeleton dynamics and intracellular redox state and NAD+-supply. Here, we focus on the role of nicotinamide phosphoribosyltransferase (NAMPT), found in extracellular form as a cytokine and growth factor, and in intracellular form as one of the key enzymes for the production of NAD+ in macrophages. Inhibition of NAD+ salvage synthesis by the NAMPT-specific drug FK866 caused a decrease in cytosolic NAD+ levels in RAW 264.7 and Maf-DKO macrophages and led to significant downregulation of the glycolytic flux without directly affecting cell viability, proliferation, ATP production capacity or mitochondrial respiratory activity. Concomitant with these differential metabolic changes, the capacity for phagocytic ingestion of particles and also substrate adhesion of macrophages were altered. Depletion of cytoplasmic NAD+ induced cell-morphological changes and impaired early adhesion in phagocytosis of zymosan particles as well as spreading performance. Restoration of NAD+ levels by NAD+, NMN, or NADP+ supplementation reversed the inhibitory effects of FK866. We conclude that direct coupling to local, actin-based, cytoskeletal dynamics is an important aspect of NAD+’s cytosolic role in the regulation of morphofunctional characteristics of macrophages. 相似文献
4.
Caspar Elo Christensen Magnus Karlsson Jakob R. Winther Pernille Rose Jensen Mathilde H. Lerche 《The Journal of biological chemistry》2014,289(4):2344-2352
Accumulating evidence suggest that the pyridine nucleotide NAD has far wider biological functions than its classical role in energy metabolism. NAD is used by hundreds of enzymes that catalyze substrate oxidation and, as such, it plays a key role in various biological processes such as aging, cell death, and oxidative stress. It has been suggested that changes in the ratio of free cytosolic [NAD+]/[NADH] reflects metabolic alterations leading to, or correlating with, pathological states. We have designed an isotopically labeled metabolic bioprobe of free cytosolic [NAD+]/[NADH] by combining a magnetic enhancement technique (hyperpolarization) with cellular glycolytic activity. The bioprobe reports free cytosolic [NAD+]/[NADH] ratios based on dynamically measured in-cell [pyruvate]/[lactate] ratios. We demonstrate its utility in breast and prostate cancer cells. The free cytosolic [NAD+]/[NADH] ratio determined in prostate cancer cells was 4 times higher than in breast cancer cells. This higher ratio reflects a distinct metabolic phenotype of prostate cancer cells consistent with previously reported alterations in the energy metabolism of these cells. As a reporter on free cytosolic [NAD+]/[NADH] ratio, the bioprobe will enable better understanding of the origin of diverse pathological states of the cell as well as monitor cellular consequences of diseases and/or treatments. 相似文献
5.
Gambini J Gomez-Cabrera MC Borras C Valles SL Lopez-Grueso R Martinez-Bello VE Herranz D Pallardo FV Tresguerres JA Serrano M Viña J 《Archives of biochemistry and biophysics》2011,(1):2008-29
Sirtuins are deacetylases involved in metabolic regulation and longevity. Our aim was to test the hypothesis that they are subjected to redox regulation by the [NADH]/[NAD+] ratio. We used NIH3T3 fibroblasts in culture, Drosophila fed with or without ethanol and exercising rats. In all three models an increase in [NADH]/[NAD+] came up with an increased expression of sirtuin mRNA and protein. PGC-1α (a substrate of sirtuins) protein level was significantly increased in fibroblasts incubated with lactate and pyruvate but this effect was lost in fibroblasts obtained from sirtuin-deficient mice.We conclude that the expression of sirtuins is subject to tight redox regulation by the [NADH]/[NAD+] ratio, which is a major sensor for metabolite availability conserved from invertebrates to vertebrates. 相似文献
6.
In the present study, we show that the extracellular addition of nicotinamide adenine dinucleotide (NAD+) induces a transient rise in [Ca2+]i in human monocytes caused by an influx of extracellular calcium. The NAD+-induced Ca2+ response was prevented by adenosine triphosphate (ATP), suggesting the involvement of ATP receptors. Of the two subtypes
of ATP receptors (P2X and P2Y), the P2X receptors were considered the most likely candidates. By the use of subtype preferential
agonists and antagonists, we identified P2X1, P2X4, and P2X7 receptors being engaged in the NAD+-induced rise in [Ca2+]i. Among the P2X receptor subtypes, the P2X7 receptor is unique in facilitating the induction of nonselective pores that allow entry of ethidium upon stimulation with
ATP. In monocytes, opening of P2X7 receptor-dependent pores strongly depends on specific ionic conditions. Measuring pore formation in response to NAD+, we found that NAD+ unlike ATP lacks the ability to induce this pore-forming response. Whereas as little as 100 μM ATP was sufficient to activate
the nonselective pore, NAD+ at concentrations up to 2 mM had no effect. Taken together, these data indicate that despite similarities in the action of
extracellular NAD+ and ATP there are nucleotide-specific variations. So far, common and distinct features of the two nucleotides are only beginning
to be understood. 相似文献
7.
Barbara Kutryb-Zajac Patrycja Koszalka Ewa M. Slominska Ryszard T. Smolenski 《Nucleosides, nucleotides & nucleic acids》2018,37(11):645-652
AbstractEndothelial cell activation and dysfunction could lead to endothelial injury that is an important factor in the development of vascular diseases. Vascular injury is strongly associated with disturbed endothelial cell energetics and pyridine nucleotide pool. This study aimed to evaluate the effects of inflammatory stimuli (IL-6, LPS), uric acid, hyperglycemia, fatty acids, flavonoids, statins and nonsteroidal anti-inflammatory drugs on cellular concentration of adenosine triphosphate (ATP), adenosine diphosphate (ADP) and nicotinamide adenine dinucleotide (NAD+) in cultured endothelial cells. Murine-immortalized heart endothelial cells (H5V cells) were treated with different concentrations of pro- and anti-atherosclerotic factors and intracellular concentration of nucleotides were measured using high performance liquid chromatography. Intracellular ATP concentration in H5V cells was not changed by inflammatory stimuli (IL-6 and LPS), uric acid, glucose, atorvastatin, acetylsalicylic acid, monounsaturated and polyunsaturated fatty acids. Only high concentration of palmitic acid (1?mM) and kaempferol (>0.1?mM) decreased intracellular ATP concentration. The concentration of intracellular ADP has not been altered by any of tested compounds. In turn, intracellular NAD+ pool was modified only by polyunsaturated fatty acids and atorvastatin. Linoleic acid, docosahexaenoic acid and atorvastatin increased cellular NAD+ concentration. Tested compounds have a small influence on murine endothelial cell energetics, but polyunsaturated fatty acids and atorvastatin increased intracellular NAD+ concentration that could be an important protective mechanism against endothelial cell injury. 相似文献
8.
Nicotinamide adenine dinucleotide (NAD+) is an essential coenzyme involved in numerous physiological processes. As an attractive product in the industrial field, NAD+ also plays an important role in oxidoreductase-catalyzed reactions, drug synthesis, and the treatment of diseases, such as dementia, diabetes, and vascular dysfunction. Currently, although the biotechnology to construct NAD+-overproducing strains has been developed, limited regulation and low productivity still hamper its use on large scales. Here, we describe multi-strategy metabolic engineering to address the NAD+-production bottleneck in E. coli. First, blocking the degradation pathway of NAD(H) increased the accumulation of NAD+ by 39%. Second, key enzymes involved in the Preiss-Handler pathway of NAD+ synthesis were overexpressed and led to a 221% increase in the NAD+ concentration. Third, the PRPP synthesis module and Preiss-Handler pathway were combined to strengthen the precursors supply, which resulted in enhancement of NAD+ content by 520%. Fourth, increasing the ATP content led to an increase in the concentration of NAD+ by 170%. Finally, with the combination of all above strategies, a strain with a high yield of NAD+ was constructed, with the intracellular NAD+ concentration reaching 26.9 μmol/g DCW, which was 834% that of the parent strain. This study presents an efficient design of an NAD+-producing strain through global regulation metabolic engineering. 相似文献
9.
Jung-Chin Chang Simei Go Eduardo H. Gilglioni Suzanne Duijst Daan M. Panneman Richard J. Rodenburg Hang Lam Li Hsu-Li Huang Lonny R. Levin Jochen Buck Arthur J. Verhoeven Ronald P.J. Oude Elferink 《BBA》2021,1862(4):148367
The evolutionarily conserved soluble adenylyl cyclase (sAC, ADCY10) mediates cAMP signaling exclusively in intracellular compartments. Because sAC activity is sensitive to local concentrations of ATP, bicarbonate, and free Ca2+, sAC is potentially an important metabolic sensor. Nonetheless, little is known about how sAC regulates energy metabolism in intact cells. In this study, we demonstrated that both pharmacological and genetic suppression of sAC resulted in increased lactate secretion and decreased pyruvate secretion in multiple cell lines and primary cultures of mouse hepatocytes and cholangiocytes. The increased extracellular lactate-to-pyruvate ratio upon sAC suppression reflected an increased cytosolic free [NADH]/[NAD+] ratio, which was corroborated by using the NADH/NAD+ redox biosensor Peredox-mCherry. Mechanistic studies in permeabilized HepG2 cells showed that sAC inhibition specifically suppressed complex I of the mitochondrial respiratory chain. A survey of cAMP effectors revealed that only selective inhibition of exchange protein activated by cAMP 1 (Epac1), but not protein kinase A (PKA) or Epac2, suppressed complex I-dependent respiration and significantly increased the cytosolic NADH/NAD+ redox state. Analysis of the ATP production rate and the adenylate energy charge showed that inhibiting sAC reciprocally affects ATP production by glycolysis and oxidative phosphorylation while maintaining cellular energy homeostasis. In conclusion, our study shows that, via the regulation of complex I-dependent mitochondrial respiration, sAC-Epac1 signaling regulates the cytosolic NADH/NAD+ redox state, and coordinates oxidative phosphorylation and glycolysis to maintain cellular energy homeostasis. As such, sAC is effectively a bioenergetic switch between aerobic glycolysis and oxidative phosphorylation at the post-translational level. 相似文献
10.
A M Requero J P Díaz M S Ayuso-Parrilla R Parrilla 《Archives of biochemistry and biophysics》1979,195(1):223-234
The intraperitoneal administration of glucagon (200 μg) to rats produced a transient increase of the hepatic polypeptide chain completion time, the increase being maximum at 5 min returning to control values at 20 min. This inhibitory effect was sustained when glucagon was constantly supplied by continuous infusion. Postmitochondrial supernatants from livers of the control group or rats treated with glucagon for 5 min showed no difference in their protein synthetic activity. After 20 min of intraperitoneal administration of the hormone, that is, when the effect on protein synthesis had vanished, the levels of cAMP were still 40% above those of the control group, and the ribosomal proteins were 110% more phosphorylated. These results suggest that the observed effect of glucagon is not due to its direct action on the protein synthesis machinery. On the other hand, the variations in the hepatic amino acid content brought about by glucagon do not appear to be quantitatively significant to account for the observed inhibition of protein synthesis. The effect of glucagon was always paralleled by a decrease in the ratio which may be responsible for the observed decrease in the rates of elongation and/or termination steps of protein synthesis. Glucagon also produced a rise in the ratio in both cellular compartments, cytosol and mitochondria, as reflected by the rise in the lactate to pyruvate and the β-hydroxybutyrate to acetoacetate ratios. This shift of the NAD+ couple to a more reduced state seems to be the result of an increased mobilization and oxidation of fatty acids brought about by the hormone. It is postulated then that the primary effect of glucagon leading to a decrease in protein synthesis is probably to increase the state of reduction of the hepatic nicotinamide nucleotide system. This point of view is supported by the fact that the nicotinamide and adenine nucleotide systems in rat liver are in equilibrium through cytosolic equilibrium reactions, so that a decrease in the ratio brought about by glucagon may be secondary to the increase in the ratio. This hypothesis is supported by the fact that glucagon was not effective in inhibiting hepatic protein synthesis in rats pretreated with a drug, 2-benzene-sulfonamido-5-(β-methoxy-ethoxy)pyrimidine, that prevents fatty acid mobilization and the subsequent changes in the and ratios. Furthermore, the administration of exogenous fatty acid brings about an inhibition of the rate of hepatic protein synthesis accompanied by a decrease in the ATP levels and an increase in the state of reduction of the NAD+ system. 相似文献
11.
Magali R. VanLinden Christian D?lle Ina K. N. Pettersen Veronika A. Kulikova Marc Niere Gennaro Agrimi Sissel E. Dyrstad Ferdinando Palmieri Andrey A. Nikiforov Karl Johan Tronstad Mathias Ziegler 《The Journal of biological chemistry》2015,290(46):27644-27659
The mitochondrial NAD pool is particularly important for the maintenance of vital cellular functions. Although at least in some fungi and plants, mitochondrial NAD is imported from the cytosol by carrier proteins, in mammals, the mechanism of how this organellar pool is generated has remained obscure. A transporter mediating NAD import into mammalian mitochondria has not been identified. In contrast, human recombinant NMNAT3 localizes to the mitochondrial matrix and is able to catalyze NAD+ biosynthesis in vitro. However, whether the endogenous NMNAT3 protein is functionally effective at generating NAD+ in mitochondria of intact human cells still remains to be demonstrated. To modulate mitochondrial NAD+ content, we have expressed plant and yeast mitochondrial NAD+ carriers in human cells and observed a profound increase in mitochondrial NAD+. None of the closest human homologs of these carriers had any detectable effect on mitochondrial NAD+ content. Surprisingly, constitutive redistribution of NAD+ from the cytosol to the mitochondria by stable expression of the Arabidopsis thaliana mitochondrial NAD+ transporter NDT2 in HEK293 cells resulted in dramatic growth retardation and a metabolic shift from oxidative phosphorylation to glycolysis, despite the elevated mitochondrial NAD+ levels. These results suggest that a mitochondrial NAD+ transporter, similar to the known one from A. thaliana, is likely absent and could even be harmful in human cells. We provide further support for the alternative possibility, namely intramitochondrial NAD+ synthesis, by demonstrating the presence of endogenous NMNAT3 in the mitochondria of human cells. 相似文献
12.
Bailin Zhao Tasmin Naila Michael R Lieber Alan E Tomkinson 《Nucleic acids research》2020,48(22):12746
As nucleotidyl transferases, formation of a covalent enzyme-adenylate intermediate is a common first step of all DNA ligases. While it has been shown that eukaryotic DNA ligases utilize ATP as the adenylation donor, it was recently reported that human DNA ligase IV can also utilize NAD+ and, to a lesser extent ADP-ribose, as the source of the adenylate group and that NAD+, unlike ATP, enhances ligation by supporting multiple catalytic cycles. Since this unexpected finding has significant implications for our understanding of the mechanisms and regulation of DNA double strand break repair, we attempted to confirm that NAD+ and ADP-ribose can be used as co-factors by human DNA ligase IV. Here, we provide evidence that NAD+ does not enhance ligation by pre-adenylated DNA ligase IV, indicating that this co-factor is not utilized for re-adenylation and subsequent cycles of ligation. Moreover, we find that ligation by de-adenylated DNA ligase IV is dependent upon ATP not NAD+ or ADP-ribose. Thus, we conclude that human DNA ligase IV cannot use either NAD+ or ADP-ribose as adenylation donor for ligation. 相似文献
13.
A. M. Surin L. R. Gorbacheva I. G. Savinkova R. R. Sharipov B. I. Khodorov V. G. Pinelis 《Biochemistry. Biokhimii?a》2014,79(2):146-157
For the first time, simultaneous monitoring of changes in the concentration of cytosolic ATP ([ATP]c), pH (pHc), and intracellular free Ca2+ concentration ([Ca2+]i) of the individual neurons challenged with toxic glutamate (Glu) concentrations was performed. To this end, the ATP-sensor AT1.03, which binds to ATP and therefore enhances the efficiency of resonance energy transfer between blue fluorescent protein (energy donor) and yellow-green fluorescent protein (energy acceptor), was expressed in cultured hippocampal neurons isolated from 1–2-day-old rat pups. Excitation of fluorescence in the acceptor protein allowed monitoring changes in pHc. Cells were loaded with fluorescent low-affinity Ca2+ indicators Fura-FF or X-rhod-FF to register [Ca2+]i. It was shown that Glu (20 μM, glycine 10 μM, Mg2+-free) produced a rapid acidification of the cytosol and decrease in [ATP]c. An approximately linear relationship (r 2 = 0.56) between the rate of [ATP]c decline and latency of glutamate-induced delayed calcium deregulation (DCD) was observed: higher rate of [ATP]c decrease corresponded to shorter DCD latency period. DCD began with a decrease in [ATP]c of as much as 15.9%. In the phase of high [Ca2+]i, the plateau of [ATP]c dropped to 10.4% compared to [ATP]c in resting neurons (100%). In the presence of the Na+/K+-ATPase inhibitor ouabain (0.5 mM), glutamate-induced reduction in [ATP]c in the phase of the high [Ca2+]i plateau was only 36.6%. Changes in [ATP]c, [Ca2+]i, mitochondrial potential, and pHc in calcium-free or sodium-free buffers, as well as in the presence of the inhibitor of Na+/K+-ATPase ouabain, led us to suggest that in addition to increase in proton conductivity and decline in [ATP]c, one of the triggering factors of DCD might be a reversion of the neuronal plasma membrane Na+/Ca2+ exchange. 相似文献
14.
The enzyme NAD+synthetase [deamido-NAD+: ammonia ligase (AMP-forming), EC 6.3.1.5] is used for the preparation of 2 μmol isotopically labelled [13N]NAD+, a radiopharmaceutical designed for positron emission tomography. To obtain a rapid and high yield synthesis of [13N]NAD+, the NAD+synthetase is immobilized on porous glass beads and packed in a column. The NAD+synthetase was obtained from Escherichia coli. Different strains were tested; the cell culture technique was optimized. A new, high yield purification was applied. A screening of different immobilization techniques was done. The selected immobilization method was further optimized to increase the enzymatic activity of the enzyme-loaded glass beads. The latter were packed into a glass column. The kinetic properties of this column were investigated and optimized. 相似文献
15.
Gluconeogenesis in rat renal cortex was measured using tissue slices incubated with or without appropriate substrates. Immediately after incubation the tissue slices were snap-frozen and the content of oxidized nicotinamide adenine dinucleotide (NAD+) was determined. Incubation with 10 mM α-ketoglutarate or L-glutamate led to enhanced glucose production and an increase in tissue content of NAD+. Quinolinate and 3-mercaptopicolinate inhibited the rate of gluconeogenesis from L-glutamate and α-ketoglutarate respectively, and decreased the tissue levels of NAD+. The enhanced rate of gluconeogenesis was associated with an increase of NAD+ in the cytosol fraction (105 × g supernatant) but not in the particulate fraction (105 × g pellet) of renal cortex homogenate. Present results indicate that NAD+ content changes in parallel with the rate of gluconeogenesis in renal cortical tissue. 相似文献
16.
Ashok K. Srivastava Rafick-Pierre Sékaly Jean-Louis Chiasson 《Molecular and cellular biochemistry》1993,121(2):127-133
The effect of nicotinamide-adenine dinucleotides (NAD+ and NADP+) on Ca2+ transport in rat liver nuclei was investigated. Ca2+ uptake and release were determined with a Ca2+ electrode. Ca2+ uptake was dependent on adenosine triphosphate (ATP; 2mM). The presence of NAD+ (2mM) or NADP+ (1 and 2mM) caused a significant inhibition of Ca2+ uptake following addition of 2mM ATP. Ca2+, which accumulated in the nuclei during 6 min after ATP addition, was significantly released by the addition of NAD+ (0.5–2mM) or NADP+ (0.1–2mM). However, the effect of NADH (2mM) or NADPH (2mM) on Ca2+ uptake and release clearly weakened in comparison with the effects of NAD+ and NADP+. Meanwhile, ryanodine (10M), thapsigargin (10M) or oxalate (0.5mM) had no effect on Ca2+ uptake and release in rat liver nuclei. These reagents did not significantly alter the effects of 2mM NAD+ on Ca2+ uptake and release. Thus, NAD+ and NADP+ had a potent effect on Ca2+ transport in rat liver nuclei. The present findings suggest that the liver cytosolic NAD+ (NADP+) is a factor in the regulation of the nuclear Ca2+ concentration. (Mol Cell Biochem121: 127–133, 1993) 相似文献
17.
During axon degeneration, NAD+ levels are largely controlled by two enzymes: nicotinamide mononucleotide adenylyltransferase 2 (NMNAT2) and sterile alpha and toll interleukin motif containing protein 1 (SARM1). NMNAT2, which catalyzes the formation of NAD+ from NMN and ATP, is actively degraded leading to decreased NAD+ levels. SARM1 activity further decreases the concentration of NAD+ by catalyzing its hydrolysis to form nicotinamide and a mixture of ADPR and cADPR. Notably, SARM1 knockout mice show decreased neurodegeneration in animal models of axon degeneration, highlighting the therapeutic potential of targeting this novel NAD+ hydrolase. This review discusses recent advances in the SARM1 field, including SARM1 structure, regulation, and catalysis as well as the identification of the first SARM1 inhibitors. 相似文献
18.
Umair S Knight JS Patchett ML Bland RJ Simpson HV 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》2012,161(3):255-260
The expression of glutamate dehydrogenase (GDH; EC 1.4.1.3) in L3 of the nematode Haemonchus contortus was confirmed by detecting GDH mRNA, contrary to earlier reports. The enzyme was active in both L3 and adult H. contortus homogenates either with NAD+/H or NADP+/H as co-factor. Although it was a dual co-factor GDH, activity was greater with NAD+/H than with NADP+/H. The rate of the aminating reaction (glutamate formation) was approximately three times higher than for the deaminating reaction (glutamate utilisation). GDH provides a pathway for ammonia assimilation, although the affinity for ammonia was low. Allosteric regulation by GTP, ATP and ADP of L3 and adult H. contortus and Teladorsagia circumcincta (Nematoda) GDH depended on the concentration of the regulators and the direction of the reaction. The effects of each nucleotide were qualitatively similar on the mammalian and parasite GDH, although the nematode enzymes were more responsive to activation by ADP and ATP and less inhibited by GTP under optimum assay condition. GTP inhibited deamination and low concentrations of ADP and ATP stimulated weakly. In the reverse direction, GTP was strongly inhibitory and ADP and ATP activated the enzyme. 相似文献
19.
《Redox report : communications in free radical research》2013,18(1):28-46
AbstractIn the twentieth century, NAD+ research generated multiple discoveries. Identification of the important role of NAD+ as a cofactor in cellular respiration and energy production was followed by discoveries of numerous NAD+ biosynthesis pathways. In recent years, NAD+ has been shown to play a unique role in DNA repair and protein deacetylation. As discussed in this review, there are close interactions between oxidative stress and immune activation, energy metabolism, and cell viability in neurodegenerative disorders and ageing. Profound interactions with regard to oxidative stress and NAD+ have been highlighted in the present work. This review emphasizes the pivotal role of NAD+ in the regulation of DNA repair, stress resistance, and cell death, suggesting that NAD+ synthesis through the kynurenine pathway and/or salvage pathway is an attractive target for therapeutic intervention in age-associated degenerative disorders. NAD+ precursors have been shown to slow down ageing and extend lifespan in yeasts, and protect severed axons from degeneration in animal models neurodegenerative diseases. 相似文献
20.
A new method is described for the determination of NAD+ in picomole amounts. An enzymatic coupling system of NAD-pyrophosphorylase and hexokinase is used to convert sodium [32P]pyrophosphate and NAD+ to [32P]ADP, glucose 6-[32P]phosphate, and NMN. The key step in this analysis is the selective adsorption of the reaction product [32P]ADP, onto activated charcoal with a solution of 1m K2HPO4:10% trichloroacetic acid (1:3, v/v, pH 2). The range of concentrations of NAD+ that can be measured is 1–200 pmol. The simplicity of the method allows as many as 180 samples to be assayed in 4–5 h. This procedure has been used to quantitate NAD+ in crude extracts of germinating wheat embryos. 相似文献