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1

Rhodanese Functions as Sulfur Supplier for Key Enzymes in Sulfur Energy Metabolism  

PubMed Central

How microorganisms obtain energy is a challenging topic, and there have been numerous studies on the mechanisms involved. Here, we focus on the energy substrate traffic in the hyperthermophilic bacterium Aquifex aeolicus. This bacterium can use insoluble sulfur as an energy substrate and has an intricate sulfur energy metabolism involving several sulfur-reducing and -oxidizing supercomplexes and enzymes. We demonstrate that the cytoplasmic rhodanese SbdP participates in this sulfur energy metabolism. Rhodaneses are a widespread family of proteins known to transfer sulfur atoms. We show that SbdP has also some unusual characteristics compared with other rhodaneses; it can load a long sulfur chain, and it can interact with more than one partner. Its partners (sulfur reductase and sulfur oxygenase reductase) are key enzymes of the sulfur energy metabolism of A. aeolicus and share the capacity to use long sulfur chains as substrate. We demonstrate a positive effect of SbdP, once loaded with sulfur chains, on sulfur reductase activity, most likely by optimizing substrate uptake. Taken together, these results lead us to propose a physiological role for SbdP as a carrier and sulfur chain donor to these key enzymes, therefore enabling channeling of sulfur substrate in the cell as well as greater efficiency of the sulfur energy metabolism of A. aeolicus. PMID:22496367

Aussignargues, Clément; Giuliani, Marie-Cécile; Infossi, Pascale; Lojou, Elisabeth; Guiral, Marianne; Giudici-Orticoni, Marie-Thérčse; Ilbert, Marianne

2012-01-01

2

Lactate dehydrogenase is the key enzyme for pneumococcal pyruvate metabolism and pneumococcal survival in blood.  

PubMed

Streptococcus pneumoniae is a fermentative microorganism and causes serious diseases in humans, including otitis media, bacteremia, meningitis, and pneumonia. However, the mechanisms enabling pneumococcal survival in the host and causing disease in different tissues are incompletely understood. The available evidence indicates a strong link between the central metabolism and pneumococcal virulence. To further our knowledge on pneumococcal virulence, we investigated the role of lactate dehydrogenase (LDH), which converts pyruvate to lactate and is an essential enzyme for redox balance, in the pneumococcal central metabolism and virulence using an isogenic ldh mutant. Loss of LDH led to a dramatic reduction of the growth rate, pinpointing the key role of this enzyme in fermentative metabolism. The pattern of end products was altered, and lactate production was totally blocked. The fermentation profile was confirmed by in vivo nuclear magnetic resonance (NMR) measurements of glucose metabolism in nongrowing cell suspensions of the ldh mutant. In this strain, a bottleneck in the fermentative steps is evident from the accumulation of pyruvate, revealing LDH as the most efficient enzyme in pyruvate conversion. An increase in ethanol production was also observed, indicating that in the absence of LDH the redox balance is maintained through alcohol dehydrogenase activity. We also found that the absence of LDH renders the pneumococci avirulent after intravenous infection and leads to a significant reduction in virulence in a model of pneumonia that develops after intranasal infection, likely due to a decrease in energy generation and virulence gene expression. PMID:25245810

Gaspar, Paula; Al-Bayati, Firas A Y; Andrew, Peter W; Neves, Ana Rute; Yesilkaya, Hasan

2014-12-01

3

Subcellular distribution of key enzymes of lipid metabolism during the euthermia-hibernation-arousal cycle  

PubMed Central

Mammalian hibernation is a natural, fully reversible hypometabolic state characterized by a drastic reduction of body temperature and metabolic activity, which ensures survival to many species under adverse environmental conditions. During hibernation, many hibernators rely for energy supply almost exclusively on lipid reserves; the shift from carbohydrate to lipid metabolism implies profound rearrangement of the anabolic and catabolic pathways of energetic substrates. However, the structural counterpart of such adaptation is not known. In this study we investigated, by using immunoelectron microscopy, the fine intracellular distribution of two key enzymes involved in lipid metabolism, namely, the fatty acid synthase (FAS) and the long-chain fatty acyl-CoA synthetase (ACSL), in hepatocytes of euthermic, hibernating and arousing hazel dormice. Our results show that the two enzymes are differentially distributed in cellular compartments (cytoplasm, mitochondria and cell nuclei) of hepatocytes during euthermia. Quantitative redistribution of both enzymes among cellular compartments takes place during hibernation and arousal, in accordance with the physiological changes. Interestingly, this redistribution follows different seasonal patterns in cytoplasm, mitochondria and nuclei. In conclusion, our data represent the first quantitative morphological evidence of lipid enzyme distribution in a true hibernator throughout the year cycle, thus providing a structural framework to biochemical changes associated with the hypometabolism of hibernation. PMID:19538638

Suozzi, Anna; Malatesta, Manuela; Zancanaro, Carlo

2009-01-01

4

Regulation of sucrose metabolism in higher plants: localization and regulation of activity of key enzymes  

NASA Technical Reports Server (NTRS)

Sucrose (Suc) plays a central role in plant growth and development. It is a major end product of photosynthesis and functions as a primary transport sugar and in some cases as a direct or indirect regulator of gene expression. Research during the last 2 decades has identified the pathways involved and which enzymes contribute to the control of flux. Availability of metabolites for Suc synthesis and 'demand' for products of sucrose degradation are important factors, but this review specifically focuses on the biosynthetic enzyme sucrose-phosphate synthase (SPS), and the degradative enzymes, sucrose synthase (SuSy), and the invertases. Recent progress has included the cloning of genes encoding these enzymes and the elucidation of posttranslational regulatory mechanisms. Protein phosphorylation is emerging as an important mechanism controlling SPS activity in response to various environmental and endogenous signals. In terms of Suc degradation, invertase-catalyzed hydrolysis generally has been associated with cell expansion, whereas SuSy-catalyzed metabolism has been linked with biosynthetic processes (e.g., cell wall or storage products). Recent results indicate that SuSy may be localized in multiple cellular compartments: (1) as a soluble enzyme in the cytosol (as traditionally assumed); (2) associated with the plasma membrane; and (3) associated with the actin cytoskeleton. Phosphorylation of SuSy has been shown to occur and may be one of the factors controlling localization of the enzyme. The purpose of this review is to summarize some of the recent developments relating to regulation of activity and localization of key enzymes involved in sucrose metabolism in plants.

Winter, H.; Huber, S. C.; Brown, C. S. (Principal Investigator)

2000-01-01

5

Neuron-astrocyte interaction enhance GABAergic synaptic transmission in a manner dependent on key metabolic enzymes  

PubMed Central

Gamma aminobutric acid (GABA) is the major inhibitory neurotransmitter in the adult brain and mechanisms of GABAergic inhibition have been intensely investigated in the past decades. Recent studies provided evidence for an important role of astrocytes in shaping GABAergic currents. One of the most obvious, but yet poorly understood, mechanisms of the cross-talk between GABAergic currents and astrocytes is metabolism including neurotransmitter homeostasis. In particular, how modulation of GABAergic currents by astrocytes depends on key enzymes involved in cellular metabolism remains largely unknown. To address this issue, we have considered two simple models of neuronal culture (NC): nominally astrocyte-free NC and neuronal-astrocytic co-cultures (ANCC). Miniature Inhibitory Postsynaptic Currents (mIPSCs) were recorded in control conditions and in the presence of different enzyme blockers. We report that enrichment of NC with astrocytes results in a marked increase in mIPSC frequency. This enhancement of GABAergic activity was accompanied by increased number of GAD65 and vGAT puncta, indicating that at least a part of the frequency enhancement was due to increased number of synaptic contacts. Inhibition of glutamine synthetase (Glns) (with MSO) strongly reduced mIPSC frequency in ANCC but had no effect in NC. Moreover, treatment of ANCC with inhibitor of glycogen phosphorylase (Gys) (BAYU6751) or with selective inhibitor of astrocytic Krebs cycle, fluoroacetate, resulted in a marked reduction of mIPSC frequency in ANCC having no effect in NC. We conclude that GABAergic synaptic transmission strongly depends on neuron-astrocyte interaction in a manner dependent on key metabolic enzymes as well as on the Krebs cycle.

Kaczor, Przemys?aw; Rakus, Dariusz; Mozrzymas, Jerzy W.

2015-01-01

6

D-pinitol attenuates the impaired activities of hepatic key enzymes in carbohydrate metabolism of streptozotocin-induced diabetic rats.  

PubMed

During diabetes mellitus, endogenous hepatic glucose production is increased as a result of impaired activities of the key enzymes of carbohydrate metabolism, which leads to the condition known as hyperglycemia. D-pinitol, a bioactive constituent isolated from soybeans, has been shown to reduce hyperglycemia in experimental diabetes. We therefore designed this study to investigate the effect of oral administration of D-pinitol (50 mg/kg b. w. for 30 days) on the activities of key enzymes in carbohydrate and glycogen metabolism in the liver tissues of streptozotocin-induced diabetic rats. The efficacy was compared with glyclazide, a standard hypoglycemic drug. Oral administration of D-pinitol to diabetic group of rats showed a marked decrease in the levels of blood glucose, glycosylated hemoglobin and an increase in plasma insulin and body weight. The activities of the hepatic enzymes such as hexokinase, pyruvate kinase, glucose-6-phosphate dehydrogenase, glycogen synthase and hepatic glycogen content were significantly (p < 0.05) increased whereas the activities of glucose-6-phosphatase, fructose-1,6-bisphosphatase, lactate dehydrogenase and glycogen phosphorylase were significantly (p < 0.05) decreased in diabetic rats treated with D-pinitol. The results suggest that alterations in the activities of key metabolic enzymes of carbohydrate metabolism could be one of the biochemical rationale by which D-pinitol attenuates the hyperglycemic effect in diabetic rats. PMID:20037188

Sivakumar, Selvaraj; Subramanian, Sorimuthu P

2009-09-01

7

Immunohistochemical Localization of Key Arachidonic Acid Metabolism Enzymes during Fracture Healing in Mice  

PubMed Central

This study investigated the localization of critical enzymes involved in arachidonic acid metabolism during the initial and regenerative phases of mouse femur fracture healing. Previous studies found that loss of cyclooxygenase-2 activity impairs fracture healing while loss of 5-lipoxygenase activity accelerates healing. These diametric results show that arachidonic acid metabolism has an essential function during fracture healing. To better understand the function of arachidonic acid metabolism during fracture healing, expression of cyclooxygenase-1 (COX-1), cyclooxygenase -2 (COX-2), 5-lipoxygenase (5-LO), and leukotriene A4 hydrolase (LTA4H) was localized by immunohistochemistry in time-staged fracture callus specimens. All four enzymes were detected in leukocytes present in the bone marrow and attending inflammatory response that accompanied the fracture. In the tissues surrounding the fracture site, the proportion of leukocytes expressing COX-1, COX-2, or LTA4H decreased while those expressing 5-LO remained high at 4 and 7 days after fracture. This may indicate an inflammation resolution function for 5-LO during fracture healing. Only COX-1 was consistently detected in fracture callus osteoblasts during the later stages of healing (day 14 after fracture). In contrast, callus chondrocytes expressed all four enzymes, though 5-LO appeared to be preferentially expressed in newly differentiated chondrocytes. Most interestingly, osteoclasts consistently and strongly expressed COX-2. In addition to bone surfaces and the growth plate, COX-2 expressing osteoclasts were localized at the chondro-osseous junction of the fracture callus. These observations suggest that arachidonic acid mediated signaling from callus chondrocytes or from callus osteoclasts at the chondro-osseous junction regulate fracture healing. PMID:24516658

Lin, Hsuan-Ni; O’Connor, J. Patrick

2014-01-01

8

Efficacy of 20-OH-ecdysone on hepatic key enzymes of carbohydrate metabolism in streptozotocin induced diabetic rats.  

PubMed

The aim of the present investigation was to evaluate the anti-diabetic activity of 20-OH-ecdysone on glucose metabolic key enzymes in control and streptozotocin induced diabetic rats. On oral administration of 20-OH-ecdysone at a dose of 5mg/kg body weight per day to diabetic rats for 30 days resulted in a significant decrease in the levels of plasma glucose, glycosylated hemoglobin (HbA1c) and an increase in the levels of insulin and hemoglobin. Administration of 20-OH-ecdysone showed significant increase in the levels of glycolytic enzyme (hexokinase) and hepatic shunt enzyme (glucose-6-phosphate dehydrogenase) whereas significant decrease in the levels of gluconeogenic enzymes (glucose-6-phosphatase and fructose-1,6-bisphosphatase) in diabetic treated rats. Furthermore, protection against body weight loss of diabetic animals also observed. This study indicates that the administration of 20-OH-ecdysone to diabetic rats resulted in alterations in the metabolism of glucose with subsequent reduction in plasma glucose levels. A comparison was made between the action of 20-OH-ecdysone and antidiabetic drug-glibenclamide. The effects produced by the 20-OH-ecdysone were comparable to that of glibenclamide. PMID:22484004

Sundaram, Ramalingam; Naresh, Rajendran; Shanthi, Palanivelu; Sachdanandam, Panchanatham

2012-06-15

9

Efficacy of azelaic acid on hepatic key enzymes of carbohydrate metabolism in high fat diet induced type 2 diabetic mice.  

PubMed

Azelaic acid (AzA), a C9 linear ?,?-dicarboxylic acid, is found in whole grains namely wheat, rye, barley, oat seeds and sorghum. The study was performed to investigate whether AzA exerts beneficial effect on hepatic key enzymes of carbohydrate metabolism in high fat diet (HFD) induced type 2 diabetic C57BL/6J mice. C57BL/6J mice were fed high fat diet for 10 weeks and subjected to intragastric administration of various doses (20 mg, 40 mg and 80 mg/kg BW) of AzA daily for the subsequent 5 weeks. Rosiglitazone (RSG) was used as reference drug. Body weight, food intake, plasma glucose, plasma insulin, blood haemoglobin (Hb), blood glycosylated haemoglobin (HbA1c), liver glycolytic enzyme (hexokinase), hepatic shunt enzyme (glucose-6-phosphate dehydrogenase), gluconeogenic enzymes(glucose-6-phosphatase and fructose-1,6-bisphosphatase), liver glycogen, plasma and liver triglycerides were examined in mice fed with normal standard diet (NC), high fat diet (HFD), HFD with AzA (HFD + AzA) and HFD with rosiglitazone (HFD + RSG). Among the three doses, 80 mg/kg BW of AzA was able to positively regulate plasma glucose, insulin, blood HbA1c and haemoglobin levels by significantly increasing the activity of hexokinase and glucose-6-phosphate dehydrogenase and significantly decreasing the activity of glucose-6-phosphatase and fructose-1,6-bisphosphatase thereby increasing the glycogen content in the liver. From this study, we put forward that AzA could significantly restore the levels of plasma glucose, insulin, HbA1c, Hb, liver glycogen and carbohydrate metabolic key enzymes to near normal in diabetic mice and hence, AzA may be useful as a biomaterial in the development of therapeutic agents against high fat diet induced T2DM. PMID:23402910

Muthulakshmi, Shanmugam; Saravanan, Ramalingam

2013-06-01

10

Shared origins of a key enzyme during the evolution of C4 and CAM metabolism  

PubMed Central

CAM and C4 photosynthesis are two key plant adaptations that have evolved independently multiple times, and are especially prevalent in particular groups of plants, including the Caryophyllales. We investigate the origin of photosynthetic PEPC, a key enzyme of both the CAM and C4 pathways. We combine phylogenetic analyses of genes encoding PEPC with analyses of RNA sequence data of Portulaca, the only plants known to perform both CAM and C4 photosynthesis. Three distinct gene lineages encoding PEPC exist in eudicots (namely ppc-1E1, ppc-1E2 and ppc-2), one of which (ppc-1E1) was recurrently recruited for use in both CAM and C4 photosynthesis within the Caryophyllales. This gene is present in multiple copies in the cacti and relatives, including Portulaca. The PEPC involved in the CAM and C4 cycles of Portulaca are encoded by closely related yet distinct genes. The CAM-specific gene is similar to genes from related CAM taxa, suggesting that CAM has evolved before C4 in these species. The similar origin of PEPC and other genes involved in the CAM and C4 cycles highlights the shared early steps of evolutionary trajectories towards CAM and C4, which probably diverged irreversibly only during the optimization of CAM and C4 phenotypes. PMID:24638902

Christin, Pascal-Antoine; Arakaki, Monica; Osborne, Colin P.; Bräutigam, Andrea; Sage, Rowan F.; Hibberd, Julian M.; Kelly, Steven; Covshoff, Sarah; Wong, Gane Ka-Shu; Hancock, Lillian; Edwards, Erika J.

2014-01-01

11

The structure of vanin 1: a key enzyme linking metabolic disease and inflammation.  

PubMed

Although part of the coenzyme A pathway, vanin 1 (also known as pantetheinase) sits on the cell surface of many cell types as an ectoenzyme, catalyzing the breakdown of pantetheine to pantothenic acid (vitamin B5) and cysteamine, a strong reducing agent. Vanin 1 was initially discovered as a protein involved in the homing of leukocytes to the thymus. Numerous studies have shown that vanin 1 is involved in inflammation, and more recent studies have shown a key role in metabolic disease. Here, the X-ray crystal structure of human vanin 1 at 2.25?Ĺ resolution is presented, which is the first reported structure from the vanin family, as well as a crystal structure of vanin 1 bound to a specific inhibitor. These structures illuminate how vanin 1 can mediate its biological roles by way of both enzymatic activity and protein-protein interactions. Furthermore, it sheds light on how the enzymatic activity is regulated by a novel allosteric mechanism at a domain interface. PMID:25478849

Boersma, Ykelien L; Newman, Janet; Adams, Timothy E; Cowieson, Nathan; Krippner, Guy; Bozaoglu, Kiymet; Peat, Thomas S

2014-12-01

12

Trans-anethole, a terpenoid ameliorates hyperglycemia by regulating key enzymes of carbohydrate metabolism in streptozotocin induced diabetic rats.  

PubMed

Trans-anethole (TA), a terpenoid and a principle constituent of many essential oils from medicinal plants possess hypoglycemic and antioxidant activities. This study was undertaken to explore beneficial effects of TA on key enzymes of carbohydrate metabolism in streptozotocin (STZ)-induced type 2 diabetic rats. Diabetes was induced in male albino Wistar rats by intraperitoneal administration of STZ (40 mg/kg BW). TA was administered to diabetic rats at a dose of 20, 40 and 80 mg/kg BW for 45 days. However, the dose at 80 mg/kg BW, resulted in a significant reduction in the levels of plasma glucose, glycosylated haemoglobin (HbA1c) and increase in the levels of insulin and haemoglobin (Hb). Upon administration of TA, the altered levels of liver glycolytic enzyme (hexokinase), hepatic shunt enzyme (glucose-6-phosphate dehydrogenase) and gluconeogenic enzymes (glucose-6-phosphatase and fructose-1,6-bisphosphatase) in the liver and kidney of diabetic rats significantly reverted to near normal levels. In addition to this, TA also improved the hepatic and muscle glycogen content in diabetic rats. The histological studies showed the ameliorative effect of TA on the ?-cells of pancreas in diabetic rats. The results were compared with glibenclamide, a standard oral hypoglycemic drug. These encouraging findings suggest that TA may be used as a propitious bioactive compound in the development of therapeutic agents against type 2 diabetes mellitus. PMID:25708856

Sheikh, Bashir Ahmad; Pari, Leelavinothan; Rathinam, Ayyasamy; Chandramohan, Ramasamy

2015-05-01

13

Key Residues Controlling Phenacetin Metabolism By Human Cytochrome P450 2A Enzymes  

SciTech Connect

Although the human lung cytochrome P450 2A13 (CYP2A13) and its liver counterpart cytochrome P450 2A6 (CYP2A6) are 94% identical in amino acid sequence, they metabolize a number of substrates with substantially different efficiencies. To determine differences in binding for a diverse set of cytochrome P450 2A ligands, we have measured the spectral binding affinities (K{sub D}) for nicotine, phenethyl isothiocyanate (PEITC), coumarin, 2{prime}-methoxyacetophenone (MAP), and 8-methoxypsoralen. The differences in the K{sub D} values for CYP2A6 versus CYP2A13 ranged from 74-fold for 2{prime}-methoxyacetophenone to 1.1-fold for coumarin, with CYP2A13 demonstrating the higher affinity. To identify active site amino acids responsible for the differences in binding of MAP, PEITC, and coumarin, 10 CYP2A13 mutant proteins were generated in which individual amino acids from the CYP2A6 active site were substituted into CYP2A13 at the corresponding position. Titrations revealed that substitutions at positions 208, 300, and 301 individually had the largest effects on ligand binding. The collective relevance of these amino acids to differential ligand selectivity was verified by evaluating binding to CYP2A6 mutant enzymes that incorporate several of the CYP2A13 amino acids at these positions. Inclusion of four CYP2A13 amino acids resulted in a CYP2A6 mutant protein (I208S/I300F/G301A/S369G) with binding affinities for MAP and PEITC much more similar to those observed for CYP2A13 than to those for CYP2A6 without altering coumarin binding. The structure-based quantitative structure-activity relationship analysis using COMBINE successfully modeled the observed mutant-ligand trends and emphasized steric roles for active site residues including four substituted amino acids and an adjacent conserved Leu{sup 370}.

DeVore, N.M.; Smith, B.D.; Urban, M.J.; Scott, E.E.

2009-05-14

14

Tyrosol, a phenolic compound, ameliorates hyperglycemia by regulating key enzymes of carbohydrate metabolism in streptozotocin induced diabetic rats.  

PubMed

The present study was designed to evaluate the effects of tyrosol, a phenolic compound, on the activities of key enzymes of carbohydrate metabolism in the control and streptozotocin-induced diabetic rats. Diabetes mellitus was induced in rats by a single intraperitoneal injection of streptozotocin (40mg/kg body weight). Experimental rats were administered tyrosol 1ml intra gastrically at the doses of 5, 10 and 20mg/kg body weight and glibenclamide 1ml at a dose of 600?g/kg body weight once a day for 45days. At the end of the experimental period, diabetic control rats exhibited significant (p<0.05) increase in plasma glucose, glycosylated hemoglobin with significant (p<0.05) decrease in plasma insulin, total hemoglobin and body weight. The activities of key enzymes of carbohydrate metabolism such as phosphoenolpyruvate carboxykinase, fructose-1,6-bisphosphatase and glucose-6-phosphatase were significantly (p<0.05) increased and the activities of hexokinase and glucose-6-phosphate dehydrogenase were significantly (p<0.05) decreased in the liver and kidney of diabetic control rats. Further, antioxidants were lowered in diabetic control rats. A significant (p<0.05) decline in glycogen level in the liver and muscle and glycogen synthase activity in the liver and a significant (p<0.05) increase in the activity of liver glycogen phosphorylase were observed in diabetic control rats compared to normal control rats. Oral administration of tyrosol to diabetic rats reversed all the above mentioned biochemical parameters to near normal in a dose dependent manner. Tyrosol at a dose of 20mg/kg body weight showed the highest significant effect than the other two doses. Immunohistochemical staining of pancreas revealed that tyrosol treated diabetic rats showed increased insulin immunoreactive ?-cells, which confirmed the biochemical findings. The observed results were compared with glibenclamide, a standard oral hypoglycemic drug. The results of the present study suggest that tyrosol decreases hyperglycemia, by its antioxidant effect. PMID:25641191

Chandramohan, Ramasamy; Pari, Leelavinothan; Rathinam, Ayyasamy; Sheikh, Bashir Ahmad

2015-03-01

15

Inhibitory Potential of Turbinaria ornata against Key Metabolic Enzymes Linked to Diabetes  

PubMed Central

One of the therapeutic approaches in treating diabetes is to reduce postprandial hyperglycemia by inhibiting major carbohydrate hydrolyzing enzymes. In the present study, crude extracts of marine seaweed, Turbinaria ornata, were tested for their antidiabetic potential using enzyme inhibitory assays (?-amylase, ?-glucosidase, and dipeptidyl peptidase-IV). Among the tested extracts, methanol and acetone extracts showed significant inhibitory effects on ?-amylase (IC50 250.9??g/mL), ?-glucosidase (535.6??g/mL), and dipeptidyl peptidase-4 (55.2??g/mL), respectively. Free radical scavenging activity of these extracts was analyzed using DPPH assay (65%). Extracts were tested for in vitro toxicity using DNA fragmentation assay, haemolytic assay, and MTT assay. None of the extracts showed toxicity in tested models. Furthermore, GC-MS analysis of lead extracts showed the presence of major compounds, hentriacontane, z, z-6, 28-heptatriactontadien-2-one, 8-heptadecene, and 1-heptacosanol. Our findings suggest that Turbinaria ornata can be used as a potential source for further in vivo studies in controlling hyperglycemia. PMID:25050371

Unnikrishnan, P. S.; Suthindhiran, K.; Jayasri, M. A.

2014-01-01

16

A key enzyme in the biogenesis of lysosomes is a protease that regulates cholesterol metabolism.  

PubMed

Mucolipidosis II is a severe lysosomal storage disorder caused by defects in the ? and ? subunits of the hexameric N-acetylglucosamine-1-phosphotransferase complex essential for the formation of the mannose 6-phosphate targeting signal on lysosomal enzymes. Cleavage of the membrane-bound ?/?-subunit precursor by an unknown protease is required for catalytic activity. Here we found that the ?/?-subunit precursor is cleaved by the site-1 protease (S1P) that activates sterol regulatory element-binding proteins in response to cholesterol deprivation. S1P-deficient cells failed to activate the ?/?-subunit precursor and exhibited a mucolipidosis II-like phenotype. Thus, S1P functions in the biogenesis of lysosomes, and lipid-independent phenotypes of S1P deficiency may be caused by lysosomal dysfunction. PMID:21719679

Marschner, Katrin; Kollmann, Katrin; Schweizer, Michaela; Braulke, Thomas; Pohl, Sandra

2011-07-01

17

Effect of T. foenumgraecum on glycogen content of tissues and the key enzymes of carbohydrate metabolism.  

PubMed

The Indian traditional system of medicine prescribed plant therapies for diseases including diabetes mellitus called madhumeh in Sanskrit. One such plant mentioned in Ayurveda is Trigonella foenumgraecum (FG). In the present study, FG (1g/kg PO) was assessed for its effect on glycogen levels of insulin dependent (skeletal muscle and liver), insulin independent tissues (kidneys and brain) and enzymes such as glucokinase (GK), hexokinase (HK), and phosphofructokinase (PFK). Administration of FG led to decrease in blood glucose levels by 14.4 and 46.64% on 15th and 30th day of the experiment. Liver and 2-kidney weight expressed as percentage of body weight was significantly increased in diabetics (P<0.0005) versus normal controls and this alteration in the renal weight (P<0.0005) but not liver weight was normalized by feeding of FG. Renal glycogen content increased by over 10 folds while hepatic and skeletal muscle glycogen content decreased by 75 and 68% in diabetic controls versus controls and these alteration in glycogen content was partly prevented by FG. Activity of HK, GK and PFK in diabetic controls was 35, 50 and 60% of the controls and FG partially corrected this alteration in PFK, HK and GK. PMID:12639747

Vats, V; Yadav, S P; Grover, J K

2003-04-01

18

Metabolic enzyme expression highlights a key role for MTHFD2 and the mitochondrial folate pathway in cancer  

NASA Astrophysics Data System (ADS)

Metabolic remodeling is now widely regarded as a hallmark of cancer, but it is not clear whether individual metabolic strategies are frequently exploited by many tumours. Here we compare messenger RNA profiles of 1,454 metabolic enzymes across 1,981 tumours spanning 19 cancer types to identify enzymes that are consistently differentially expressed. Our meta-analysis recovers established targets of some of the most widely used chemotherapeutics, including dihydrofolate reductase, thymidylate synthase and ribonucleotide reductase, while also spotlighting new enzymes, such as the mitochondrial proline biosynthetic enzyme PYCR1. The highest scoring pathway is mitochondrial one-carbon metabolism and is centred on MTHFD2. MTHFD2 RNA and protein are markedly elevated in many cancers and correlated with poor survival in breast cancer. MTHFD2 is expressed in the developing embryo, but is absent in most healthy adult tissues, even those that are proliferating. Our study highlights the importance of mitochondrial compartmentalization of one-carbon metabolism in cancer and raises important therapeutic hypotheses.

Nilsson, Roland; Jain, Mohit; Madhusudhan, Nikhil; Sheppard, Nina Gustafsson; Strittmatter, Laura; Kampf, Caroline; Huang, Jenny; Asplund, Anna; Mootha, Vamsi K.

2014-01-01

19

Metabolic enzyme expression highlights a key role for MTHFD2 and the mitochondrial folate pathway in cancer  

PubMed Central

Metabolic remodeling is now widely regarded as a hallmark of cancer, but it is not clear whether individual metabolic strategies are frequently exploited by many tumours. Here we compare messenger RNA profiles of 1,454 metabolic enzymes across 1,981 tumours spanning 19 cancer types to identify enzymes that are consistently differentially expressed. Our meta-analysis recovers established targets of some of the most widely used chemotherapeutics, including dihydrofolate reductase, thymidylate synthase and ribonucleotide reductase, while also spotlighting new enzymes, such as the mitochondrial proline biosynthetic enzyme PYCR1. The highest scoring pathway is mitochondrial one-carbon metabolism and is centred on MTHFD2. MTHFD2 RNA and protein are markedly elevated in many cancers and correlated with poor survival in breast cancer. MTHFD2 is expressed in the developing embryo, but is absent in most healthy adult tissues, even those that are proliferating. Our study highlights the importance of mitochondrial compartmentalization of one-carbon metabolism in cancer and raises important therapeutic hypotheses. PMID:24451681

Nilsson, Roland; Jain, Mohit; Madhusudhan, Nikhil; Sheppard, Nina Gustafsson; Strittmatter, Laura; Kampf, Caroline; Huang, Jenny; Asplund, Anna; Mootha, Vamsi K

2014-01-01

20

Reduced lipid intake leads to changes in digestive enzymes in the intestine but has minor effects on key enzymes of hepatic intermediary metabolism in rainbow trout ( Oncorhynchus mykiss).  

PubMed

For sustainable aquaculture, the removal of marine resource ingredients in fish diets is an important objective. While most studies focus on the replacement of fish oil by vegetable oil, little is known on the nutritional effects of presence (which corresponds to the control diet) or absence of dietary fish oil. We studied fatty acid composition of brush-border membranes and digestive enzyme activities of the intestine and measured the expression and activities of several enzymes involved in the hepatic intermediary metabolism of rainbow trout (Oncorhynchus mykiss) fed for 7 weeks with or without fish oil. The diets were pair-fed to ensure that fish fed either diet had comparable carbohydrate and protein intakes. Absence of fish oil significantly reduced growth rate, protein efficiency and plasma lipid components. Activities of intestinal digestive enzymes were significantly decreased in the anterior intestine in fish fed without fish oil. In liver, dietary fish oil removal did not affect the transcript levels or activities of the main enzymes involved in lipogenesis (fatty acid synthase) and fatty acid ?-oxidation (3-hydroxyacyl-CoA dehydrogenase), glycolysis or amino acid oxidation. It lowered the expression of the genes coding for gluconeogenic enzymes (glucose-6-phosphatase and phosphoenolpyruvate carboxykinase), but their enzyme activities were not affected. The activities, but not gene expression of lipogenic enzymes, involved in NADPH and malonyl-CoA formation were also modified after fish oil removal as reflected by higher activities of isocitrate dehydrogenase/glucose-6-phosphate dehydrogenase and acetyl-CoA carboxylase enzymes. Overall, our results indicate that the intestinal digestive capacity was strongly modified by dietary fish oil removal, while hepatic intermediary metabolism was only marginally affected, in fed rainbow trout. PMID:22444883

Ducasse-Cabanot, S; Zambonino-Infante, J; Richard, N; Medale, F; Corraze, G; Mambrini, M; Robin, J; Cahu, C; Kaushik, S; Panserat, S

2007-10-01

21

In vitro inhibitory potential of Cynara scolymus, Silybum marianum, Taraxacum officinale, and Peumus boldus on key enzymes relevant to metabolic syndrome.  

PubMed

Boldocynara®, a proprietary dietary supplement product consisting of the plants Cynara scolymus, Silybum marianum, Taraxacum officinale, and Peumus boldus, used to promote functions of the liver and the gallbladder. It was the aim of the present study to look from a different perspective at the product by investigating the in vitro potential of Boldocynara® as a combination product and its individual extracts on key enzymes relevant to metabolic syndrome. Peumus boldus extract exhibited pronounced inhibitory activities on ?-glucosidase (80% inhibition at 100 µg/ml, IC50: 17.56 µg/ml). Silybum marianum had moderate pancreatic lipase (PL) inhibitory activities (30% at 100 µg/ml) whereas Cynara scolymus showed moderate ACE inhibitory activity (31% at 100 µg/ml). The combination had moderate to weak effects on the tested enzymes. In conclusion, our results indicate some moderate potential of the dietary supplement Boldocynara® and its single ingredients for the prevention of metabolic disorders. PMID:25636882

Villiger, Angela; Sala, Filippo; Suter, Andy; Butterweck, Veronika

2015-01-15

22

The metabolic acclimation of Arabidopsis thaliana to arsenate is sensitized by the loss of mitochondrial LIPOAMIDE DEHYDROGENASE2, a key enzyme in oxidative metabolism.  

PubMed

Mitochondrial lipoamide dehydrogenase is essential for the activity of four mitochondrial enzyme complexes central to oxidative metabolism. The reduction in protein amount and enzyme activity caused by disruption of mitochondrial LIPOAMIDE DEHYDROGENASE2 enhanced the arsenic sensitivity of Arabidopsis thaliana. Both arsenate and arsenite inhibited root elongation, decreased seedling size and increased anthocyanin production more profoundly in knockout mutants than in wild-type seedlings. Arsenate also stimulated lateral root formation in the mutants. The activity of lipoamide dehydrogenase in isolated mitochondria was sensitive to arsenite, but not arsenate, indicating that arsenite could be the mediator of the observed phenotypes. Steady-state metabolite abundances were only mildly affected by mutation of mitochondrial LIPOAMIDE DEHYDROGENASE2. In contrast, arsenate induced the remodelling of metabolite pools associated with oxidative metabolism in wild-type seedlings, an effect that was enhanced in the mutant, especially around the enzyme complexes containing mitochondrial lipoamide dehydrogenase. These results indicate that mitochondrial lipoamide dehydrogenase is an important protein for determining the sensitivity of oxidative metabolism to arsenate in Arabidopsis. PMID:23961884

Chen, Weihua; Taylor, Nicolas L; Chi, Yingjun; Millar, A Harvey; Lambers, Hans; Finnegan, Patrick M

2014-03-01

23

Modulatory effect of green tea extract on hepatic key enzymes of glucose metabolism in streptozotocin and high fat diet induced diabetic rats.  

PubMed

The study was undertaken to evaluate the antidiabetic effect of green tea extract on carbohydrate metabolic key enzymes in control and streptozotocin high fat diet -induced diabetic rats. The daily oral treatment of green tea extract (300 mg/kg body weight) to diabetic rats for 30 days resulted in a significant reduction in the levels of plasma glucose, glycosylated hemoglobin (HbA1c) and increase in the levels of insulin and hemoglobin. The altered activities of the key enzymes of carbohydrate metabolism such as hexokinase, pyruvate kinase, lactate dehydrogenase, glucose-6-phosphatase, fructose-1,6-bisphosphatase, glucose-6-phosphate dehydrogenase, glycogen synthase and glycogen phosphorylase in liver of diabetic rats were significantly reverted to near normal levels by the administration of green tea extract. Further, green tea extract administration to diabetic rats improved muscle and hepatic glycogen content suggesting the antihyperglycemic potential of green tea extract in diabetic rats. The obtained results were compared with metformin, a standard oral hypoglycemic drug. Thus, this study indicates that the administration of green tea extract to diabetic rats resulted in alterations in the metabolism of glucose with subsequent reduction in plasma glucose levels. PMID:23453307

Sundaram, Ramalingam; Naresh, Rajendran; Shanthi, Palanivelu; Sachdanandam, Panchanatham

2013-05-15

24

Effects of myogenin on muscle fiber types and key metabolic enzymes in gene transfer mice and C2C12 myoblasts.  

PubMed

Skeletal muscle fiber type composition is one of the important factors influencing muscle growth and meat quality. As a member of the myogenic transcription factors, myogenin (MyoG) is required for embryonic myoblast differentiation, but the expression of MyoG continues in mature muscle tissue of adult animals, especially in oxidative metabolic muscle, which suggests that MyoG may play a more extended role. Therefore, using MyoG gene transfer mice and C2C12 myoblasts as in vivo and in vitro models, respectively, we elected to study the role of MyoG in muscle fiber types and oxidative metabolism by using overexpression and siRNA suppression strategies. The overexpression of MyoG by DNA electroporation in mouse gastrocnemius muscle had no significant effect on fiber type composition but upregulated the mRNA expression (P<0.01) and enzyme activity (P<0.05) of oxidative succinic dehydrogenase (SDH). In addition, downregulation of the activity of the glycolytic enzymes lactate dehydrogenase (LDH, P<0.05) and pyruvate kinase (PK, P<0.05) was observed in MyoG gene transfer mice. In vitro experiments verified the results obtained in mice. Stable MyoG-transfected differentiating C2C12 cells showed higher mRNA expression levels of myosin heavy chain (MyHC) isoform IIX (P<0.01) and SDH (P<0.05), while the LDH mRNA was attenuated. The enzyme activities of SDH (P<0.01) and LDH (P<0.05) were similarly altered at the mRNA level. When MyoG was knocked down in C2C12 cells, MyHC IIX expression (P<0.05) was decreased, but the mRNA level (P<0.05) and the enzyme activity (P<0.05) of SDH were increased. Downregulating MyoG also increased the activity of the glycolytic enzymes PK (P<0.05) and hexokinase (HK, P<0.05). Based on those results, we concluded that MyoG barely changes the MyHC isoforms, except MyHC IIX, in differentiating myoblasts but probably influences the shift from glycolytic metabolism towards oxidative metabolism both in vivo and in vitro. These results contribute to further understand the role of MyoG in skeletal muscle energy metabolism and also help to explore the key genes that regulate meat quality. PMID:24055422

Zhu, Lin-Na; Ren, Yang; Chen, Jing-Qing; Wang, Yi-Zhen

2013-12-15

25

Dithioerythritol (DTE) prevents inhibitory effects of triphenyltin (TPT) on the key enzymes of the human sex steroid hormone metabolism  

Microsoft Academic Search

Organotins are known to induce imposex (pseudohermaphroditism) in marine neogastropods and are suggested to act as specific endocrine disruptors, inhibiting the enzyme-mediated conversion of steroid hormones. Therefore, we investigated the in vitro effects of triphenyltin (TPT) on human 5?-reductase type 2 (5?-Re 2), cytochrome P450 aromatase (P450arom), 17?-hydroxysteroid dehydrogenase type 3 (17?-HSD 3), 3?-HSD type 2 and 17?-HSD type 1

Susan Lo; Axel Alléra; Peter Albers; Jörg Heimbrecht; Eckard Jantzen; Dietrich Klingmüller; Stephan Steckelbroeck

2003-01-01

26

Effect of feeding aqueous extract of Pterocarpus marsupium on glycogen content of tissues and the key enzymes of carbohydrate metabolism.  

PubMed

The Indian traditional system of medicine prescribed plant therapies for diseases including diabetes mellitus called madhumeh in Sanskrit. One such plant mentioned in Ayurveda is Pterocarpus marsupium (PM). In the present study, aqueous extract of PM (1 g/kg PO) was assessed for its effect on glycogen levels of insulin dependent (skeletal muscle and liver), insulin-independent tissues (kidneys and brain) and enzymes such as glucokinase (GK), hexokinase (HK), and phosphofructokinase (PFK). Administration of PM led to decrease in blood glucose levels by 38 and 60% on 15th and 30th day of the experiment. Liver and 2-kidney weight expressed as percentage of body-weight was significantly increased in diabetics (p < 0.0005) vs. normal controls and this alteration in the renal weight (p < 0.0005) but not liver weight was normalized by feeding of PM extract. Renal glycogen content increased by over 10-fold while hepatic and skeletal muscle glycogen content decreased by 75 and 68% in diabetic controls vs. controls and these alteration in glycogen content was partly prevented by PM. Activity of HK, GK and PFK in diabetic controls was 35,50 and 60% of the controls and PM completely corrected this alteration in PFK and only partly in HK and GK. PMID:12482025

Grover, Jagdish Kumari; Vats, Vikrant; Yadav, Satyapal

2002-12-01

27

Chicken Cytochrome P450 1A5 Is the Key Enzyme for Metabolizing T-2 Toxin to 3?OH-T-2  

PubMed Central

The transmission of T-2 toxin and its metabolites into the edible tissues of poultry has potential effects on human health. We report that T-2 toxin significantly induces CYP1A4 and CYP1A5 expression in chicken embryonic hepatocyte cells. The enzyme activity assays of CYP1A4 and CYP1A5 heterologously expressed in HeLa cells indicate that only CYP1A5 metabolizes T-2 to 3?OH-T-2 by the 3?-hydroxylation of isovaleryl groups. In vitro enzyme assays of recombinant CYP1A5 expressed in DH5? further confirm that CYP1A5 can convert T-2 into TC-1 (3?OH-T-2). Therefore, CYP1A5 is critical for the metabolism of trichothecene mycotoxin in chickens. PMID:23702848

Shang, Shufeng; Jiang, Jun; Deng, Yiqun

2013-01-01

28

A Quaternary Mechanism Enables the Complex Biological Functions of Octameric Human UDP-glucose Pyrophosphorylase, a Key Enzyme in Cell Metabolism.  

PubMed

In mammals, UDP-glucose pyrophosphorylase (UGP) is the only enzyme capable of activating glucose-1-phosphate (Glc-1-P) to UDP-glucose (UDP-Glc), a metabolite located at the intersection of virtually all metabolic pathways in the mammalian cell. Despite the essential role of its product, the molecular basis of UGP function is poorly understood. Here we report the crystal structure of human UGP in complex with its product UDP-Glc. Beyond providing first insight into the active site architecture, we describe the substrate binding mode and intermolecular interactions in the octameric enzyme that are crucial to its activity. Importantly, the quaternary mechanism identified for human UGP in this study may be common for oligomeric sugar-activating nucleotidyltransferases. Elucidating such mechanisms is essential for understanding nucleotide sugar metabolism and opens the perspective for the development of drugs that specifically inhibit simpler organized nucleotidyltransferases in pathogens. PMID:25860585

Führing, Jana Indra; Cramer, Johannes Thomas; Schneider, Julia; Baruch, Petra; Gerardy-Schahn, Rita; Fedorov, Roman

2015-01-01

29

Effect of nitrogen and phosphorus deficiency on transcriptional regulation of genes encoding key enzymes of starch metabolism in duckweed (Landoltia punctata).  

PubMed

The production of starch by plants influences their use as biofuels. Nitrogen (N) and phosphorus (P) regulate starch gene expression during plant growth and development, yet the role of key enzymes such as ADP-glucose pyrophosphorylase (E.C. 2.7.7.27 AGPase) in starch metabolism during N- and P-deficiency remains unknown. We investigated the effect of N- and P-deficiency on the expression of large (LeAPL1, LeAPL2, and LeAPL3) and small (LeAPS) subunits of AGPase in duckweed (Landoltia punctata) and their correlation with starch content. We first isolated the full-length cDNA encoding LeAPL1 (GenBank Accession No. KJ603244) and LeAPS (GenBank Accession No. KJ603243); they contained open reading frames of 1554 bp (57.7-kDa polypeptide of 517 amino acids) and 1578 bp (57.0 kDa polypeptide of 525 amino acids), respectively. Real-time PCR analysis revealed that LeAPL1 and LeAPL3 were highly expressed during early stages of N-deficiency, while LeAPL2 was only expressed during late stage. However, in response to P-deficiency, LeAPL1 and LeAPL2 were upregulated during early stages and LeAPL3 was primarily expressed in the late stage. Interestingly, LeAPS was highly expressed following N-deficiency during both stages, but was only upregulated in the early stage after P-deficiency. The activities of AGPase and soluble starch synthesis enzyme (SSS EC 2.4.1.21) were positively correlated with changes in starch content. Furthermore, LeAPL3 and LeSSS (SSS gene) were positively correlated with changes in starch content during N-deficiency, while LeAPS and LeSSS were correlated with starch content in response to P-deficiency. These results elevate current knowledge of the molecular mechanisms underlying starch synthesis. PMID:25438139

Zhao, Zhao; Shi, Hui-juan; Wang, Mao-lin; Cui, Long; Zhao, Hai; Zhao, Yun

2015-01-01

30

21 CFR 862.3360 - Drug metabolizing enzyme genotyping system.  

Code of Federal Regulations, 2011 CFR

...2011-04-01 false Drug metabolizing enzyme genotyping system. 862.3360 Section...Systems § 862.3360 Drug metabolizing enzyme genotyping system. (a) Identification . A drug metabolizing enzyme genotyping system is a device...

2011-04-01

31

21 CFR 862.3360 - Drug metabolizing enzyme genotyping system.  

Code of Federal Regulations, 2014 CFR

...2014-04-01 false Drug metabolizing enzyme genotyping system. 862.3360 Section...Systems § 862.3360 Drug metabolizing enzyme genotyping system. (a) Identification. A drug metabolizing enzyme genotyping system is a device...

2014-04-01

32

21 CFR 862.3360 - Drug metabolizing enzyme genotyping system.  

Code of Federal Regulations, 2013 CFR

...2013-04-01 false Drug metabolizing enzyme genotyping system. 862.3360 Section...Systems § 862.3360 Drug metabolizing enzyme genotyping system. (a) Identification . A drug metabolizing enzyme genotyping system is a device...

2013-04-01

33

Enzyme and Microbial Technology 35 (2004) 654662 In silico analysis of lactate producing metabolic  

E-print Network

Enzyme and Microbial Technology 35 (2004) 654­662 In silico analysis of lactate producing metabolic observed the effect of enzymes on lactate production using flux distribution analysis, metabolic control results because of their characteristics but some key enzymes for lactate production were found from them

34

Arthrobacter P 1, a fast growing versatile methylotroph with amine oxidase as a key enzyme in the metabolism of methylated amines  

Microsoft Academic Search

A facultative methylotrophic bacterium was isolated from enrichment cultures containing methylamine as the sole carbon source.\\u000a It was tentatively identified as an Arthrobacter species. Extracts of cells grown on methylamine or ethylamine contained high levels of amine oxidase (E.C. 1.4.3.) activity.\\u000a Glucose- or choline-grown cells lacked this enzyme. Oxidation of primary amines by the enzyme resulted in the formation of

P. R. Levering; J. P. van Dijken; M. Veenhuis; W. Harder

1981-01-01

35

Enzymes of glucose metabolism in Frankia sp.  

PubMed

Enzymes of glucose metabolism were assayed in crude cell extracts of Frankia strains HFPArI3 and HFPCcI2 as well as in isolated vesicle clusters from Alnus rubra root nodules. Activities of the Embden-Meyerhof-Parnas pathway enzymes glucokinase, phosphofructokinase, and pyruvate kinase were found in Frankia strain HFPArI3 and glucokinase and pyruvate kinase were found in Frankia strain HFPCcI2 and in the vesicle clusters. An NADP+-linked glucose 6-phosphate dehydrogenase and an NAD-linked 6-phosphogluconate dehydrogenase were found in all of the extracts, although the role of these enzymes is unclear. No NADP+-linked 6-phosphogluconate dehydrogenase was found. Both dehydrogenases were inhibited by adenosine 5-triphosphate, and the apparent Km's for glucose 6-phosphate and 6-phosphogluconate were 6.86 X 10(-4) and 7.0 X 10(-5) M, respectively. In addition to the enzymes mentioned above, an NADP+-linked malic enzyme was detected in the pure cultures but not in the vesicle clusters. In contrast, however, the vesicle clusters had activity of an NAD-linked malic enzyme. The possibility that this enzyme resulted from contamination from plant mitochondria trapped in the vesicle clusters could not be discounted. None of the extracts showed activities of the Entner-Doudoroff enzymes or the gluconate metabolism enzymes gluconate dehydrogenase or gluconokinase. Propionate- versus trehalose-grown cultures of strain HFPArI3 showed similar activities of most enzymes except malic enzyme, which was higher in the cultures grown on the organic acid. Nitrogen-fixing cultures of strain HFPArI3 showed higher specific activities of glucose 6-phosphate and 6-phosphogluconate dehydrogenases and phosphofructokinase than ammonia-grown cultures. PMID:3980434

Lopez, M F; Torrey, J G

1985-04-01

36

Enzyme kinetics in drug metabolism: fundamentals and applications.  

PubMed

Enzymes are protein catalysts that lower the energy barrier for a reaction and speed the rate of a chemical change. The kinetics of reactions catalyzed by enzymes, as well as several mechanisms underlying the kinetics, have been comprehensively studied and written in textbooks (1, 2). The importance of quantitative evaluation of enzymatic processes has been recognized in many fields of study, including biochemistry, molecular biology, and pharmaceutical sciences to name a few. In pharmaceutical sciences, the applications of enzyme kinetics range from hit finding efforts for new chemical entities on a pharmacological target to concentration effect relationships to large-scale biosynthesis. The study of the science of drug metabolism has two principal concepts-rate and extent. While understanding disposition pathways and identification of metabolites provides an insight into the extent of metabolism, kinetics of depletion of substrates (endogenous or exogenous) and formation of metabolites deals with the rate of metabolism. The current textbook specifically focuses on kinetics of drug-metabolizing enzymes, detailing specific enzyme classes, and discusses kinetics as they apply to drug transporters. This textbook also outlines additional factors that contribute to the kinetics of reactions catalyzed by these proteins such as variability in isoforms (pharmacogenomics) and experimental factors including key concepts such as alterations of substrate concentrations due to binding. Applications of these approaches in predicting kinetic parameters and alternative approaches for enzymes (systems biology) and transporters are also discussed. The final section focuses on real-life examples (case studies) to try and exemplify the applications of enzyme kinetic principles. This chapter provides a brief overview outlining some key concepts within each of the sections and the chapters within this textbook. PMID:24523105

Nagar, Swati; Argikar, Upendra A; Tweedie, Donald J

2014-01-01

37

Metaproteogenomic analysis of a sulfate-reducing enrichment culture reveals genomic organization of key enzymes in the m-xylene degradation pathway and metabolic activity of proteobacteria.  

PubMed

This study aimed to ascertain the functional and phylogenetic relationships within an m-xylene degrading sulfate-reducing enrichment culture, which had been maintained for several years in the laboratory with m-xylene as the sole source of carbon and energy. Previous studies indicated that a phylotype affiliated to the Desulfobacteraceae was the main m-xylene assimilating organism. In the present study, genes and gene products were identified by a metaproteogenomic approach using LC-MS/MS analysis of the microbial community, and 2426 peptides were identified from 576 proteins. In the metagenome of the community, gene clusters encoding enzymes involved in fumarate addition to a methyl moiety of m-xylene (nms, bss), as well as gene clusters coding for enzymes involved in modified beta-oxidation to (3-methyl)benzoyl-CoA (bns), were identified in two separate contigs. Additionally, gene clusters containing homologues to bam genes encoding benzoyl-CoA reductase (Bcr) class II, catalyzing the dearomatization of (3-methyl)benzoyl-CoA, were identified. Time-resolved protein stable isotope probing (protein-SIP) experiments using (13)C-labeled m-xylene showed that the respective gene products were highly (13)C-labeled. The present data suggested the identification of gene products that were similar to those involved in methylnaphthalene degradation even though the consortium was not capable of growing in the presence of naphthalene, methylnaphthalene or toluene as substrates. Thus, a novel branch of enzymes was found that was probably specific for anaerobic m-xylene degradation. PMID:25156802

Bozinovski, Dragana; Taubert, Martin; Kleinsteuber, Sabine; Richnow, Hans-Hermann; von Bergen, Martin; Vogt, Carsten; Seifert, Jana

2014-10-01

38

Isolation and structural characterization of 2R, 3R taxifolin 3-O-rhamnoside from ethyl acetate extract of Hydnocarpus alpina and its hypoglycemic effect by attenuating hepatic key enzymes of glucose metabolism in streptozotocin-induced diabetic rats.  

PubMed

Hydnocarpus alpina Wt. (Flacourtiaceae) (H. alpina) is a large tree traditionally used to treat leprosy; it also posses antidiabetic property. The present study was undertaken to isolate, characterize and to evaluate the antidiabetic effect of 2R, 3R taxifolin 3-O-rhamnoside. (rhamnoside) and its impact on carbohydrate metabolic key enzymes in control and streptozotocin (STZ)-induced diabetic rats. Diabetes mellitus was induced by a single intraperitoneal injection of streptozotocin (STZ) (40 mg/kg). Oral administration of rhamnoside for 21 days significantly reduced food intake, calorie intake, blood glucose and glycosylated hemoglobin levels, and improved plasma insulin levels. Administration of rhamnoside showed significant increase in the body weight, body composition (Lean body weight (LBW) and retro body fat), glycolytic hexokinase, glucose-6-phophate dehydrogenase and pyruvate kinase levels where as significant decrease was observed in the levels of glucose-6-phosphatase fructose-1, 6-bisphosphatase and lactate dehydrogenase in diabetic treated rats. Further, administration of rhamnoside significantly improved the glycogen content, glycogen synthase and glycogen phosphorylase, suggesting the antihyperglycemic potential of rhamnoside in diabetic rats. The results obtained were compared with glibenclamide a standard hypoglycaemic drug. Immunohistopathological study of pancreas revealed increased number of ?-cells and insulin granules in diabetes-induced rats after treatment with rhamnoside for 21 days. Furthermore, Co-administration of rhamnoside (50 mg/kg) with nifedipine (13.6 mg/kg), a Ca(2+)ion channel blocker, or nicorandil (6.8 mg/kg), an ATP-sensitive K(+) ion channel opener, reveals the insulin secretion property of rhamnoside via a K(+)-ATP channels dependent pathway in diabetic rats. In conclusion, rhamnoside normalized blood glucose, glycosylated hemoglobin, key hepatic enzymes and glycogen content by increasing insulin secretion via K(+)-ATP channels dependent signaling pathway. The results suggest that the rhamnoside from H. alpina could be used as a therapeutic agent to treat diabetes mellitus. PMID:25698613

Balamurugan, Rangachari; Vendan, Subramanian Ezhil; Aravinthan, Adithan; Kim, Jong-Hoon

2015-04-01

39

Expression of Enzymes that Metabolize Medications  

NASA Technical Reports Server (NTRS)

Most pharmaceuticals are metabolized by the liver. Clinically-used medication doses are given with normal liver function in mind. A drug overdose can result if the liver is damaged and removing pharmaceuticals from the circulation at a rate slower than normal. Alternatively, if liver function is elevated and removing drugs from the system more quickly than usual, it would be as if too little drug had been given for effective treatment. Because of the importance of the liver in drug metabolism we want to understand the effects of spaceflight on the enzymes of the liver.

Wotring, Virginia E.; Peters, C. P.

2012-01-01

40

Modulating effects of hesperidin on key carbohydrate-metabolizing enzymes, lipid profile, and membrane-bound adenosine triphosphatases against 7,12-dimethylbenz(a)anthracene-induced breast carcinogenesis.  

PubMed

The aim of this study was to document the effect of hesperidin on the key enzyme activities of carbohydrate metabolism, lipid profile, and membrane-bound adenosine triphosphatases (ATPases) during 7,12-dimethylbenz(a)anthracene (DMBA)-induced breast carcinogenesis. Hesperidin has been reported to have multiple biological properties. Breast cancer was induced by single dose of DMBA (20 mg/kg body weight (bw)). The results revealed that there was a significant increase in the activities of hexokinase, phosphoglucoisomerase, and aldolase and a concomitant decrease in the activities of glucose-6-phosphatase and fructose-1,6-diphosphatase in cancer-induced animals. The activities of ATPases were found to be decreased both in erythrocyte membrane and in the liver of mammary cancer-bearing animals. The lipid profiles such as total cholesterol, free cholesterol, phospholipids, triglycerides, and free fatty acids significantly increased and in contrast the ester cholesterol in plasma was found to be decreased, whereas it was found to be elevated in the liver of cancer-bearing groups. The altered levels of the above-mentioned biochemical parameters in cancer-bearing animals were significantly ameliorated by the administration of hesperidin at the dosage of 30 mg/kg bw for 45 days. The histopathological analysis of breast and liver tissues were well supported the modulatory property of hesperidin, and this might be associated with normalizing the gluconeogenesis process, stabilization of cell membranes, and modulation of lipid biosynthesis. PMID:23690228

Nandakumar, N; Rengarajan, T; Balamurugan, A; Balasubramanian, M P

2014-05-01

41

Metabolic profiling reveals key metabolic features of renal cell carcinoma  

PubMed Central

Abstract Recent evidence suggests that metabolic changes play a pivotal role in the biology of cancer and in particular renal cell carcinoma (RCC). Here, a global metabolite profiling approach was applied to characterize the metabolite pool of RCC and normal renal tissue. Advanced decision tree models were applied to characterize the metabolic signature of RCC and to explore features of metastasized tumours. The findings were validated in a second independent dataset. Vitamin E derivates and metabolites of glucose, fatty acid, and inositol phosphate metabolism determined the metabolic profile of RCC. ?-tocopherol, hippuric acid, myoinositol, fructose-1-phosphate and glucose-1-phosphate contributed most to the tumour/normal discrimination and all showed pronounced concentration changes in RCC. The identified metabolic profile was characterized by a low recognition error of only 5% for tumour versus normal samples. Data on metastasized tumours suggested a key role for metabolic pathways involving arachidonic acid, free fatty acids, proline, uracil and the tricarboxylic acid cycle. These results illustrate the potential of mass spectroscopy based metabolomics in conjunction with sophisticated data analysis methods to uncover the metabolic phenotype of cancer. Differentially regulated metabolites, such as vitamin E compounds, hippuric acid and myoinositol, provide leads for the characterization of novel pathways in RCC. PMID:19845817

Catchpole, Gareth; Platzer, Alexander; Weikert, Cornelia; Kempkensteffen, Carsten; Johannsen, Manfred; Krause, Hans; Jung, Klaus; Miller, Kurt; Willmitzer, Lothar; Selbig, Joachim; Weikert, Steffen

2011-01-01

42

Tracing metabolic pathways from enzyme data.  

PubMed

The IUBMB Enzyme List is widely used by other databases as a source for avoiding ambiguity in the recognition of enzymes as catalytic entities. However, it was never designed for activities such as pathway tracing, which have become increasingly important in systems biology. This is because it often relies on generic or representative reactions to show the reactions catalysed by enzymes of wide specificity. It is necessary to go to databases such as BRENDA to find further, more detailed, information on what is known about the range of substrates for any particular enzyme. In order to provide a framework for tracing pathways involving any specific enzyme or metabolite, we have created a Reactions Database from the material in the Enzyme List. This allows reactions to be searched by substrate/product and pathways to be traced from any selected starting/seed substrate. An extensive synonym glossary allows searches by many of the alternative names, including accepted abbreviations, by which a chemical compound may be known. This database was necessary for the development of the application Reaction Explorer (http://www.reaction-explorer.org), which was written in REALbasic to search the Reactions Database and draw metabolic pathways from reactions selected by the user. Having input the name of the starting compound (the "seed"), the user is presented with a list of all reactions containing that compound and then selects the product of interest as the next point on the ensuing graph. The pathway diagram is then generated as the process iterates. A contextual menu is provided, which allows the user to (i) remove a compound from the graph, along with all associated links; (ii) search the reactions database again for additional reactions involving the compound and (iii) search for the compound within the Enzyme List. PMID:19563919

McDonald, Andrew G; Tipton, Keith F; Boyce, Sinéad

2009-09-01

43

A core metabolic enzyme mediates resistance to phosphine gas.  

PubMed

Phosphine is a small redox-active gas that is used to protect global grain reserves, which are threatened by the emergence of phosphine resistance in pest insects. We find that polymorphisms responsible for genetic resistance cluster around the redox-active catalytic disulfide or the dimerization interface of dihydrolipoamide dehydrogenase (DLD) in insects (Rhyzopertha dominica and Tribolium castaneum) and nematodes (Caenorhabditis elegans). DLD is a core metabolic enzyme representing a new class of resistance factor for a redox-active metabolic toxin. It participates in four key steps of core metabolism, and metabolite profiles indicate that phosphine exposure in mutant and wild-type animals affects these steps differently. Mutation of DLD in C. elegans increases arsenite sensitivity. This specific vulnerability may be exploited to control phosphine-resistant insects and safeguard food security. PMID:23139334

Schlipalius, David I; Valmas, Nicholas; Tuck, Andrew G; Jagadeesan, Rajeswaran; Ma, Li; Kaur, Ramandeep; Goldinger, Anita; Anderson, Cameron; Kuang, Jujiao; Zuryn, Steven; Mau, Yosep S; Cheng, Qiang; Collins, Patrick J; Nayak, Manoj K; Schirra, Horst Joachim; Hilliard, Massimo A; Ebert, Paul R

2012-11-01

44

The enzymes of human diphosphoinositol polyphosphate metabolism  

PubMed Central

Diphospho-myo-inositol polyphosphates have many roles to play, including roles in apoptosis, vesicle trafficking, the response of cells to stress, the regulation of telomere length and DNA damage repair, and inhibition of the cyclin-dependent kinase Pho85 system that monitors phosphate levels. This review focuses on the three classes of enzymes involved in the metabolism of these compounds: inositol hexakisphosphate kinases, inositol hexakisphosphate and diphosphoinositol-pentakisphosphate kinases and diphosphoinositol polyphosphate phosphohydrolases. However, these enzymes have roles beyond being mere catalysts, and their interactions with other proteins have cellular consequences. Through their interactions, the three inositol hexakisphosphate kinases have roles in exocytosis, diabetes, the response to infection, and apoptosis. The two inositol hexakisphosphate and diphosphoinositol-pentakisphosphate kinases influence the cellular response to phosphatidylinositol (3,4,5)-trisphosphate and the migration of pleckstrin homology domain-containing proteins to the plasma membrane. The five diphosphoinositol polyphosphate phosphohydrolases interact with ribosomal proteins and transcription factors, as well as proteins involved in membrane trafficking, exocytosis, ubiquitination and the proteasomal degradation of target proteins. Possible directions for future research aiming to determine the roles of these enzymes are highlighted. PMID:24152294

Thomas, Mark P; Potter, Barry V L

2014-01-01

45

Expression of Enzymes that Metabolize Medications  

NASA Technical Reports Server (NTRS)

INTRODUCTION: Increased exposure to radiation is one physiological stressor associated with spaceflight and it is feasible to conduct ground experiments using known radiation exposures. The health of the liver, especially the activity rate of its metabolic enzymes, determines the concentration of circulating drugs as well as the duration of their efficacy. While radiation is known to alter normal physiological function, how radiation affects liver metabolism of administered medications is unclear. Crew health could be affected if the actions of medications used in spaceflight deviated from expectations formed during terrestrial medication use. This study is an effort to identify liver metabolic enzymes whose expression is altered by spaceflight or by radiation exposures that mimic features of the spaceflight environment. METHODS: Using procedures approved by the Animal Care and Use Committee, mice were exposed to either 137Cs (controls, 50 mGy, 6Gy, or 50 mGy + 6Gy separated by 24 hours) or 13 days of spaceflight on STS 135. Animals were anesthetized and sacrificed at several time points (4 hours, 24 hours or 7 days) after their last radiation exposure, or within 6 hours of return to Earth for the STS 135 animals. Livers were removed immediately and flash-frozen in liquid nitrogen. Tissue was homogenized, RNA extracted, purified and quality-tested. Complementary DNA was prepared from high-quality RNA samples, and used in RT-qPCR experiments to determine relative expression of a wide variety of genes involved in general metabolism and drug metabolism. RESULTS: Results of the ground radiation exposure experiments indicated 65 genes of the 190 tested were significantly affected by at least one of the radiation doses. Many of the affected genes are involved in the metabolism of drugs with hydrophobic or steroid-like structures, maintenance of redox homeostasis and repair of DNA damage. Most affected genes returned to near control expression levels by 7 days post-treatment. Not all recovered completely by the final time point tested: with 6 Gy exposure, metallothionein expression was 132-fold more than control at the 4 hr time point, and fell at each later time point (11-fold at 24 hrs, and 8-fold at 7 days). In contrast, there were other genes whose expression was altered and remained relatively constant through the 7 day period we tested. One examples is Cyp17a1, which showed a 4-fold elevation at 4 hrs after exposure and remained constant for 7 days after the last treatment. Spaceflight samples evaluated with similar methods and comparisons will be made between the radiation-treated groups and the spaceflight samples. CONCLUSION It seems likely that radiation exposure triggers homeostatic mechanisms, which could include alterations of gene expression. Better understanding of these pathways could aid in optimizing medications doses given to crewmembers who require treatment and eventually, to development of new countermeasures to ameliorate or prevent radiation-induced damage to cells and tissues.

Wotring, V. E.; Peters, C. P.

2011-01-01

46

Tradeoff between enzyme and metabolite efficiency maintains metabolic homeostasis upon perturbations in enzyme capacity  

PubMed Central

What is the relationship between enzymes and metabolites, the two major constituents of metabolic networks? We propose three alternative relationships between enzyme capacity and metabolite concentration alterations based on a Michaelis–Menten kinetic; that is enzyme capacities, metabolite concentrations, or both could limit the metabolic reaction rates. These relationships imply different correlations between changes in enzyme capacity and metabolite concentration, which we tested by quantifying metabolite, transcript, and enzyme abundances upon local (single-enzyme modulation) and global (GCR2 transcription factor mutant) perturbations in Saccharomyces cerevisiae. Our results reveal an inverse relationship between fold-changes in substrate metabolites and their catalyzing enzymes. These data provide evidence for the hypothesis that reaction rates are jointly limited by enzyme capacity and metabolite concentration. Hence, alteration in one network constituent can be efficiently buffered by converse alterations in the other constituent, implying a passive mechanism to maintain metabolic homeostasis upon perturbations in enzyme capacity. PMID:20393576

Fendt, Sarah-Maria; Buescher, Joerg Martin; Rudroff, Florian; Picotti, Paola; Zamboni, Nicola; Sauer, Uwe

2010-01-01

47

Metabolic Syndrome -- Lifestyle Changes are Key  

MedlinePLUS

By Jill Pluhar R.D., L.D.N. Brigham and Women's Hospital What Is Metabolic Syndrome? What Puts Me at Risk for Developing Metabolic ... of metabolic syndrome. Back to top Jill Pluhar R.D., L.D.N. is a registered dietitian working ...

48

21 CFR 862.3360 - Drug metabolizing enzyme genotyping system.  

Code of Federal Regulations, 2010 CFR

...Identification . A drug metabolizing enzyme genotyping system is a device intended for use in testing deoxyribonucleic acid (DNA) extracted from clinical samples to identify the presence or absence of human genotypic markers encoding a drug...

2010-04-01

49

21 CFR 862.3360 - Drug metabolizing enzyme genotyping system.  

Code of Federal Regulations, 2012 CFR

...Identification . A drug metabolizing enzyme genotyping system is a device intended for use in testing deoxyribonucleic acid (DNA) extracted from clinical samples to identify the presence or absence of human genotypic markers encoding a drug...

2012-04-01

50

Key Applications of Plant Metabolic Engineering  

PubMed Central

Great strides have been made in plant metabolic engineering over the last two decades, with notable success stories including Golden rice. Here, we discuss the field's progress in addressing four long-standing challenges: creating plants that satisfy their own nitrogen requirement, so reducing or eliminating the need for nitrogen fertilizer; enhancing the nutrient content of crop plants; engineering biofuel feed stocks that harbor easy-to-access fermentable saccharides by incorporating self-destructing lignin; and increasing photosynthetic efficiency. We also look to the future at emerging areas of research in this field. PMID:24915445

Lau, Warren; Fischbach, Michael A.; Osbourn, Anne; Sattely, Elizabeth S.

2014-01-01

51

Key applications of plant metabolic engineering.  

PubMed

Great strides have been made in plant metabolic engineering over the last two decades, with notable success stories including Golden rice. Here, we discuss the field's progress in addressing four long-standing challenges: creating plants that satisfy their own nitrogen requirement, so reducing or eliminating the need for nitrogen fertilizer; enhancing the nutrient content of crop plants; engineering biofuel feed stocks that harbor easy-to-access fermentable saccharides by incorporating self-destructing lignin; and increasing photosynthetic efficiency. We also look to the future at emerging areas of research in this field. PMID:24915445

Lau, Warren; Fischbach, Michael A; Osbourn, Anne; Sattely, Elizabeth S

2014-06-01

52

Enzyme heterozygosity, metabolism, and developmental stability  

Microsoft Academic Search

Developmental homeostasis, measured as either fluctuating asymmetry or variance of morphological characters, increases with\\u000a enzyme heterozygosity in many, but not all, natural populations. These results have been reported forDrosophila, monarch butterflies, honeybees, blue mussels, side-blotched lizards, killifish, salmonid fishes, guppies, Sonoran topminnows,\\u000a herring, rufous-collared sparrows, house sparrows, brown hares, white-tailed deer, and humans. Because heterozygosity at a\\u000a few loci can

Jeffry B. Mitton

1993-01-01

53

Experiment K-7-21: Effect of Microgravity on 1: Metabolic Enzymes of Type 1 and Type 2 Muscle Fibers, and on 2: Metabolic Enzymes, Neurotransmitter Amino Acids, and Neurotransmitter Associated Enzymes in Selected Regions of the Central Nervous System. Part 2; The Distribution of Selected Enzymes and Amino Acids in the Hippocampal Formation  

NASA Technical Reports Server (NTRS)

Six key metabolic enzymes plus glutaminase and glutamate decarboxylase, as well as glutamate, aspartate and GABA, were measured in 11 regions of the hippocampal formation of synchronous, flight and tail suspension rats. Major differences were observed in the normal distribution patterns of each enzyme and amino acid, but no substantive effects of either microgravity or tail suspension on these patterns were clearly demonstrated.

Lowry, O. H.; Krasnov, I.; Ilyina-Kakueva, E. I.; Nemeth, P. M.; McDougal, D. B., Jr.; Choksi, R.; Carter, J. G.; Chi, M. M. Y.; Manchester, J. K.; Pusateri, M. E.

1994-01-01

54

Bifidobacterial Enzymes Involved in the Metabolism of Human Milk Oligosaccharides123  

PubMed Central

Intestinal colonization of bifidobacteria is important for the health of infants. Human milk oligosaccharides (HMO) have been identified as growth factors for bifidobacteria. Recently, a bifidobacterial enzymatic system to metabolize HMO was identified. 1,3-?-Galactosyl-N-acetylhexosamine phosphorylase (GLNBP, EC 2.4.1.211), which catalyzes the reversible phosphorolysis of galacto-N-biose (GNB) (Gal?1?3GalNAc)] and lacto-N-biose I (LNB) (Gal?1?3GlcNAc), is a key enzyme to explain the metabolism of HMO. Infant-type bifidobacteria possess the intracellular pathway to specifically metabolize GNB and LNB (GNB/LNB pathway). Bifidobacterium bifidum possesses extracellular enzymes to liberate LNB from HMO. However, Bifidobacterium longum subsp. infantis imports intact HMO to be hydrolyzed by intracellular enzymes. Bifidobacterial enzymes related to the metabolism of HMO are useful tools for preparing compounds related to HMO. For instance, LNB and GNB were produced from sucrose and GlcNAc/GalNAc in 1 pot using 4 bifidobacterial enzymes, including GLNBP. LNB is expected to be a selective bifidus factor for infant-type strains. PMID:22585921

Kitaoka, Motomitsu

2012-01-01

55

Regulation of Gene Expression by a Metabolic Enzyme  

E-print Network

enzymatic pro- teins can directly control gene expression has not been extensively investigated. Butow, bound specific se- quences, or had a direct role in affecting the expression of specific genes. Zheng etRegulation of Gene Expression by a Metabolic Enzyme David A. Hall,1 Heng Zhu,2 * Xiaowei Zhu,3

Gerstein, Mark

56

2006 Nature Publishing Group A trehalose metabolic enzyme controls  

E-print Network

© 2006 Nature Publishing Group A trehalose metabolic enzyme controls inflorescence architecture Inflorescence branching is a major yield trait in crop plants controlled by the developmental fate of axillary (inflorescences) and affect crop yield by influencing seed number or harvesting ability2,3 . Several growth

Jackson, David

57

How nutritional status signalling coordinates metabolism and lignocellulolytic enzyme secretion.  

PubMed

The utilisation of lignocellulosic plant biomass as an abundant, renewable feedstock for green chemistries and biofuel production is inhibited by its recalcitrant nature. In the environment, lignocellulolytic fungi are naturally capable of breaking down plant biomass into utilisable saccharides. Nonetheless, within the industrial context, inefficiencies in the production of lignocellulolytic enzymes impede the implementation of green technologies. One of the primary causes of such inefficiencies is the tight transcriptional control of lignocellulolytic enzymes via carbon catabolite repression. Fungi coordinate metabolism, protein biosynthesis and secretion with cellular energetic status through the detection of intra- and extra-cellular nutritional signals. An enhanced understanding of the signals and signalling pathways involved in regulating the transcription, translation and secretion of lignocellulolytic enzymes is therefore of great biotechnological interest. This comparative review describes how nutrient sensing pathways regulate carbon catabolite repression, metabolism and the utilisation of alternative carbon sources in Saccharomyces cerevisiae and ascomycete fungi. PMID:25011009

Brown, Neil Andrew; Ries, Laure Nicolas Annick; Goldman, Gustavo Henrique

2014-11-01

58

Enzyme kinetics and metabolic control. A method to test and quantify the effect of enzymic properties on metabolic variables.  

PubMed Central

It is usual to study the sensitivity of metabolic variables to small (infinitesimal) changes in the magnitudes of individual parameters such as an enzyme concentration. Here, the effect that a simultaneous change in all the enzyme concentrations by the same factor alpha (Co-ordinate-Control Operation, CCO) has on the variables of time-dependent metabolic systems is investigated. This factor alpha can have any arbitrary large value. First, we assume, for each enzyme measured in isolation, the validity of the steady-state approximation and the proportionality between reaction rate and enzyme concentration. Under these assumptions, any time-invariant variable may behave like a metabolite concentration, i.e. S alpha = Sr (S-type), or like a flux, i.e. J alpha = alpha Jr (J-type). The subscripts r and alpha correspond to the values of the variable before and after the CCO respectively. Similarly, time-dependent variables may behave according to S alpha (t/alpha) = Sr (t) (S-type) or to J alpha (t/alpha) = alpha J r (t) (J-type). A method is given to test these relationships in experimental systems, and to quantify deviations from the predicted behaviour. A positive test for deviations proves the violation of some of the assumptions made. However, the breakdown of the assumptions in an enzyme-catalysed reaction, studied in isolation, may or may not affect significantly the behaviour of the system when the component reaction is embedded in the metabolic network. PMID:2390063

Acerenza, L; Kacser, H

1990-01-01

59

Filament formation by metabolic enzymes is a specific adaptation to an advanced state of cellular starvation  

PubMed Central

One of the key questions in biology is how the metabolism of a cell responds to changes in the environment. In budding yeast, starvation causes a drop in intracellular pH, but the functional role of this pH change is not well understood. Here, we show that the enzyme glutamine synthetase (Gln1) forms filaments at low pH and that filament formation leads to enzymatic inactivation. Filament formation by Gln1 is a highly cooperative process, strongly dependent on macromolecular crowding, and involves back-to-back stacking of cylindrical homo-decamers into filaments that associate laterally to form higher order fibrils. Other metabolic enzymes also assemble into filaments at low pH. Hence, we propose that filament formation is a general mechanism to inactivate and store key metabolic enzymes during a state of advanced cellular starvation. These findings have broad implications for understanding the interplay between nutritional stress, the metabolism and the physical organization of a cell. DOI: http://dx.doi.org/10.7554/eLife.02409.001 PMID:24771766

Petrovska, Ivana; Nüske, Elisabeth; Munder, Matthias C; Kulasegaran, Gayathrie; Malinovska, Liliana; Kroschwald, Sonja; Richter, Doris; Fahmy, Karim; Gibson, Kimberley; Verbavatz, Jean-Marc; Alberti, Simon

2014-01-01

60

Inhibitors of testosterone biosynthetic and metabolic activation enzymes.  

PubMed

The Leydig cells of the testis have the capacity to biosynthesize testosterone from cholesterol. Testosterone and its metabolically activated product dihydrotestosterone are critical for the development of male reproductive system and spermatogenesis. At least four steroidogenic enzymes are involved in testosterone biosynthesis: Cholesterol side chain cleavage enzyme (CYP11A1) for the conversion of cholesterol into pregnenolone within the mitochondria, 3?-hydroxysteroid dehydrogenase (HSD3B), for the conversion of pregnenolone into progesterone, 17?-hydroxylase/17,20-lyase (CYP17A1) for the conversion of progesterone into androstenedione and 17?-hydroxysteroid dehydrogenase (HSD17B3) for the formation of testosterone from androstenedione. Testosterone is also metabolically activated into more potent androgen dihydrotestosterone by two isoforms 5?-reductase 1 (SRD5A1) and 2 (SRD5A2) in Leydig cells and peripheral tissues. Many endocrine disruptors act as antiandrogens via directly inhibiting one or more enzymes for testosterone biosynthesis and metabolic activation. These chemicals include industrial materials (perfluoroalkyl compounds, phthalates, bisphenol A and benzophenone) and pesticides/biocides (methoxychlor, organotins, 1,2-dibromo-3-chloropropane and prochloraz) and plant constituents (genistein and gossypol). This paper reviews these endocrine disruptors targeting steroidogenic enzymes. PMID:22138857

Ye, Leping; Su, Zhi-Jian; Ge, Ren-Shan

2011-01-01

61

Radiation Exposure Alters Expression of Metabolic Enzyme Genes in Mice  

NASA Technical Reports Server (NTRS)

Most administered pharmaceuticals are metabolized by the liver. The health of the liver, especially the rate of its metabolic enzymes, determines the concentration of circulating drugs as well as the duration of their efficacy. Most pharmaceuticals are metabolized by the liver, and clinically-used medication doses are given with normal liver function in mind. A drug overdose can result in the case of a liver that is damaged and removing pharmaceuticals from the circulation at a rate slower than normal. Alternatively, if liver function is elevated and removing drugs from the system more quickly than usual, it would be as if too little drug had been given for effective treatment. Because of the importance of the liver in drug metabolism, we want to understand the effects of spaceflight on the enzymes of the liver and exposure to cosmic radiation is one aspect of spaceflight that can be modeled in ground experiments. Additionally, it has been previous noted that pre-exposure to small radiation doses seems to confer protection against later and larger radiation doses. This protective power of pre-exposure has been called a priming effect or radioadaptation. This study is an effort to examine the drug metabolizing effects of radioadaptation mechanisms that may be triggered by early exposure to low radiation doses.

Wotring, V. E.; Mangala, L. S.; Zhang, Y.; Wu, H.

2011-01-01

62

Radiation Exposure Alters Expression of Metabolic Enzyme Genes In Mice  

NASA Technical Reports Server (NTRS)

Most pharmaceuticals are metabolized by the liver. The health of the liver, especially the rate of its metabolic enzymes, determines the concentration of circulating drugs as well as the duration of their efficacy. Because of the importance of the liver in drug metabolism it is important to understand the effects of spaceflight on the enzymes of the liver. Exposure to cosmic radiation is one aspect of spaceflight that can be modeled in ground experiments. This study is an effort to examine the effects of adaptive mechanisms that may be triggered by early exposure to low radiation doses. Using procedures approved by the JSC Animal Care & Use Committee, C57 male mice were exposed to Cs-137 in groups: controls, low dose (50 mGy), high dose (6Gy) and a fourth group that received both radiation doses separated by 24 hours. Animals were anesthetized and sacrificed 4 hours after their last radiation exposure. Livers were removed immediately and flash-frozen in liquid nitrogen. Tissue was homogenized, RNA extracted and purified (Absolutely RNA, Agilent). Quality of RNA samples was evaluated (Agilent Bioanalyzer 2100). Complementary DNA was prepared from high-quality RNA samples, and used to run RT-qPCR screening arrays for DNA Repair and Drug Metabolism (SuperArray, SABiosciences/Qiagen; BioRad Cfx96 qPCR System). Of 91 drug metabolism genes examined, expression of 7 was altered by at least one treatment condition. Genes that had elevated expression include those that metabolize promethazine and steroids (4-8-fold), many that reduce oxidation products, and one that reduces heavy metal exposure (greater than 200-fold). Of the 91 DNA repair and general metabolism genes examined, expression of 14 was altered by at least one treatment condition. These gene expression changes are likely homeostatic and could lead to development of new radioprotective countermeasures.

Wotring, Virginia E.; Mangala, L. S.; Zhang, Y.; Wu, H.

2010-01-01

63

Clinically Relevant Genetic Variations in Drug Metabolizing Enzymes  

PubMed Central

In the field of pharmacogenetics, we currently have a few markers to guide physicians as to the best course of therapy for patients. For the most part, these genetic variants are within a drug metabolizing enzyme that has a large effect on the degree or rate at which a drug is converted to its metabolites. For many drugs, response and toxicity are multi-genic traits and understanding relationships between a patient's genetic variation in drug metabolizing enzymes and the efficacy and/or toxicity of a medication offers the potential to optimize therapies. This review will focus on variants in drug metabolizing enzymes with predictable and relatively large impacts on drug efficacy and/or toxicity; some of these drug/gene variant pairs have impacted drug labels by the United States Food and Drug Administration. The challenges in identifying genetic markers and implementing clinical changes based on known markers will be discussed. In addition, the impact of next generation sequencing in identifying rare variants will be addressed. PMID:21453273

Pinto, Navin; Dolan, M. Eileen

2011-01-01

64

Regulation of liver metabolism by enzyme phosphorylation during mammalian hibernation.  

PubMed

Kinetic properties of regulatory enzymes of glycolysis in liver of the mouse, Zapus hudsonius, were modified during hibernation, the probable mechanism being covalent modification. Liver glycogen phosphorylase activity was strongly depressed during both short (less than 24 h) and long (5-8 days) term hibernation, the mechanism involving a decrease in both the percentage of enzyme in the active a form and the total amount (a + b) of enzyme expressed. Phosphofructokinase showed kinetic changes (a 2.5-fold increase in Ka for fructose-2,6-P2, 4- and 3.7-fold decreases in I50 values for ATP and citrate, compared to euthermic controls) in liver of hibernators indicative of phosphorylation inactivation of the enzyme. Measured levels of fructose-2,6-P2 in liver did not change during hibernation. Changes in pyruvate kinase kinetics in liver from long term hibernators similarly indicated enzyme phosphorylation in the depressed state (Ka for fructose-1,6-P2 increased 4.4-fold, I50 for L-alanine decreased 6.3-fold). Apparent covalent modification of glycolytic enzymes during hibernation may serve two functions: depression of glycolytic activity as part of the general metabolic rate depression of hibernation, or reorganization of fuel use in the hibernating state to limit carbohydrate catabolism and promote gluconeogenesis. PMID:2948958

Storey, K B

1987-02-01

65

Elucidation of metabolic pathways from enzyme classification data.  

PubMed

The IUBMB Enzyme List is widely used by other databases as a source for avoiding ambiguity in the recognition of enzymes as catalytic entities. However, it was not designed for metabolic pathway tracing, which has become increasingly important in systems biology. A Reactions Database has been created from the material in the Enzyme List to allow reactions to be searched by substrate/product, and pathways to be traced from any selected starting/seed substrate. An extensive synonym glossary allows searches by many of the alternative names, including accepted abbreviations, by which a chemical compound may be known. This database was necessary for the development of the application Reaction Explorer ( http://www.reaction-explorer.org ), which was written in Real Studio ( http://www.realsoftware.com/realstudio/ ) to search the Reactions Database and draw metabolic pathways from reactions selected by the user. Having input the name of the starting compound (the "seed"), the user is presented with a list of all reactions containing that compound and then selects the product of interest as the next point on the ensuing graph. The pathway diagram is then generated as the process iterates. A contextual menu is provided, which allows the user: (1) to remove a compound from the graph, along with all associated links; (2) to search the reactions database again for additional reactions involving the compound; (3) to search for the compound within the Enzyme List. PMID:24218216

McDonald, Andrew G; Tipton, Keith F

2014-01-01

66

Gene expression analysis of membrane transporters and drug-metabolizing enzymes in the lung of healthy and COPD subjects  

PubMed Central

This study describes for the first time the expression levels of genes encoding membrane transporters and drug-metabolizing enzymes in the lungs of ex-smoking patients with chronic obstructive pulmonary disease (COPD). Membrane transporters and drug-metabolizing enzymes are key determinants of drug uptake, metabolism, and elimination for systemically administered as well as inhaled drugs, with consequent influence on clinical efficacy and patient safety. In this study, while no difference in gene expression was found between healthy and COPD subjects, we identified a significant regional difference in mRNA expression of both membrane transporters and drug-metabolizing enzymes between central and peripheral tissue in both healthy and COPD subjects. The majority of the differentially expressed genes were higher expressed in the central airways such as the transporters SLC2A1 (GLUT1), SLC28A3 (CNT3), and SLC22A4 (OCTN1) and the drug-metabolizing enzymes GSTZ1, GSTO2, and CYP2F1. Together, this increased knowledge of local pharmacokinetics in diseased and normal lung may improve modeling of clinical outcomes of new chemical entities intended for inhalation therapy delivered to COPD patients. In addition, based on the similarities between COPD and healthy subjects regarding gene expression of membrane transporters and drug-metabolizing enzymes, our results suggest that clinical pharmacological studies in healthy volunteers could be a valid model of COPD patients regarding drug disposition of inhaled drugs in terms of drug metabolism and drug transporters. PMID:25505599

Berg, Tove; Hegelund Myrbäck, Tove; Olsson, Marita; Seidegĺrd, Janeric; Werkström, Viktoria; Zhou, Xiao-Hong; Grunewald, Johan; Gustavsson, Lena; Nord, Magnus

2014-01-01

67

Metabolic modeling of muscle metabolism identifies key reactions linked to insulin resistance phenotypes  

PubMed Central

Objective Dysregulated muscle metabolism is a cardinal feature of human insulin resistance (IR) and associated diseases, including type 2 diabetes (T2D). However, specific reactions contributing to abnormal energetics and metabolic inflexibility in IR are unknown. Methods We utilize flux balance computational modeling to develop the first systems-level analysis of IR metabolism in fasted and fed states, and varying nutrient conditions. We systematically perturb the metabolic network to identify reactions that reproduce key features of IR-linked metabolism. Results While reduced glucose uptake is a major hallmark of IR, model-based reductions in either extracellular glucose availability or uptake do not alter metabolic flexibility, and thus are not sufficient to fully recapitulate IR-linked metabolism. Moreover, experimentally-reduced flux through single reactions does not reproduce key features of IR-linked metabolism. However, dual knockdowns of pyruvate dehydrogenase (PDH), in combination with reduced lipid uptake or lipid/amino acid oxidation (ETFDH), does reduce ATP synthesis, TCA cycle flux, and metabolic flexibility. Experimental validation demonstrates robust impact of dual knockdowns in PDH/ETFDH on cellular energetics and TCA cycle flux in cultured myocytes. Parallel analysis of transcriptomic and metabolomics data in humans with IR and T2D demonstrates downregulation of PDH subunits and upregulation of its inhibitory kinase PDK4, both of which would be predicted to decrease PDH flux, concordant with the model. Conclusions Our results indicate that complex interactions between multiple biochemical reactions contribute to metabolic perturbations observed in human IR, and that the PDH complex plays a key role in these metabolic phenotypes. PMID:25737951

Nogiec, Christopher; Burkart, Alison; Dreyfuss, Jonathan M.; Lerin, Carles; Kasif, Simon; Patti, Mary-Elizabeth

2015-01-01

68

Impact of expression of EMP enzymes on glucose metabolism in Zymomonas mobilis.  

PubMed

Zymomonas mobilis is the only known microorganism that utilizes the Entner-Doudoroff (ED) pathway anaerobically. In this work, we investigated whether the overexpression of a phosphofructokinase (PFK), the only missing Embden-Meyerhof-Parnas (EMP) pathway enzyme, could establish the pathway in this organism. Introduction of a pyrophosphate-dependent PFK, along with co-expression of homologous fructose-1,6-bisphosphate aldolase and triosephosphate isomerase, did not result in an EMP flux to any appreciable level. However, the metabolism of glucose was impacted significantly. Eight percent of glucose was metabolized to form a new metabolite, dihydroxyacetone. Reducing flux through the ED pathway by as much as 40 % through antisense of a key enzyme, ED aldolase, did not result in a fully functional EMP pathway, suggesting that the ED pathway, especially the lower arm, downstream from glyceraldehyde-3-phosphate, is very rigid, possibly due to redox balance. PMID:23613118

Chen, Rachel Ruizhen; Agrawal, Manoj; Mao, Zichao

2013-06-01

69

In vivo enzyme activity in inborn errors of metabolism  

SciTech Connect

Low-dose continuous infusions of (2H5)phenylalanine, (1-13C)propionate, and (1-13C)leucine were used to quantitate phenylalanine hydroxylation in phenylketonuria (PKU, four subjects), propionate oxidation in methylmalonic acidaemia (MMA, four subjects), and propionic acidaemia (PA, four subjects) and leucine oxidation in maple syrup urine disease (MSUD, four subjects). In vivo enzyme activity in PKU, MMA, and PA subjects was similar to or in excess of that in adult controls (range of phenylalanine hydroxylation in PKU, 3.7 to 6.5 mumol/kg/h, control 3.2 to 7.9, n = 7; propionate oxidation in MMA, 15.2 to 64.8 mumol/kg/h, and in PA, 11.1 to 36.0, control 5.1 to 19.0, n = 5). By contrast, in vivo leucine oxidation was undetectable in three of the four MSUD subjects (less than 0.5 mumol/kg/h) and negligible in the remaining subject (2 mumol/kg/h, control 10.4 to 15.7, n = 6). These results suggest that significant substrate removal can be achieved in some inborn metabolic errors either through stimulation of residual enzyme activity in defective enzyme systems or by activation of alternate metabolic pathways. Both possibilities almost certainly depend on gross elevation of substrate concentrations. By contrast, only minimal in vivo oxidation of leucine appears possible in MSUD.

Thompson, G.N.; Walter, J.H.; Leonard, J.V.; Halliday, D. (Clinical Research Centre, Harrow (England))

1990-08-01

70

Up-regulation of photosynthesis and sucrose metabolism enzymes in young expanding leaves of sugarcane under elevated growth CO2  

Technology Transfer Automated Retrieval System (TEKTRAN)

Leaf photosynthesis (CER), levels of soluble protein and chlorophyll (Chl), and activities of key enzymes involved in C4 photosynthesis and sucrose metabolism were determined during leaf ontogeny for sugarcane grown at 360 and 720 ppm CO2. After leaf emergence, although leaf CER of both CO2 treatmen...

71

The RNA world and the origin of metabolic enzymes  

PubMed Central

An RNA world has been placed centre stage for explaining the origin of life. Indeed, RNA is the most plausible molecule able to form both a (self)-replicator and to inherit information, necessities for initiating genetics. However, in parallel with self-replication, the proto-organism had to obtain the ability to catalyse supply of its chemical constituents, including the ribonucleotide metabolites required to replicate RNA. Although the possibility of an RNA-catalysed metabolic network has been considered, it is to be questioned whether RNA molecules, at least on their own, possess the required catalytic capacities. An alternative scenario for the origin of metabolism involves chemical reactions that are based on environmental catalysts. Recently, we described a non-enzymatic glycolysis and pentose phosphate pathway-like reactions catalysed by metal ions [mainly Fe(II)] and phosphate, simple inorganic molecules abundantly found in Archaean sediments. While the RNA world can serve to explain the origin of genetics, the origin of the metabolic network might thus date back to constraints of environmental chemistry. Interestingly, considering a metal-catalysed origin of metabolism gives rise to an attractive hypothesis about how the first enzymes could have formed: simple RNA or (poly)peptide molecules could have bound the metal ions, and thus increased their solubility, concentration and accessibility. In a second step, this would have allowed substrate specificity to evolve. PMID:25109990

Ralser, Markus

2014-01-01

72

Activity of Glutathione-Metabolizing and Antioxidant Enzymes in Malignant and Benign Tumors of Human Lungs  

Microsoft Academic Search

We measured the content of glutathione and activity of glutathione-metabolizing and antioxidant enzymes superoxide dismutase and catalase in samples obtained from 52 patients with malignant lung tumors and 20 patients with benign lung tumors. The content of glutathione and activity of glutathione-metabolizing enzymes underwent similar changes, but these changes were most pronounced in malignant tumors. Antioxidant enzyme activity changed insignificantly

R. N. Korotkina; G. N. Matskevich; A. Sh. Devlikanova; A. A. Vishnevskii; A. G. Kunitsyn; A. A. Karelin

2002-01-01

73

The enzymes of biotin dependent CO2 metabolism: What structures reveal about  

E-print Network

REVIEW The enzymes of biotin dependent CO2 metabolism: What structures reveal about their reaction cofactor involved in carbon dioxide metabolism. Indeed, biotin- dependent enzymes are ubiquitous in nature. It is the ureido ring that functions as the CO2 carrier. A complete understanding of biotin-dependent enzymes

Holden, Hazel

74

Evolution of Domain Architectures and Catalytic Functions of Enzymes in Metabolic Systems  

E-print Network

Evolution of Domain Architectures and Catalytic Functions of Enzymes in Metabolic Systems Summit architectures and catalytic functions of enzymes constitute the centerpieces of a metabolic network. These types. In contrast, prokaryotic enzymes become more versatile by catalyzing multiple reactions with similar chemical

Yeang, Chen-Hsiang

75

Flavanone 3beta-hydroxylases from rice: key enzymes for favonol and anthocyanin biosynthesis.  

PubMed

Flavanone 3beta-hydroxylases (F3H) are key enzymes in the synthesis of flavonol and anthocyanin. In this study, three F3H cDNAs from Oryza sativa (OsF3H-1 approximately 3) were cloned by RT-PCR and expressed in E. coli as gluthatione S-transferase (GST) fusion proteins. The purified recombinant OsF3Hs used flavanone, naringenin and eriodictyol as substrates. The reaction products with naringen and eriodictyol were determined by nuclear magnetic resonance spectroscopy to be dihydrokaempferol and taxifolin, respectively. OsF3H-1 had the highest enzymatic activity whereas the overall expression of OsF3H-2 was highest in all tissues except seeds. Flavanone 3beta-hydroxylase could be a useful target for flavonoid metabolic engineering in rice. PMID:18413994

Kim, Jeong Ho; Lee, Yoon Jung; Kim, Bong Gyu; Lim, Yoongho; Ahn, Joong-Hoon

2008-04-30

76

FERULOYL ESTERASE - A KEY ENZYME IN BIOMASS DEGRADATION  

Technology Transfer Automated Retrieval System (TEKTRAN)

Feruloyl esterase forms a part of the enzyme complex that acts collectively and synergistically to completely hydrolyze xylan to its monomers. The enzyme has found potential uses in a wide variety of applications of interest to the agri-food and pharmaceutical industries. This review describes the...

77

Key Building Blocks via Enzyme-Mediated Synthesis  

NASA Astrophysics Data System (ADS)

Biocatalytic approaches to valuable building blocks in organic synthesis have emerged as an important tool in the last few years. While first applications were mainly based on hydrolases, other enzyme classes such as oxidoreductases or lyases moved into the focus of research. Nowadays, a vast number of biotransformations can be found in the chemical and pharmaceutical industries delivering fine chemicals or drugs. The mild reaction conditions, high stereo-, regio-, and chemoselectivities, and the often shortened reaction pathways lead to economical and ecological advantages of enzymatic conversions. Due to the enormous number of enzyme-mediated syntheses, the present chapter is not meant to be a complete review, but to deliver comprehensive insights into well established enzymatic systems and recent advances in the application of enzymes in natural product synthesis. Furthermore, it is focused on the most frequently used enzymes or enzyme classes not covered elsewhere in the present volume.

Fischer, Thomas; Pietruszka, Jörg

78

Hormonal Regulation of Hepatic Drug Metabolizing Enzyme Activity During Adolescence  

PubMed Central

Activities of drug metabolizing enzymes (DME) are known to change throughout the course of physical and sexual maturation with the greatest variability noted during infancy and adolescence. The mechanisms responsible for developmental regulation of DME are currently unknown. However, the hormonal changes of puberty/adolescence provide a theoretical framework for understanding biochemical regulation of DME activity during growth and maturation. Important information regarding potential influences of growth and sex hormones can also be extrapolated from studies evaluating changes in activities of DMEs occurring as a consequence of physiologic, pathologic and/or pharmacologic hormonal fluctuations. Collectively, current data support the hypothesis that isoform-specific alterations in DME activity during adolescence are mediated via sex and/or growth hormones. Characterization of the underlying biochemical alterations responsible for developmental changes in DME activity will require additional studies in which relationships between DME and important hormonal axes are evaluated during the course of pubertal development. PMID:18971926

Kennedy, M.J.

2009-01-01

79

Maternal Smoking, Xenobiotic Metabolizing Enzyme Gene Variants, and Gastroschisis Risk  

PubMed Central

Maternal smoking during pregnancy is one proposed risk factor for gastroschisis, but reported associations have been modest, suggesting that differences in genetic susceptibility might play a role. We included 108 non-Hispanic white and 62 Hispanic families who had infants with gastroschisis, and 1147 non-Hispanic white and 337 Hispanic families who had liveborn infants with no major structural birth defects (controls) in these analyses. DNA was extracted from buccal cells collected from infants and mothers, and information on periconceptional smoking history was obtained from maternal interviews, as part of the National Birth Defects Prevention Study. We analyzed five polymorphisms in three genes that code for enzymes involved in metabolism of some cigarette smoke constituents (CYP1A1, CYP1A2, and NAT2). Logistic regression models were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) independently for maternal smoking and maternal and infant gene variants, and to assess joint associations of maternal smoking and maternal or infant gene variants with gastroschisis. In analyses adjusted for maternal age at delivery and stratified by maternal race-ethnicity, we identified three suggestive associations among 30 potential associations with sufficient numbers to calculate ORs: CYP1A1*2A for non-Hispanic white mothers who smoked periconceptionally (aOR=0.38, 95% CI 0.15-0.98), and NAT2*6 for Hispanic non-smoking mothers (aOR=2.17, 95% CI 1.12-4.19) and their infants (aOR=2.11, 95% CI 1.00-4.48). This analysis does not support the occurrence of effect modification between periconceptional maternal smoking and most of the xenobiotic metabolizing enzyme gene variants assessed. PMID:24668907

Jenkins, Mary M.; Reefhuis, Jennita; Gallagher, Margaret L.; Mulle, Jennifer G.; Hoffmann, Thomas J.; Koontz, Deborah A.; Sturchio, Cynthia; Rasmussen, Sonja A.; Witte, John S.; Richter, Patricia; Honein, Margaret A.

2015-01-01

80

Maternal smoking, xenobiotic metabolizing enzyme gene variants, and gastroschisis risk.  

PubMed

Maternal smoking during pregnancy is one proposed risk factor for gastroschisis, but reported associations have been modest, suggesting that differences in genetic susceptibility might play a role. We included 108 non-Hispanic white and 62 Hispanic families who had infants with gastroschisis, and 1,147 non-Hispanic white and 337 Hispanic families who had liveborn infants with no major structural birth defects (controls) in these analyses. DNA was extracted from buccal cells collected from infants and mothers, and information on periconceptional smoking history was obtained from maternal interviews, as part of the National Birth Defects Prevention Study. We analyzed five polymorphisms in three genes that code for enzymes involved in metabolism of some cigarette smoke constituents (CYP1A1, CYP1A2, and NAT2). Logistic regression models were used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) independently for maternal smoking and maternal and infant gene variants, and to assess joint associations of maternal smoking and maternal or infant gene variants with gastroschisis. In analyses adjusted for maternal age at delivery and stratified by maternal race-ethnicity, we identified three suggestive associations among 30 potential associations with sufficient numbers to calculate ORs: CYP1A1*2A for non-Hispanic white mothers who smoked periconceptionally (aOR = 0.38, 95% CI 0.15-0.98), and NAT2*6 for Hispanic non-smoking mothers (aOR = 2.17, 95% CI 1.12-4.19) and their infants (aOR = 2.11, 95% CI 1.00-4.48). This analysis does not support the occurrence of effect modification between periconceptional maternal smoking and most of the xenobiotic metabolizing enzyme gene variants assessed. PMID:24668907

Jenkins, Mary M; Reefhuis, Jennita; Gallagher, Margaret L; Mulle, Jennifer G; Hoffmann, Thomas J; Koontz, Deborah A; Sturchio, Cynthia; Rasmussen, Sonja A; Witte, John S; Richter, Patricia; Honein, Margaret A

2014-06-01

81

Herb-drug interactions: focus on metabolic enzymes and transporters.  

PubMed

As the uses of herbal medicines from traditional natural products are increased, the need for pharmacokinetic studies and relevant data are also increased for safe pharmacotherapy. The market entry for the traditional herbal medicine is easier compared with that for synthetic drugs because of a lower regulatory barrier. Thus, the exact mechanisms for the absorption, distribution, metabolism and excretion of active components in herbal medicines and the potential herb-drug interactions are not always fully understood. Recently, there has been an increasing interest in pharmacokinetic studies of herbal remedies and relevant data of commonly used herbal remedies are accumulating in this field. In this review, the effects of nine botanicals (ginkgo, green tea, grapes, licorice, saw palmetto, garlic milk thistle, ginseng and St. John's wort) on metabolic enzymes and transporters affecting absorption and disposition of herbal products are summarized. The source of samples (extracts and individual components), the species (human and animal) and in vivo and in vitro systems were separately reviewed for a better understanding of herb-drug interactions. PMID:22139685

Choi, Young Hee; Chin, Young-Won; Kim, Yoon Gyoon

2011-11-01

82

Ammonium Metabolism Enzymes Aid Helicobacter pylori Acid Resistance  

PubMed Central

The gastric pathogen Helicobacter pylori possesses a highly active urease to support acid tolerance. Urea hydrolysis occurs inside the cytoplasm, resulting in the production of NH3 that is immediately protonated to form NH4+. This ammonium must be metabolized or effluxed because its presence within the cell is counterproductive to the goal of raising pH while maintaining a viable proton motive force (PMF). Two compatible hypotheses for mitigating intracellular ammonium toxicity include (i) the exit of protonated ammonium outward via the UreI permease, which was shown to facilitate diffusion of both urea and ammonium, and/or (ii) the assimilation of this ammonium, which is supported by evidence that H. pylori assimilates urea nitrogen into its amino acid pools. We investigated the second hypothesis by constructing strains with altered expression of the ammonium-assimilating enzymes glutamine synthetase (GS) and glutamate dehydrogenase (GDH) and the ammonium-evolving periplasmic enzymes glutaminase (Ggt) and asparaginase (AsnB). H. pylori strains expressing elevated levels of either GS or GDH are more acid tolerant than the wild type, exhibit enhanced ammonium production, and are able to alkalize the medium faster than the wild type. Strains lacking the genes for either Ggt or AsnB are acid sensitive, have 8-fold-lower urea-dependent ammonium production, and are more acid sensitive than the parent. Additionally, we found that purified H. pylori GS produces glutamine in the presence of Mg2+ at a rate similar to that of unadenylated Escherichia coli GS. These data reveal that all four enzymes contribute to whole-cell acid resistance in H. pylori and are likely important for assimilation and/or efflux of urea-derived ammonium. PMID:24936052

Miller, Erica F.

2014-01-01

83

Ghrelin O Acyl Transferase (GOAT) as a Novel Metabolic Regulatory Enzyme  

PubMed Central

Background: Obesity and Type 2 Diabetes Mellitus (T2DM) presents a growing threat to the global health. Evidences highlight an important role of ghrelin as a key regulator of glucose metabolism. The physiological functions of ghrelin are mediated by enzyme ghrelin-O-acyltransferase (GOAT) which is capable of generating the active form of this metabolic hormone. However, its exact mechanism of action and influence on energy balance and glucose metabolism is yet to be explored. Objectives: To review the physiological role of GOAT in the regulation of energy balance and glucose metabolism and explore the potential therapeutic avenues of modulators of GOAT to counter the progression of obesity and T2DM. Methods: Publications were sought through electronic searches. The bibliographies of all papers, book, chapters and editorials were scanned and hand searches were also conducted for journals, and conference proceedings. Conclusion: GOAT peptide modulates the insulin secretion as well as insulin sensitivity. Modulators of GOAT signaling like inhibitors of GOAT increases insulin secretion, enhance peripheral insulin sensitivity and thus counters obesity and T2DM. Modulators of GOAT can be a probable therapy for modifying food intake and for countering obesity and T2DM.

Gaidhane, Shilpa; Gaidhane, Abhay M.; Simkhada, Padam; Zahiruddin, Quazi Syed

2015-01-01

84

Effect of Chromium(VI) Toxicity on Enzymes of Nitrogen Metabolism in Clusterbean (Cyamopsis tetragonoloba L.)  

PubMed Central

Heavy metals are the intrinsic component of the environment with both essential and nonessential types. Their excessive levels pose a threat to plant growth and yield. Also, some heavy metals are toxic to plants even at very low concentrations. The present investigation (a pot experiment) was conducted to determine the affects of varying chromium(VI) levels (0.0, 0.5, 1.0, 2.0, and 4.0?mg chromium(VI)?kg?1 soil in the form of potassium dichromate) on the key enzymes of nitrogen metabolism in clusterbean. Chromium treatment adversely affect nitrogenase, nitrate reductase, nitrite reductase, glutamine synthetase, and glutamate dehydrogenase in various plant organs at different growth stages as specific enzyme activity of these enzymes decreased with an increase in chromium(VI) levels from 0 to 2.0?mg chromium(VI)?kg?1 soil and 4.0?mg chromium(VI)?kg?1 soil was found to be lethal to clusterbean plants. In general, the enzyme activity increased with advancement of growth to reach maximum at flowering stage and thereafter decreased at grain filling stage. PMID:24744916

Sangwan, Punesh; Joshi, U. N.

2014-01-01

85

(Pyruvate decarboxylase: A key enzyme for alcohol production)  

SciTech Connect

The overall objectives of this grant are to investigate the properties and regulation of pyruvate decarboxylase from Z. mobilis and to identify the unique features which allow its high level of expression. Steps in the project include enzyme purification and antibody production, characterization of physical and kinetic properties, investigations of how environmental conditions affect the level of pyruvate decarboxylase, its biosynthesis and turnover, cloning and sequencing, investigations of the effects of increased gene dosage, control of m-RNA synthesis and turnover, and promoter structure. 16 refs.

Not Available

1989-01-01

86

A QUANTITATIVE MODEL FOR XENOBIOTIC METABOLIZING ENZYME (XME) INDUCTION REGULATED BY THE PREGNANE X RECEPTOR (PXR)  

EPA Science Inventory

The nuclear receptor, PXR, is an integral part of the regulation of hepatic metabolism. It has been shown to regulate specific CYPs (phase I drug-metabolizing enzymes) as well as certain phase II drug metabolism activities, including UDP-glucuronosyl transferase (UGT), sulfotran...

87

Fermentation, Respiration & Enzyme Specificity: A Simple Device & Key Experiments with Yeast.  

ERIC Educational Resources Information Center

Using graphs and diagrams, the authors describe a simple fermentation chamber and provide key experiments that can be used in the classroom to give students meaningful insight into metabolic processes. (ZWH)

Reinking, Larry N.; And Others

1994-01-01

88

Mutations in Phosphoinositide Metabolizing Enzymes and Human Disease  

NSDL National Science Digital Library

Phosphoinositides are implicated in the regulation of a wide variety of cellular functions. Their importance in cellular and organismal physiology is underscored by the growing number of human diseases linked to perturbation of kinases and phosphatases that catalyze interconversion from one phosphoinositide to another. Many such enzymes are attractive targets for therapeutic interventions. Here, we review diseases linked to inheritable or somatic mutations of these enzymes. Phosphatidylinositol (PtdIns), a membrane phospholipid, can be reversibly phosphorylated at the 3, 4, and 5 positions of the inositol ring to generate seven phosphoinositides [PI3P, PI4P, PI5P, PI(3,4)P2, PI(4,5)P2, PI(3,5)P2, and PI(3,4,5)P3] (FIGURE 1A). The importance of this metabolism in cell regulation was first established in the context of studies on stimulus-secretion coupling. It was found that many stimuli that trigger secretion also trigger enhanced turnover of PtdIns and phosphoinositides (42). Subsequently, it became clear that phospholipase C-dependent hydrolysis of PI(4,5)P2 to generate the second messenger molecules diacyl glycerol and Ins(1,4,5)P3 (IP3) is a mechanism through which many cell surface receptors, including many receptors that stimulate secretion, transduce their signals (10). Diacyl glycerol binds and regulates protein kinase C and a variety of other effectors, whereas IP3 triggers calcium release from the endoplasmic reticulum (10, 42). In another signal transduction pathway, PI(4,5)P2 is cleaved by phospholipase A2 to generate arachidonic acid, a precursor of many signaling molecules.

Heather J. McCrea (Yale University)

2009-02-01

89

Methanol metabolism in yeasts: Regulation of the synthesis of catabolic enzymes  

Microsoft Academic Search

The regulation of the synthesis of four dissimilatory enzymes involved in methanol metabolism, namely alcohol oxidase, formaldehyde dehydrogenase, formate dehydrogenase and catalase was investigated in the yeasts Hansenula polymorpha and Kloeckera sp. 2201. Enzyme profiles in cell-free extracts of the two organisms grown under glucose limitation at various dilution rates, suggested that the synthesis of these enzymes is controlled by

Th. Egli; J. P. van Dijken; M. Veenhuis; W. Harder; A. Fiechter

1980-01-01

90

Extraction and study of enzymes linked to malate metabolism in tree leaves  

E-print Network

Extraction and study of enzymes linked to malate metabolism in tree leaves D. Gerant, A. Citerne, C, 1984). Enzymes implicated in synthesis and catabolism of this substrate are widely distributed in the cellular compartments. If these enzymes have become well known in herbaceous plants (Macrae, 1971; Davis

Paris-Sud XI, Université de

91

Molecular characterization of genes encoding enzymes of the metabolic pathways in Neocallimastix frontalis.  

E-print Network

Molecular characterization of genes encoding enzymes of the metabolic pathways in Neocallimastix. Enzyme activities leading to the formation of succinate, lactate, formate and ethanol are associated with the cytoplasmic fraction, while the enzymes leading to the formation of the main fermentation products H2, C02

Boyer, Edmond

92

Specificity of Non-Michaelis-Menten Enzymes: Necessary Information for Analyzing Metabolic Pathways  

E-print Network

of enzyme kinetics and mechanism, including pH8 and temperature dependence,9 progress curve analysis10 consider only effects of competing substrates on the rates of individual enzyme-catalyzed reactionsSpecificity of Non-Michaelis-Menten Enzymes: Necessary Information for Analyzing Metabolic Pathways

93

A Multiscale Approach to Modelling Drug Metabolism by Membrane-Bound Cytochrome P450 Enzymes  

PubMed Central

Cytochrome P450 enzymes are found in all life forms. P450s play an important role in drug metabolism, and have potential uses as biocatalysts. Human P450s are membrane-bound proteins. However, the interactions between P450s and their membrane environment are not well-understood. To date, all P450 crystal structures have been obtained from engineered proteins, from which the transmembrane helix was absent. A significant number of computational studies have been performed on P450s, but the majority of these have been performed on the solubilised forms of P450s. Here we present a multiscale approach for modelling P450s, spanning from coarse-grained and atomistic molecular dynamics simulations to reaction modelling using hybrid quantum mechanics/molecular mechanics (QM/MM) methods. To our knowledge, this is the first application of such an integrated multiscale approach to modelling of a membrane-bound enzyme. We have applied this protocol to a key human P450 involved in drug metabolism: CYP3A4. A biologically realistic model of CYP3A4, complete with its transmembrane helix and a membrane, has been constructed and characterised. The dynamics of this complex have been studied, and the oxidation of the anticoagulant R-warfarin has been modelled in the active site. Calculations have also been performed on the soluble form of the enzyme in aqueous solution. Important differences are observed between the membrane and solution systems, most notably for the gating residues and channels that control access to the active site. The protocol that we describe here is applicable to other membrane-bound enzymes. PMID:25033460

Sansom, Mark S. P.; Mulholland, Adrian J.

2014-01-01

94

MIT-led study finds turning on key enzyme blocks tumor formation  

Cancer.gov

Unlike ordinary cells, cancer cells devote most of their energy to reproducing themselves. To do this, they must trigger alternative metabolic pathways that produce new cellular building blocks, such as DNA, carbohydrates and lipids. Chemical compounds that disrupt an enzyme critical to this metabolic diversion prevent tumors from forming in mice, according to an MIT-led study appearing online in Nature Chemical Biology on Aug. 26. MIT is home to the David H. Koch Institute for Integrative Cancer Research.

95

communication between distal protein sites and the enzyme catalytic core Key feature of the catalytic cycle of TNF-{alpha} converting enzyme involves  

E-print Network

of the catalytic cycle of TNF-{alpha} converting enzyme involves Ariel Solomon, Barak Akabayov, Anatoly Frenkel.pnas.org/misc/reprints.shtml To order reprints, see: Notes: #12;Key feature of the catalytic cycle of TNF- converting enzyme involvescommunication between distal protein sites and the enzyme catalytic core Key feature

Frenkel, Anatoly

96

Evidence for key enzymatic controls on metabolism of Arctic river organic matter.  

PubMed

Permafrost thaw in the Arctic driven by climate change is mobilizing ancient terrigenous organic carbon (OC) into fluvial networks. Understanding the controls on metabolism of this OC is imperative for assessing its role with respect to climate feedbacks. In this study, we examined the effect of inorganic nutrient supply and dissolved organic matter (DOM) composition on aquatic extracellular enzyme activities (EEAs) in waters draining the Kolyma River Basin (Siberia), including permafrost-derived OC. Reducing the phenolic content of the DOM pool resulted in dramatic increases in hydrolase EEAs (e.g., phosphatase activity increased >28-fold) supporting the idea that high concentrations of polyphenolic compounds in DOM (e.g., plant structural tissues) inhibit enzyme synthesis or activity, limiting OC degradation. EEAs were significantly more responsive to inorganic nutrient additions only after phenolic inhibition was experimentally removed. In controlled mixtures of modern OC and thawed permafrost endmember OC sources, respiration rates per unit dissolved OC were 1.3-1.6 times higher in waters containing ancient carbon, suggesting that permafrost-derived OC was more available for microbial mineralization. In addition, waters containing ancient permafrost-derived OC supported elevated phosphatase and glucosidase activities. Based on these combined results, we propose that both composition and nutrient availability regulate DOM metabolism in Arctic aquatic ecosystems. Our empirical findings are incorporated into a mechanistic conceptual model highlighting two key enzymatic processes in the mineralization of riverine OM: (i) the role of phenol oxidase activity in reducing inhibitory phenolic compounds and (ii) the role of phosphatase in mobilizing organic P. Permafrost-derived DOM degradation was less constrained by this initial 'phenolic-OM' inhibition; thus, informing reports of high biological availability of ancient, permafrost-derived DOM with clear ramifications for its metabolism in fluvial networks and feedbacks to climate. PMID:24115585

Mann, Paul J; Sobczak, William V; Larue, Madeleine M; Bulygina, Ekaterina; Davydova, Anna; Vonk, Jorien E; Schade, John; Davydov, Sergei; Zimov, Nikita; Holmes, Robert M; Spencer, Robert G M

2014-04-01

97

Peroxisome proliferators alter the expression of estrogen-metabolizing enzymes.  

PubMed

Exposure to some peroxisome proliferator chemicals (PPC) leads to toxic effects on sex organ function possibly by alterations of steroid hormone metabolism. A systematic search for genes whose mRNA levels are modulated by the PPC WY-14643 (WY) was carried out in rat liver, a site of steroid hormone metabolism. The sequence of one up-regulated cDNA (2480 bp) was predicted to encode a protein of 735 amino acids with 82% identity to the porcine 17 beta-hydroxysteroid dehydrogenase type IV (HSD IV) originally isolated as a 17 beta-estradiol dehydrogenase. The rat HSD IV was localized to peroxisomes and was regulated by diverse PPC by two distinct mechanisms. Induction of HSD IV and acyl-CoA oxidase (ACO) proteins in rat liver at different treatment times and concentrations of gemfibrozil (GEM) and di-n-butyl phthalate (DBP) were almost identical, suggesting that HSD IV mRNA induction involves the peroxisome proliferator-activated receptor alpha, a regulator of ACO. In contrast, HSD IV protein levels were only weakly induced by WY, a strong inducer of ACO protein, even though the levels of both HSD IV and ACO mRNA were strongly stimulated by WY. Thus HSD IV protein levels were uniquely regulated pretranslationally by WY. In addition to HSD IV we also identified the male-specific alpha 2u-globulin as a PPC down-regulated gene. This prompted us to examine the expression of another male-specific gene, CYP2C11, that catalyzes the hydroxylations of estradiol at the 2 and 16 alpha positions. Cyp2C11 protein expression in rat liver was either decreased or completely abolished after a 3-week treatment by GEM or WY, respectively. Decreased expression of enzymes which inactivate estradiol including Cyp2C11, and the reported increased expression of aromatase may explain why male rats exposed to diverse PPC have higher serum estradiol levels. These higher estradiol levels in male rats have been thought to be mechanistically linked to Leydig cell hyperplasia and adenomas. Increased conversion of estradiol to the less active estrone by HSD IV induction may explain how exposure to the phthalate di-(2-ethylhexyl) phthalate leads to decreases in serum estradiol levels and suppression of ovulation in female rats. PMID:9209713

Corton, J C; Bocos, C; Moreno, E S; Merritt, A; Cattley, R C; Gustafsson, J A

1997-01-01

98

New enzymes involved in aerobic benzoate metabolism in Azoarcus evansii.  

PubMed

A new principle of aerobic aromatic metabolism has been postulated, which is in contrast to the known pathways. In various bacteria the aromatic substrate benzoate is first converted to its coenzyme A (CoA) thioester, benzoyl-CoA, which is subsequently attacked by an oxygenase, followed by a non-oxygenolytic fission of the ring. We provide evidence for this hypothesis and show that benzoyl-CoA conversion in the bacterium Azoarcus evansii requires NADPH, O(2) and two protein components, BoxA and BoxB. BoxA is a homodimeric 46 kDa iron-sulphur-flavoprotein, which acts as reductase. In the absence of BoxB, BoxA catalyses the benzoyl-CoA stimulated artificial transfer of electrons from NADPH to O(2) via free FADH(2) to produce H(2)O(2). Physiologically, BoxA uses NADPH to reduce BoxB, a monomeric 55 kDa iron-protein that acts as benzoyl-CoA oxygenase. The product of benzoyl-CoA oxidation was identified by NMR spectroscopy as its dihydrodiol derivative, 2,3-dihydro-2,3-dihydroxybenzoyl-CoA. This suggests that BoxBA act as a benzoyl-CoA dioxygenase/reductase. Unexpectedly, benzoyl-CoA transformation by BoxBA was greatly stimulated when another enoyl-CoA hydratase/isomerase-like protein, BoxC, was added that catalysed the further transformation of the dihydrodiol product formed from benzoyl-CoA. The benzoyl-CoA oxygenase system has very low similarity to known (di)oxygenase systems and is the first member of a new enzyme family. PMID:15458418

Zaar, Annette; Gescher, Johannes; Eisenreich, Wolfgang; Bacher, Adelbert; Fuchs, Georg

2004-10-01

99

Fungal colonization and enzyme-mediated metabolism of waste coal by Neosartorya fischeri strain ECCN 84.  

PubMed

Colonization and oxidative metabolism of South African low-rank discard coal by the fungal strain ECCN 84 previously isolated from a coal environment and identified as Neosartorya fischeri was investigated. Results show that waste coal supported fungal growth. Colonization of waste coal particles by N. fischeri ECCN 84 was associated with the formation of compact spherical pellets or sclerotia-like structures. Dissection of the pellets from liquid cultures revealed a nucleus of "engulfed" coal which when analyzed by energy dispersive X-ray spectroscopy showed a time-dependent decline in weight percentage of elemental carbon and an increase in elemental oxygen. Proliferation of peroxisomes in hyphae attached to coal particles and increased extracellular laccase activity occurred after addition of waste coal to cultures of N. fischeri ECCN 84. These results support a role for oxidative enzyme action in the biodegradation of coal and suggest that extracellular laccase is a key component in this process. PMID:25273148

Sekhohola, Lerato Mary; Isaacs, Michelle Louise; Cowan, Ashton Keith

2014-10-01

100

Articular cartilage chondrocytes express aromatase and use enzymes involved in estrogen metabolism  

PubMed Central

Introduction Sex hormones, especially estrogens, have been implicated in articular cartilage metabolism and the pathogenesis of postmenopausal osteoarthritis. The conversion by aromatase (CYP19A1) of androstenedione into estrone (E1) and of testosterone into 17?-estradiol (E2) plays a key role in the endogenous synthesis of estrogens in tissue. Methods We analyzed the expression of aromatase (CYP19A1) in immortalized C-28/I2 and T/C-28a2 chondrocytes, as well as in cultured primary human articular chondrocytes and human articular cartilage tissue, by means of RT-PCR, Western blotting and immunohistochemistry. By means of quantitative RT-PCR and enzyme-linked immunosorbent assay, we also determined whether the aromatase inhibitor letrozole influences estrogen metabolism of cultured chondrocytes in immortalized C-28/I2 chondrocytes. Results Aromatase mRNA was detected in both immortalized chondrocyte cell lines, in cultured primary human chondrocytes, and in human articular cartilage tissue. By means of Western blot analysis, aromatase was detected at the protein level in articular cartilage taken from various patients of both sexes and different ages. Cultured primary human articular chondrocytes, C-28/I2 and T/C-28a2, and human articular cartilage tissue reacted with antibodies for aromatase. Incubation of C-28/I2 chondrocytes with 10?11 M to 10?7 M letrozole as an aromatase inhibitor revealed significantly increased amounts of the mRNAs of the enzyme cytochrome P4501A1 (CYP1A1), which is involved in the catagen estrogen metabolism, and of the estrogen receptors ER-? and ER-?. Concomitantly, synthesis of estrone (E1) was significantly downregulated after incubation with letrozole. Conclusions We demonstrate that human articular cartilage expresses aromatase at the mRNA and protein levels. Blocking of estrone synthesis by the aromatase inhibitor letrozole is counteracted by an increase in ER-? and ER-?. In addition, CYP1A1, an enzyme involved in catabolic estrogen metabolism, is upregulated. This suggests that articular chondrocytes use ERs functionally. The role of endogenous synthesized estrogens in articular cartilage health remains to be elucidated. PMID:24725461

2014-01-01

101

Comparative genomic and phylogenetic investigation of the xenobiotic metabolizing arylamine N-acetyltransferase enzyme family  

Technology Transfer Automated Retrieval System (TEKTRAN)

Arylamine N-acetyltransferases (NATs) are xenobiotic metabolizing enzymes characterized in several bacteria and eukaryotic organisms. We report a comprehensive phylogenetic analysis employing an exhaustive dataset of NAT-homologous sequences recovered through inspection of 2445 genomes. We describe ...

102

Sensor potency of the moonlighting enzyme-decorated cytoskeleton: the cytoskeleton as a metabolic sensor  

PubMed Central

Background There is extensive evidence for the interaction of metabolic enzymes with the eukaryotic cytoskeleton. The significance of these interactions is far from clear. Presentation of the hypothesis In the cytoskeletal integrative sensor hypothesis presented here, the cytoskeleton senses and integrates the general metabolic activity of the cell. This activity depends on the binding to the cytoskeleton of enzymes and, depending on the nature of the enzyme, this binding may occur if the enzyme is either active or inactive but not both. This enzyme-binding is further proposed to stabilize microtubules and microfilaments and to alter rates of GTP and ATP hydrolysis and their levels. Testing the hypothesis Evidence consistent with the cytoskeletal integrative sensor hypothesis is presented in the case of glycolysis. Several testable predictions are made. There should be a relationship between post-translational modifications of tubulin and of actin and their interaction with metabolic enzymes. Different conditions of cytoskeletal dynamics and enzyme-cytoskeleton binding should reveal significant differences in local and perhaps global levels and ratios of ATP and GTP. The different functions of moonlighting enzymes should depend on cytoskeletal binding. Implications of the hypothesis The physical and chemical effects arising from metabolic sensing by the cytoskeleton would have major consequences on cell shape, dynamics and cell cycle progression. The hypothesis provides a framework that helps the significance of the enzyme-decorated cytoskeleton be determined. PMID:23398642

2013-01-01

103

Biochemical strategies of overwintering in the gall gly larva, Eurosta solidaginis : Effect of low temperature acclimation on the activities of enzymes of intermediary metabolism  

Microsoft Academic Search

1.The activities of some enzymes of intermediary metabolism, including those involved in glycerol and sorbitol synthesis, were measured in the third instar larvae of the gall fly,Eurosta solidaginis sampled during a controlled (1°C per day decrease) low temperature acclimation of the larvae from 15 to 30°C.2.Low temperature acclimation resulted in increased activities of three key enzymes of carbohydrate catabolism: phosphorylase,

Kenneth B. Storey; Janet M. Storey

1981-01-01

104

Serum Lipids, Lipoproteins, and Lipid Metabolizing Enzymes in Identical Twins Discordant for Obesity  

Microsoft Academic Search

Obesity is associated with adverse changes in plasma lipoprotein metabolism, but it is not known completely how this association is modified by genetic factors. We assessed the contribution of obesity to serum lipid and lipoprotein levels and lipid metabolizing enzyme activities by examining 23 identical twin pairs (9 male, 14 female) who had, on the average, an 18-kg intrapair difference

TAPANI RONNEMAA; JUKKA MARNIEMI; MARKKU J. SAVOLAINEN; Y. ANTERO KESANIEMI; CHRISTIAN EHNHOLM; CLAUDE BOUCHARD; MARKKU KOSKENVUO

2010-01-01

105

RESVERATROL IN HUMAN HEPATOMA HEPG2 CELLS: METABOLISM AND INDUCIBILITY OF DETOXIFYING ENZYMES  

E-print Network

1 RESVERATROL IN HUMAN HEPATOMA HEPG2 CELLS: METABOLISM AND INDUCIBILITY OF DETOXIFYING ENZYMES-Université de Bourgogne, Dijon, France Running title page : Resveratrol metabolism in human liver-derived HepG2 transcriptase polymerase chain reaction. ABSTRACT trans-Resveratrol is a polyphenol present in several plant

Paris-Sud XI, Université de

106

Activities of amino acid metabolizing enzymes in the stomach and small intestine of developing rats  

E-print Network

Activities of amino acid metabolizing enzymes in the stomach and small intestine of developing rats amino acids. Introduction. The stomach and small intestine clearly have a common embryonic origin ; both, involded in amino acid metabolism. The stomach and intestine play an important role in the post

Boyer, Edmond

107

Axonal and dendritic localization of mRNAs for glycogen-metabolizing enzymes in cultured rodent neurons  

PubMed Central

Background Localization of mRNAs encoding cytoskeletal or signaling proteins to neuronal processes is known to contribute to axon growth, synaptic differentiation and plasticity. In addition, a still increasing spectrum of mRNAs has been demonstrated to be localized under different conditions and developing stages thus reflecting a highly regulated mechanism and a role of mRNA localization in a broad range of cellular processes. Results Applying fluorescence in-situ-hybridization with specific riboprobes on cultured neurons and nervous tissue sections, we investigated whether the mRNAs for two metabolic enzymes, namely glycogen synthase (GS) and glycogen phosphorylase (GP), the key enzymes of glycogen metabolism, may also be targeted to neuronal processes. If it were so, this might contribute to clarify the so far enigmatic role of neuronal glycogen. We found that the mRNAs for both enzymes are localized to axonal and dendritic processes in cultured lumbar spinal motoneurons, but not in cultured trigeminal neurons. In cultured cortical neurons which do not store glycogen but nevertheless express glycogen synthase, the GS mRNA is also subject to axonal and dendritic localization. In spinal motoneurons and trigeminal neurons in situ, however, the mRNAs could only be demonstrated in the neuronal somata but not in the nerves. Conclusions We could demonstrate that the mRNAs for major enzymes of neural energy metabolism can be localized to neuronal processes. The heterogeneous pattern of mRNA localization in different culture types and developmental stages stresses that mRNA localization is a versatile mechanism for the fine-tuning of cellular events. Our findings suggest that mRNA localization for enzymes of glycogen metabolism could allow adaptation to spatial and temporal energy demands in neuronal events like growth, repair and synaptic transmission. PMID:24898526

2014-01-01

108

The Emerging Significance of Drug Transporters and Metabolizing Enzymes to Ophthalmic Drug Design  

Microsoft Academic Search

SUMMARY Ophthalmic drugs typically achieve less than 10% ocular bioavailability. A drug applied to the surface of the eye may cross ocular–blood barriers, where it may encounter cellular transporters and metabolizing enzymes, before it is distributed to the site of action. Characterization of ocular membrane transporters, enzyme systems and their respective substrate selectivity has provided new insight into the roles

Mayssa Attar; Jie Shen

109

Xenobiotic metabolizing enzyme (XME) expression in aging humans.  

EPA Science Inventory

In the presence of foreign compounds, metabolic homeostasis of the organism is maintained by the liver?s ability to detoxify and eliminate these xenobiotics. This is accomplished, in part, by the expression of XMEs, which metabolize xenobiotics and determine whether exposure will...

110

Decoding key nodes in the metabolism of cancer cells: sugar & spice and all things nice  

PubMed Central

In the past 5 years, a convergence of studies has resulted in a broad appreciation in the cancer research community that reprogramming of cellular metabolism may be more central to cancer than appreciated in the past 30 years. The re-emergence of cancer metabolism stems in part from discoveries that a number of common oncogenes and tumor suppressor genes more directly control cell metabolism than previously thought. In addition, a number of what would previously have been called “card-carrying” metabolic enzymes have been identified as human tumor suppressors or oncogenes, causally mutated in a variety of human cancers. This growing appreciation of the role of altered cell metabolism has led to further investigation into the rate-limiting proteins involved in different aspects of the unique metabolism of tumor cells. Targeting cancer metabolism with drugs requires a therapeutic window in which tumor cells, compared to normal tissues, have a greater dependence on specific metabolic enzymes. Themes that have emerged in the past decade of developing oncogene-targeted cancer therapeutics suggest that tumors with distinct oncogenic lesions are likely to require drugs that target distinct metabolic pathways. Ultimately, the hope is that detailed knowledge of oncogene and tumor suppressor gene functions and their effects on metabolism will lead to drug combinations that will be far more effective in treating cancers. PMID:22242042

2012-01-01

111

De novo transcriptome characterization of Lilium 'Sorbonne' and key enzymes related to the flavonoid biosynthesis.  

PubMed

Lily is an important cut-flower and bulb crop in the commercial market. Here, transcriptome profiling of Lilium 'Sorbonne' was conducted through de novo sequencing based on Illumina platform. This research aims at revealing basic information and data that can be used for applied purposes especially the molecular regulatory information on flower color formation in lily. In total, 36,920,680 short reads which corresponded to 3.32 GB of total nucleotides, were produced through transcriptome sequencing. These reads were assembled into 39,636 Unigenes, of which 30,986 were annotated in Nr, Nt, Swiss-Prot, KEGG, COG, GO databases. Based on the three public protein databases, a total of 32,601 coding sequences were obtained. Meanwhile, 19,242 Unigenes were assigned to 128 KEGG pathways. Those with the greatest representation by unique sequences were for ''metabolic pathways'' (5,406 counts, 28.09 %). Our transcriptome revealed 156 Unigenes that encode key enzymes in the flavonoid biosynthesis pathway including CHS, CHI, F3H, FLS, DFR, etc. MISA software identified 2,762 simple sequence repeats, from which 1,975 primers pairs were designed. Over 2,762 motifs were identified, of which the most frequent was AG/CT (659, 23.86 %), followed by A/T (615, 22.27 %) and CCG/CGG (416, 15.06 %). Based on the results, we believe that the color formation of the Lilium 'Sorbonne' flower was mainly controlled by the flavonoid biosynthesis pathway. Additionally, this research provides initial genetic resources that will be valuable to the lily community for other molecular biology research, and the SSRs will facilitate marker-assisted selection in lily breeding. PMID:25307066

Zhang, Ming-Fang; Jiang, Ling-Min; Zhang, Dong-Mei; Jia, Gui-Xia

2015-02-01

112

Predicting Metabolic Pathways of Small Molecules and Enzymes Based on Interaction Information of Chemicals and Proteins  

PubMed Central

Metabolic pathway analysis, one of the most important fields in biochemistry, is pivotal to understanding the maintenance and modulation of the functions of an organism. Good comprehension of metabolic pathways is critical to understanding the mechanisms of some fundamental biological processes. Given a small molecule or an enzyme, how may one identify the metabolic pathways in which it may participate? Answering such a question is a first important step in understanding a metabolic pathway system. By utilizing the information provided by chemical-chemical interactions, chemical-protein interactions, and protein-protein interactions, a novel method was proposed by which to allocate small molecules and enzymes to 11 major classes of metabolic pathways. A benchmark dataset consisting of 3,348 small molecules and 654 enzymes of yeast was constructed to test the method. It was observed that the first order prediction accuracy evaluated by the jackknife test was 79.56% in identifying the small molecules and enzymes in a benchmark dataset. Our method may become a useful vehicle in predicting the metabolic pathways of small molecules and enzymes, providing a basis for some further analysis of the pathway systems. PMID:23029334

Feng, Kai-Yan; Huang, Tao; Jiang, Yang

2012-01-01

113

Computational prediction of metabolism: sites, products, SAR, P450 enzyme dynamics, and mechanisms.  

PubMed

Metabolism of xenobiotics remains a central challenge for the discovery and development of drugs, cosmetics, nutritional supplements, and agrochemicals. Metabolic transformations are frequently related to the incidence of toxic effects that may result from the emergence of reactive species, the systemic accumulation of metabolites, or by induction of metabolic pathways. Experimental investigation of the metabolism of small organic molecules is particularly resource demanding; hence, computational methods are of considerable interest to complement experimental approaches. This review provides a broad overview of structure- and ligand-based computational methods for the prediction of xenobiotic metabolism. Current computational approaches to address xenobiotic metabolism are discussed from three major perspectives: (i) prediction of sites of metabolism (SOMs), (ii) elucidation of potential metabolites and their chemical structures, and (iii) prediction of direct and indirect effects of xenobiotics on metabolizing enzymes, where the focus is on the cytochrome P450 (CYP) superfamily of enzymes, the cardinal xenobiotics metabolizing enzymes. For each of these domains, a variety of approaches and their applications are systematically reviewed, including expert systems, data mining approaches, quantitative structure-activity relationships (QSARs), and machine learning-based methods, pharmacophore-based algorithms, shape-focused techniques, molecular interaction fields (MIFs), reactivity-focused techniques, protein-ligand docking, molecular dynamics (MD) simulations, and combinations of methods. Predictive metabolism is a developing area, and there is still enormous potential for improvement. However, it is clear that the combination of rapidly increasing amounts of available ligand- and structure-related experimental data (in particular, quantitative data) with novel and diverse simulation and modeling approaches is accelerating the development of effective tools for prediction of in vivo metabolism, which is reflected by the diverse and comprehensive data sources and methods for metabolism prediction reviewed here. This review attempts to survey the range and scope of computational methods applied to metabolism prediction and also to compare and contrast their applicability and performance. PMID:22339582

Kirchmair, Johannes; Williamson, Mark J; Tyzack, Jonathan D; Tan, Lu; Bond, Peter J; Bender, Andreas; Glen, Robert C

2012-03-26

114

Computational Prediction of Metabolism: Sites, Products, SAR, P450 Enzyme Dynamics, and Mechanisms  

PubMed Central

Metabolism of xenobiotics remains a central challenge for the discovery and development of drugs, cosmetics, nutritional supplements, and agrochemicals. Metabolic transformations are frequently related to the incidence of toxic effects that may result from the emergence of reactive species, the systemic accumulation of metabolites, or by induction of metabolic pathways. Experimental investigation of the metabolism of small organic molecules is particularly resource demanding; hence, computational methods are of considerable interest to complement experimental approaches. This review provides a broad overview of structure- and ligand-based computational methods for the prediction of xenobiotic metabolism. Current computational approaches to address xenobiotic metabolism are discussed from three major perspectives: (i) prediction of sites of metabolism (SOMs), (ii) elucidation of potential metabolites and their chemical structures, and (iii) prediction of direct and indirect effects of xenobiotics on metabolizing enzymes, where the focus is on the cytochrome P450 (CYP) superfamily of enzymes, the cardinal xenobiotics metabolizing enzymes. For each of these domains, a variety of approaches and their applications are systematically reviewed, including expert systems, data mining approaches, quantitative structure–activity relationships (QSARs), and machine learning-based methods, pharmacophore-based algorithms, shape-focused techniques, molecular interaction fields (MIFs), reactivity-focused techniques, protein–ligand docking, molecular dynamics (MD) simulations, and combinations of methods. Predictive metabolism is a developing area, and there is still enormous potential for improvement. However, it is clear that the combination of rapidly increasing amounts of available ligand- and structure-related experimental data (in particular, quantitative data) with novel and diverse simulation and modeling approaches is accelerating the development of effective tools for prediction of in vivo metabolism, which is reflected by the diverse and comprehensive data sources and methods for metabolism prediction reviewed here. This review attempts to survey the range and scope of computational methods applied to metabolism prediction and also to compare and contrast their applicability and performance. PMID:22339582

2012-01-01

115

Problems and perspectives of phenotyping for drug-metabolizing enzymes in man.  

PubMed

Pronounced interindividual differences in drug disposition are mainly caused by differences in the activity of liver drug-metabolizing enzymes. These depend on known and unknown covariates, including genetic as well as environmental factors. Phenotyping, i.e. assessment of enzyme activities in vivo after administration of a test dose, seems to be a promising tool for determining actual metabolic capacities. Although it is a well-established experimental approach, phenotyping has not yet found its way into clinical practice. Main reasons for this are lack of validation for many probes and assays used, complicated procedures, invasiveness, semi-quantitative test results, non-compliance on behalf of the subjects tested, high costs, and lack of prospective clinical studies to assess the benefit of phenotyping for patients. Problems and perspectives of phenotyping are exemplified for the cytochrome P-450 enzymes CYP1A2 and CYP3A4, two major human drug-metabolizing enzymes. PMID:10667830

Zaigler, M; Tantcheva-Poór, I; Fuhr, U

2000-01-01

116

Intermittent hypoxic resistance training: is metabolic stress the key moderator?  

PubMed

Traditionally, researchers and practitioners have manipulated acute resistance exercise variables to elicit the desired responses to training. However, recent research indicates that altering the muscular environment during resistance training, namely by implementing a hypoxic stimulus, can augment muscle hypertrophy and strength. Intermittent hypoxic resistance training (IHRT), whereby participants inspire hypoxic air during resistance training, has been previously demonstrated to increase muscle cross-sectional area and maximum strength by significantly greater amounts than the equivalent training in normoxia. However, some recent evidence has provided conflicting results, reporting that the use of systemic hypoxia during resistance training provided no added benefit. While the definitive mechanisms that may augment muscular responses to IHRT are not yet fully understood, an increased metabolic stress is thought to be important for moderating many downstream processes related to hypertrophy. It is likely that methodological differences between conflicting IHRT studies have resulted in different degrees of metabolic stress during training, particularly when considering the inter-set recovery intervals used. Given that the most fundamental physiological stresses resulting from hypoxia are disturbances to oxidative metabolism, it becomes apparent that resistance training may only benefit from additional hypoxia if the exercise is structured to elicit a strong metabolic response. We hypothesize that for IHRT to be more effective in producing muscular hypertrophy and increasing strength than the equivalent normoxic training, exercise should be performed with relatively brief inter-set recovery periods, with the aim of providing a potent metabolic stimulus to enhance anabolic responses. PMID:25547781

Scott, Brendan R; Slattery, Katie M; Dascombe, Ben J

2015-02-01

117

Experiment K-6-21. Effect of microgravity on 1) metabolic enzymes of type 1 and type 2 muscle fibers and on 2) metabolic enzymes, neutransmitter amino acids, and neurotransmitter associated enzymes in motor and somatosensory cerebral cortex. Part 1: Metabolic enzymes of individual muscle fibers; part 2: metabolic enzymes of hippocampus and spinal cord  

NASA Technical Reports Server (NTRS)

The individual fibers of any individual muscle vary greatly in enzyme composition, a fact which is obscured when enzyme levels of a whole muscle are measured. The purpose of this study was therefore to assess the changes due to weightless on the enzyme patterns composed by the individual fibers within the flight muscles. In spite of the limitation in numbers of muscles examined, it is apparent that: (1) that the size of individual fibers (i.e., their dry weight) was reduced about a third, (2) that this loss in dry mass was accompanied by changes in the eight enzymes studied, and (3) that these changes were different for the two muscles, and different for the two enzyme groups. In the soleus muscle the absolute amounts of the three enzymes of oxidative metabolism decreased about in proportion to the dry weight loss, so that their concentration in the atrophic fibers was almost unchanged. In contrast, there was little loss among the four enzymes of glycogenolysis - glycolysis so that their concentrations were substantially increased in the atrophic fibers. In the TA muscle, these seven enzymes were affected in just the opposite direction. There appeared to be no absolute loss among the oxidative enzymes, whereas the glycogenolytic enzymes were reduced by nearly half, so that the concentrations of the first metabolic group were increased within the atrophic fibers and the concentrations of the second group were only marginally decreased. The behavior of hexokinase was exceptional in that it did not decrease in absolute terms in either type of muscle and probably increased as much as 50 percent in soleus. Thus, their was a large increase in concentration of this enzyme in the atrophied fibers of both muscles. Another clear-cut finding was the large increase in the range of activities of the glycolytic enzymes among individual fibers of TA muscles. This was due to the emergence of TA fibers with activities for enzymes of this group extending down to levels as low as those found in control soleus muscles. It would be interesting to know if this represents a transition stage, and whether with prolonged weightlessness most of the fibers would be transformed into a low glycogenolytic type.

Lowry, O.; Mcdougal, D., Jr.; Nemeth, Patti M.; Maggie, M.-Y. Chi; Pusateri, M.; Carter, J.; Manchester, J.; Norris, Beverly; Krasnov, I.

1990-01-01

118

Enzyme II Glc contributes to trehalose metabolism in Bacillus subtilis  

Microsoft Academic Search

The tre operon of Bacillus subtilis is regulated at the transcriptional level by the Tre repressor (TreR) and by carbon catabolite repression. The molecular inducer of TreR is trehalose-6-phosphate, which is generated by transport of trehalose across the membrane by a specific Enzyme IITre (TreP) and simultaneous phosphorylation. It has been shown that TreP lacks the Enzyme IIA domain which

Michael K Dahl

1997-01-01

119

PGC-1{alpha}: a key regulator of energy metabolism  

NSDL National Science Digital Library

Peroxisome proliferator-activated receptor-{gamma} coactivator (PGC)-1{alpha} is a member of a family of transcription coactivators that plays a central role in the regulation of cellular energy metabolism. This makes it an inviting target for pharmacological intervention in the treatment of obesity and Type 2 diabetes.

Huiyun Liang (University of Texas Health Science Center Department of Cellular and Structural Biology, Barshop Institute for Longevity and Aging Studies)

2006-12-01

120

Heterologous Expression and Maturation of an NADP-Dependent [NiFe]-Hydrogenase: A Key Enzyme in Biofuel Production  

PubMed Central

Hydrogen gas is a major biofuel and is metabolized by a wide range of microorganisms. Microbial hydrogen production is catalyzed by hydrogenase, an extremely complex, air-sensitive enzyme that utilizes a binuclear nickel-iron [NiFe] catalytic site. Production and engineering of recombinant [NiFe]-hydrogenases in a genetically-tractable organism, as with metalloprotein complexes in general, has met with limited success due to the elaborate maturation process that is required, primarily in the absence of oxygen, to assemble the catalytic center and functional enzyme. We report here the successful production in Escherichia coli of the recombinant form of a cytoplasmic, NADP-dependent hydrogenase from Pyrococcus furiosus, an anaerobic hyperthermophile. This was achieved using novel expression vectors for the co-expression of thirteen P. furiosus genes (four structural genes encoding the hydrogenase and nine encoding maturation proteins). Remarkably, the native E. coli maturation machinery will also generate a functional hydrogenase when provided with only the genes encoding the hydrogenase subunits and a single protease from P. furiosus. Another novel feature is that their expression was induced by anaerobic conditions, whereby E. coli was grown aerobically and production of recombinant hydrogenase was achieved by simply changing the gas feed from air to an inert gas (N2). The recombinant enzyme was purified and shown to be functionally similar to the native enzyme purified from P. furiosus. The methodology to generate this key hydrogen-producing enzyme has dramatic implications for the production of hydrogen and NADPH as vehicles for energy storage and transport, for engineering hydrogenase to optimize production and catalysis, as well as for the general production of complex, oxygen-sensitive metalloproteins. PMID:20463892

Jenney, Francis E.; McTernan, Patrick M.; Adams, Michael W. W.

2010-01-01

121

Effects of boron deficiency on major metabolites, key enzymes and gas exchange in leaves and roots of Citrus sinensis seedlings.  

PubMed

Boron (B) deficiency is a widespread problem in many crops, including Citrus. The effects of B-deficiency on gas exchange, carbohydrates, organic acids, amino acids, total soluble proteins and phenolics, and the activities of key enzymes involved in organic acid and amino acid metabolism in 'Xuegan' [Citrus sinensis (L.) Osbeck] leaves and roots were investigated. Boron-deficient leaves displayed excessive accumulation of nonstructural carbohydrates and much lower CO2 assimilation, demonstrating feedback inhibition of photosynthesis. Dark respiration, concentrations of most organic acids [i.e., malate, citrate, oxaloacetate (OAA), pyruvate and phosphoenolpyruvate] and activities of enzymes [i.e., phosphoenolpyruvate carboxylase (PEPC), NAD-malate dehydrogenase, NAD-malic enzyme (NAD-ME), NADP-ME, pyruvate kinase (PK), phosphoenolpyruvate phosphatase (PEPP), citrate synthase (CS), aconitase (ACO), NADP-isocitrate dehydrogenase (NADP-IDH) and hexokinase] involved in glycolysis, the tricarboxylic acid (TCA) cycle and the anapleurotic reaction were higher in B-deficient leaves than in controls. Also, total free amino acid (TFAA) concentration and related enzyme [i.e., NADH-dependent glutamate 2-oxoglutarate aminotransferase (NADH-GOGAT) and glutamate OAA transaminase (GOT)] activities were enhanced in B-deficient leaves. By contrast, respiration, concentrations of nonstructural carbohydrates and three organic acids (malate, citrate and pyruvate), and activities of most enzymes [i.e., PEPC, NADP-ME, PK, PEPP, CS, ACO, NAD-isocitrate dehydrogenase, NADP-IDH and hexokinase] involved in glycolysis, the TCA cycle and the anapleurotic reaction, as well as concentration of TFAA and activities of related enzymes (i.e., nitrate reductase, NADH-GOGAT, glutamate pyruvate transaminase and glutamine synthetase) were lower in B-deficient roots than in controls. Interestingly, leaf and root concentration of total phenolics increased, whereas that of total soluble protein decreased, in response to B-deficiency. In conclusion, respiration, organic acid (i.e., glycolysis and the TCA cycle) metabolism, the anapleurotic pathway and amino acid biosynthesis were upregulated in B-deficient leaves with excessive accumulation of carbohydrates to 'consume' the excessive carbon available, but downregulated in B-deficient roots with less accumulation of carbohydrates to maintain the net carbon balance. PMID:24957048

Lu, Yi-Bin; Yang, Lin-Tong; Li, Yan; Xu, Jing; Liao, Tian-Tai; Chen, Yan-Bin; Chen, Li-Song

2014-06-01

122

Metabolic and Bactericidal Effects of Targeted Suppression of NadD and NadE Enzymes in Mycobacteria  

PubMed Central

ABSTRACT Mycobacterium tuberculosis remains a major cause of death due to the lack of treatment accessibility, HIV coinfection, and drug resistance. Development of new drugs targeting previously unexplored pathways is essential to shorten treatment time and eliminate persistent M. tuberculosis. A promising biochemical pathway which may be targeted to kill both replicating and nonreplicating M. tuberculosis is the biosynthesis of NAD(H), an essential cofactor in multiple reactions crucial for respiration, redox balance, and biosynthesis of major building blocks. NaMN adenylyltransferase (NadD) and NAD synthetase (NadE), the key enzymes of NAD biosynthesis, were selected as promising candidate drug targets for M. tuberculosis. Here we report for the first time kinetic characterization of the recombinant purified NadD enzyme, setting the stage for its structural analysis and inhibitor development. A protein knockdown approach was applied to validate bothNadD and NadE as target enzymes. Induced degradation of either target enzyme showed a strong bactericidal effect which coincided with anticipated changes in relative levels of NaMN and NaAD intermediates (substrates of NadD and NadE, respectively) and ultimate depletion of the NAD(H) pool. A metabolic catastrophe predicted as a likely result of NAD(H) deprivation of cellular metabolism was confirmed by 13C biosynthetic labeling followed by gas chromatography-mass spectrometry (GC-MS) analysis. A sharp suppression of metabolic flux was observed in multiple NAD(P)(H)-dependent pathways, including synthesis of many amino acids (serine, proline, aromatic amino acids) and fatty acids. Overall, these results provide strong validation of the essential NAD biosynthetic enzymes, NadD and NadE, as antimycobacterial drug targets. PMID:24549842

Rodionova, Irina A.; Schuster, Brian M.; Guinn, Kristine M.; Sorci, Leonardo; Scott, David A.; Li, Xiaoqing; Kheterpal, Indu; Shoen, Carolyn; Cynamon, Michael; Locher, Christopher; Rubin, Eric J.; Osterman, Andrei L.

2014-01-01

123

Oral cancer cells may rewire alternative metabolic pathways to survive from siRNA silencing of metabolic enzymes  

PubMed Central

Background Cancer cells may undergo metabolic adaptations that support their growth as well as drug resistance properties. The purpose of this study is to test if oral cancer cells can overcome the metabolic defects introduced by using small interfering RNA (siRNA) to knock down their expression of important metabolic enzymes. Methods UM1 and UM2 oral cancer cells were transfected with siRNA to transketolase (TKT) or siRNA to adenylate kinase (AK2), and Western blotting was used to confirm the knockdown. Cellular uptake of glucose and glutamine and production of lactate were compared between the cancer cells with either TKT or AK2 knockdown and those transfected with control siRNA. Statistical analysis was performed with student T-test. Results Despite the defect in the pentose phosphate pathway caused by siRNA knockdown of TKT, the survived UM1 or UM2 cells utilized more glucose and glutamine and secreted a significantly higher amount of lactate than the cells transferred with control siRNA. We also demonstrated that siRNA knockdown of AK2 constrained the proliferation of UM1 and UM2 cells but similarly led to an increased uptake of glucose/glutamine and production of lactate by the UM1 or UM2 cells survived from siRNA silencing of AK2. Conclusions Our results indicate that the metabolic defects introduced by siRNA silencing of metabolic enzymes TKT or AK2 may be compensated by alternative feedback metabolic mechanisms, suggesting that cancer cells may overcome single defective pathways through secondary metabolic network adaptations. The highly robust nature of oral cancer cell metabolism implies that a systematic medical approach targeting multiple metabolic pathways may be needed to accomplish the continued improvement of cancer treatment. PMID:24666435

2014-01-01

124

Thyroid regulation of resting metabolic rate and intermediary metabolic enzymes in a lizard (Sceloporus occidentalis).  

PubMed

This study investigates the effects of physiological increments in plasma thyroxine (T4) at three levels of biological organization in thyroid-intact and thyroidectomized captive western fence lizards, Sceloporus occidentalis. Two doses of T4-loaded pellets elevated plasma T4 in thyroid-intact lizards from 4.8 +/- 0.47 to 10.7 +/- 2.25 and 20.4 +/- 5.77 ng/ml (mean +/- SE). Surgical thyroidectomy reduced T4 to 1.8 +/- 0.23 ng/ml, and subsequent T4 pellet implantation raised T4 to 14.8 +/- 4.30 ng/ml. Minimal resting metabolic rate (= standard metabolic rate; SMR), a common organismal metric of thyroid perturbation, was reduced 31% (P less than 0.0001) by thyroidectomy and was restored by T4 replacement but was not stimulated by T4 supplementation in thyroid-intact lizards. In T4-replaced, thyroidectomized lizards, SMR was significantly correlated with plasma T4 (r2 = 0.626, P = 0.003, n = 11). At the organ level, liver mass was not changed by any treatment; heart mass was decreased by thyroid deficiency and restored by T4 replacement. At the molecular level, citrate synthase activity was significantly reduced by thyroidectomy and was returned to control levels by T4 replacement in liver and skeletal muscle (gastrocnemius) but was not changed in cardiac muscle. Citrate synthase was not affected in any tissue by T4 supplementation in thyroid-intact lizards. Pyruvate kinase activity was not affected by any of the treatments in any of the tissues. Cytosolic alpha-glycerophosphate dehydrogenase was significantly reduced in liver by all treatments and in skeletal muscle by T4 replacement after thyroidectomy. These results indicate that SMR and cardiac muscle mass in lizards are dependent on normal thyroid function and are expressed maximally in euthyroid animals. The stimulatory effect of T4 on SMR in thyroid-intact lizards, which has been reported previously by several investigators, is a nonphysiological response to pharmacological T4 levels, at least in these captive lizards. Molecular responses are tissue and enzyme dependent and cannot be generalized. Pellet implantation is an effective means of inducing physiological increments in plasma T4 and should replace previously used injection protocols. This new method can be used in capture-recapture experiments involving field-active lizards. PMID:2295423

John-Alder, H B

1990-01-01

125

Evolution of the key alkaloid enzyme putrescine N-methyltransferase from spermidine synthase  

PubMed Central

Putrescine N-methyltransferases (PMTs) are the first specific enzymes of the biosynthesis of nicotine and tropane alkaloids. PMTs transfer a methyl group onto the diamine putrescine from S-adenosyl-l-methionine (SAM) as coenzyme. PMT proteins have presumably evolved from spermidine synthases (SPDSs), which are ubiquitous enzymes of polyamine metabolism. SPDSs use decarboxylated SAM as coenzyme to transfer an aminopropyl group onto putrescine. In an attempt to identify possible and necessary steps in the evolution of PMT from SPDS, homology based modeling of Datura stramonium SPDS1 and PMT was employed to gain deeper insight in the preferred binding positions and conformations of the substrate and the alternative coenzymes. Based on predictions of amino acids responsible for the change of enzyme specificities, sites of mutagenesis were derived. PMT activity was generated in D. stramonium SPDS1 after few amino acid exchanges. Concordantly, Arabidopsis thaliana SPDS1 was mutated and yielded enzymes with both, PMT and SPDS activities. Kinetic parameters were measured for enzymatic characterization. The switch from aminopropyl to methyl transfer depends on conformational changes of the methionine part of the coenzyme in the binding cavity of the enzyme. The rapid generation of PMT activity in SPDS proteins and the wide-spread occurrence of putative products of N-methylputrescine suggest that PMT activity is present frequently in the plant kingdom. PMID:23908659

Junker, Anne; Fischer, Juliane; Sichhart, Yvonne; Brandt, Wolfgang; Dräger, Birgit

2013-01-01

126

Metaproteomic Analysis of a Chemosynthetic Hydrothermal Vent Community Reveals Insights into Key-Metabolic Processes  

NASA Astrophysics Data System (ADS)

In 2005 researchers at the Centre for Geobiology, University of Bergen, Norway, discovered two active vent fields at the southwestern Mohns Ridge in the Norwegian-Greenland Sea. The fields harbours both low-temperature iron deposits and high-temperature white smoker vents. Distinct microbial mats were abundantly present and located in close vicinity to the hydrothermal vent sites. Characteristics of the mat environment were steep physical and chemical gradients with temperatures ranging from 10°C in the top layer to 90°C at 10 cm bsf and high concentrations of hydrogen sulfide and methane. The work presented here focus on the In situ community activities, and is part of an integrated strategy combining metagenomics, metatranscriptomics and metaproteomics to in-depth characterise these newly discovered hydrothermal vent communities. Extracted proteins were separated via SDS-PAGE. Peptides extracted after In-gel tryptic digest was injected into an Ultimate 3000 nanoLC system connected to a linear quadropole ion trap-orbitrap (LTQ-Orbitrap XL) mass spectrometer equipped with a nanoelectrospray ion source. A custom database of open reading frames (ORFs) from the combined metatranscriptome and metagenome datasets was implemented and searched against using Mascot 2.2; the IRMa tool box [1] was used in peptide validation. Validated ORFs were subjected to a Blastp search against Refseq with an E-value cut-off of 0.001. A total of 1097 proteins with ? 2 peptides were identified of which 921 gave a hit against Refseq, containing 519 unique proteins. Key enzymes of the sulfur oxidation pathway (sox) were found, which were taxonomically affiliated to Epsilonproteobacteria. In addition, this group actively expressed hydrogenases and membrane proteins involved in aerobic and anaerobic respiratory chains. Enzymes of dissimilatory sulfate-reduction (APS-reductase, AprAB and DsrA2) were found with closest hit to members of the Deltaproteobacteria. These findings indicate an internal sulfur cycle within the community. The community contained expressed enzymes of a variety of carbon metabolism pathways. Key enzymes of the reverse TCA cycle for fixation of CO2 and the Wood-Ljungdahl pathway for oxidation of acetyl-CoA and / or the fixation of CO2 were found. Key enzymes of aerobic and anaerobic methane-oxidation pathways were identified as well, namely particulate methane monooxygenase and methyl-Coenzyme M reductase. Various house-keeping gene-products, like cold- and heat shock proteins as well as ribosomal proteins and ATP synthases were identified. This approach has a future potential of broadening our understanding of environmental complexity and regulation in response to geochemical constraints. [1] Dupierris, V., Masselon, C., Court, M., Kieffer-Jaquinod, S., and Bruley, C. (2009) A toolbox for validation of mass spectrometry peptides identification and generation of database: IRMa. Bioinformatics 25, 1980-1981.

Steen, I.; Stokke, R.; Lanzen, A.; Pedersen, R.; Řvreĺs, L.; Urich, T.

2010-12-01

127

Relevance of induction of human drug-metabolizing enzymes: pharmacological and toxicological implications  

PubMed Central

1Human drug-metabolizing systems can be induced, or activated, by a large number of exogenous agents including drugs, alcohol, components in the diet and cigarette smoke, as well as by endogenous factors. 2Such perturbation of enzyme activity undoubtedly contributes to both intra- and inter-individual variation both with respect to the rate and route of metabolism for a particular drug. Induction may, in theory, either attenuate the pharmacological response or exacerbate the toxicity of a particular drug, or both. 3The clinical impact of enzyme induction will depend upon the number of different enzyme isoforms affected and the magnitude of the inductive response within an individual, and also on the therapeutic indices of the affected substrates. 4The toxicological implications will be determined either by any change in the route of metabolism, or by a disturbance of the balance between activation and detoxication processes, which may be isozyme selective. PMID:8799511

PARK, B. K.; KITTERINGHAM, N. R.; PIRMOHAMED, M.; TUCKER, G. T.

1996-01-01

128

Cyclophosphamide- metabolizing enzyme polymorphisms and survival outcomes after adjuvant chemotherapy for node-positive breast cancer: a retrospective cohort study  

E-print Network

chemotherapy for node- positive breast cancer: a retrospective cohort study Breast Cancer ResearchResearch article Open Access Cyclophosphamide- metabolizing enzyme polymorphisms and survival outcomes after adjuvant chemotherapy

2010-01-01

129

A gold-containing drug against parasitic polyamine metabolism: the X-ray structure of trypanothione reductase from Leishmania infantum in complex with auranofin reveals a dual mechanism of enzyme inhibition  

Microsoft Academic Search

Auranofin is a gold(I)-containing drug in clinical use as an antiarthritic agent. Recent studies showed that auranofin manifests\\u000a interesting antiparasitic actions very likely arising from inhibition of parasitic enzymes involved in the control of the\\u000a redox metabolism. Trypanothione reductase is a key enzyme of Leishmania infantum polyamine-dependent redox metabolism, and a validated target for antileishmanial drugs. As trypanothione reductase contains

Andrea Ilari; Paola Baiocco; Luigi Messori; Annarita Fiorillo; Alberto Boffi; Marina Gramiccia; Trentina Di Muccio; Gianni Colotti

130

Survey of Human Oxidoreductases and Cytochrome P450 Enzymes Involved in the Metabolism of Chemicals.  

PubMed

Analyzing the literature resources used in our previous reports, we calculated the fractions of the oxidoreductase enzymes FMO (microsomal flavin-containing monooxygenase), AKR (aldo-keto reductase), MAO (monoamine oxidase), and cytochrome P450 participating in metabolic reactions. The calculations show that the fractions of P450s involved in metabolism of all chemicals (general chemicals, natural and physiological compounds, and drugs) are rather consistent in the findings that > 90% of enzymatic reactions are catalyzed by P450s. Regarding drug metabolism, three-fourths of the human P450 reactions can be accounted for by a set of five P450s: 1A2, 2C9, 2C19, 2D6, and 3A4, and the largest fraction of the P450 reactions is catalyzed by P450 3A enzymes. P450 3A4 participation in metabolic reactions of drugs varied from 13% for general chemicals to 27% for drugs. PMID:25485457

Rendic, Slobodan Petar; Guengerich, F Peter

2014-12-01

131

Metabolic engineering is key to a sustainable chemical industry.  

PubMed

The depletion of fossil fuel stocks will prohibit their use as the main feedstock of future industrial processes. Biocatalysis is being increasingly used to reduce fossil fuel reliance and to improve the sustainability, efficiency and cost of chemical production. Even with their current small market share, biocatalyzed processes already generate approximately US$50 billion and it has been estimated that they could be used to produce up to 20% of fine chemicals by 2020. Until the advent of molecular biological technologies, the compounds that were readily accessible from renewable biomass were restricted to naturally-occurring metabolites. However, metabolic engineering has considerably broadened the range of compounds now accessible, providing access to compounds that cannot be otherwise reliably sourced, as well as replacing established chemical processes. This review presents the case for continued efforts to promote the adoption of biocatalyzed processes, highlighting successful examples of industrial chemical production from biomass and/or via biocatalyzed processes. A selection of emerging technologies that may further extend the potential and sustainability of biocatalysis are also presented. As the field matures, metabolic engineering will be increasingly crucial in maintaining our quality of life into a future where our current resources and feedstocks cannot be relied upon. PMID:21666928

Murphy, Annabel C

2011-08-01

132

Biotransformation of anthelmintics and the activity of drug-metabolizing enzymes in the tapeworm Moniezia expansa.  

PubMed

SUMMARY The sheep tapeworm Moniezia expansa is very common parasite, which affects ruminants such as sheep, goats as well as other species. The benzimidazole anthelmintics albendazole (ABZ), flubendazole (FLU) and mebendazole (MBZ) are often used to treat the infection. The drug-metabolizing enzymes of helminths may alter the potency of anthelmintic treatment. The aim of our study was to assess the activity of the main drug-metabolizing enzymes and evaluate the metabolism of selected anthelmintics (ABZ, MBZ and FLU) in M. expansa. Activities of biotransformation enzymes were determined in subcellular fractions. Metabolites of the anthelmintics were detected and identified using high performance liquid chromatography/ultra-violet/VIS/fluorescence or ultra-high performance liquid chromatography/mass spectrometry. Reduction of MBZ, FLU and oxidation of ABZ were proved as well as activities of various metabolizing enzymes. Despite the fact that the conjugation enzymes glutathione S-transferase, UDP-glucuronosyl transferase and UDP-glucosyl transferase were active in vitro, no conjugated metabolites of anthelmintics were identified either ex vivo or in vitro. The obtained results indicate that sheep tapeworm is able to deactivate the administered anthelmintics, and thus protects itself against their action. PMID:25373326

Prchal, Lukáš; Bártíková, Hana; Be?anová, Aneta; Jirásko, Robert; Vok?ál, Ivan; Stuchlíková, Lucie; Skálová, Lenka; Kubí?ek, Vladimír; Lamka, Ji?í; Trejtnar, František; Szotáková, Barbora

2014-11-01

133

Allelic Variants of Drug Metabolizing Enzymes as Risk Factors in Psoriasis  

Microsoft Academic Search

The onset or exacerbation of psoriasis, a T-cell-dependent skin disease with autoimmune features, can be triggered by drugs such as antimalarials and beta-blockers. Xenobiotics may also play a role in idiopathic psoriasis. It has been hypothesized that different metabolic efficiencies caused by variant alleles of xenobiotic metabolizing enzymes could lead to the accumulation of xenobiotics or their reactive metabolites in

Dagmar Richter-Hintz; Ricarda Their; Swantje Steinwachs; Sven Kronenberg; Ellen Fritsche; Bernd Sachs; Marty Wulferink; Torsten Tonn; Charlotte Esser

2003-01-01

134

Altitude-related changes in activities of carbon metabolism enzymes in Rumex nepalensis  

Microsoft Academic Search

Activities of some enzymes related to carbon metabolism were studied in different ecotypes of Rumex nepalensis growing at 1 300, 2 250, and 3 250 m above mean sea level. Activities of ribulose-1,5-bisphosphate carboxylase\\/oxygenase,\\u000a phosphoenolpyruvate carboxylase, aspartate aminotransferase, and glutamine synthetase increased with altitude, whereas activities of\\u000a malate dehydrogenase, NAD-malic enzyme, and citrate synthase did not show a significant difference

N. Kumar; S. Kumar Vats; S. Kumar; P. S. Ahuja

2008-01-01

135

Hydrogenosome Metabolism Is the Key Target for Antiparasitic Activity of Resveratrol against Trichomonas vaginalis  

PubMed Central

Metronidazole (MDZ) and related 5-nitroimidazoles are the recommended drugs for treatment of trichomoniasis, a sexually transmitted disease caused by the protozoan parasite Trichomonas vaginalis. However, novel treatment options are needed, as recent reports have claimed resistance to these drugs in T. vaginalis isolates. In this study, we analyzed for the first time the in vitro effects of the natural polyphenol resveratrol (RESV) on T. vaginalis. At concentrations of between 25 and 100 ?M, RESV inhibited the in vitro growth of T. vaginalis trophozoites; doses of 25 ?M exerted a cytostatic effect, and higher doses exerted a cytotoxic effect. At these concentrations, RESV caused inhibition of the specific activity of a 120-kDa [Fe]-hydrogenase (Tvhyd). RESV did not affect Tvhyd gene expression and upregulated pyruvate-ferredoxin oxidoreductase (a hydrogenosomal enzyme) gene expression only at a high dose (100 ?M). At doses of 50 to 100 ?M, RESV also caused overexpression of heat shock protein 70 (Hsp70), a protective protein found in the hydrogenosome of T. vaginalis. The results demonstrate the potential of RESV as an antiparasitic treatment for trichomoniasis and suggest that the mechanism of action involves induction of hydrogenosomal dysfunction. In view of the results, we propose hydrogenosomal metabolism as a key target in the design of novel antiparasitic drugs. PMID:23478970

Mallo, Natalia; Lamas, Jesús

2013-01-01

136

In vivo co-localization of enzymes on RNA scaffolds increases metabolic production in a geometrically dependent manner.  

PubMed

Co-localization of biochemical processes plays a key role in the directional control of metabolic fluxes toward specific products in cells. Here, we employ in vivo scaffolds made of RNA that can bind engineered proteins fused to specific RNA binding domains. This allows proteins to be co-localized on RNA scaffolds inside living Escherichia coli. We assembled a library of eight aptamers and corresponding RNA binding domains fused to partial fragments of fluorescent proteins. New scaffold designs could co-localize split green fluorescent protein fragments to produce activity as measured by cell-based fluorescence. The scaffolds consisted of either single bivalent RNAs or RNAs designed to polymerize in one or two dimensions. The new scaffolds were used to increase metabolic output from a two-enzyme pentadecane production pathway that contains a fatty aldehyde intermediate, as well as three and four enzymes in the succinate production pathway. Pentadecane synthesis depended on the geometry of enzymes on the scaffold, as determined through systematic reorientation of the acyl-ACP reductase fusion by rotation via addition of base pairs to its cognate RNA aptamer. Together, these data suggest that intra-cellular scaffolding of enzymatic reactions may enhance the direct channeling of a variety of substrates. PMID:25034694

Sachdeva, Gairik; Garg, Abhishek; Godding, David; Way, Jeffrey C; Silver, Pamela A

2014-08-01

137

In vivo co-localization of enzymes on RNA scaffolds increases metabolic production in a geometrically dependent manner  

PubMed Central

Co-localization of biochemical processes plays a key role in the directional control of metabolic fluxes toward specific products in cells. Here, we employ in vivo scaffolds made of RNA that can bind engineered proteins fused to specific RNA binding domains. This allows proteins to be co-localized on RNA scaffolds inside living Escherichia coli. We assembled a library of eight aptamers and corresponding RNA binding domains fused to partial fragments of fluorescent proteins. New scaffold designs could co-localize split green fluorescent protein fragments to produce activity as measured by cell-based fluorescence. The scaffolds consisted of either single bivalent RNAs or RNAs designed to polymerize in one or two dimensions. The new scaffolds were used to increase metabolic output from a two-enzyme pentadecane production pathway that contains a fatty aldehyde intermediate, as well as three and four enzymes in the succinate production pathway. Pentadecane synthesis depended on the geometry of enzymes on the scaffold, as determined through systematic reorientation of the acyl-ACP reductase fusion by rotation via addition of base pairs to its cognate RNA aptamer. Together, these data suggest that intra-cellular scaffolding of enzymatic reactions may enhance the direct channeling of a variety of substrates. PMID:25034694

Sachdeva, Gairik; Garg, Abhishek; Godding, David; Way, Jeffrey C.; Silver, Pamela A.

2014-01-01

138

Gene Expression of Glutamate Metabolizing Enzymes in the Hippocampal Formation in Human Temporal Lobe Epilepsy  

PubMed Central

Summary PURPOSE Increased interictal concentrations of extracellular hippocampal glutamate have been implicated in the pathophysiology of temporal lobe epilepsy (TLE) in humans. Recent studies suggest that perturbations of the glutamate metabolizing enzymes glutamine synthetase (GS) and phosphate activated glutaminase (PAG) may underlie the glutamate excess in TLE. However, the molecular mechanism of the enzyme perturbations remains unclear. A better understanding of the regulatory mechanisms of GS and PAG could facilitate the discovery of novel therapeutics for TLE. METHODS We used in situ hybridization on histological sections to assess the distribution and quantity of mRNA for GS and PAG in subfields of hippocampal formations from: (a) patients with TLE and concomitant hippocampal sclerosis, (b) patients with TLE and no hippocampal sclerosis, and (c) non-epilepsy autopsy subjects. KEY FINDINGS GS mRNA was increased by approximately 50% in the CA3 in TLE patients without hippocampal sclerosis vs. in TLE patients with sclerosis and in non-epilepsy subjects. PAG mRNA was increased by more than 100% in the subiculum in both TLE patient categories vs. in non-epilepsy subjects. PAG mRNA was also increased in the CA1, CA2, CA3 and dentate hilus in TLE without hippocampal sclerosis vs. in TLE with sclerosis. Finally, PAG mRNA was increased in the dentate gyrus in TLE with sclerosis vs. in non-epilepsy subjects, and also increased in the hilus in TLE without sclerosis vs. in TLE with sclerosis. SIGNIFICANCE These findings demonstrate complex changes in the expression of mRNAs for GS and PAG in the hippocampal formation in TLE, and raise the possibility that both transcriptional and post-transcriptional mechanisms may underlie the regulation of GS and PAG proteins in the epileptic brain. PMID:23384343

Eid, Tore; Lee, Tih-Shih W; Wang, Yue; Peréz, Edgar; Drummond, Jana; Lauritzen, Fredrik; Bergersen, Linda H; Woodruff, James H Meador; Spencer, Dennis D; de Lanerolle, Nihal C; McCullumsmith, Robert E

2012-01-01

139

2D SMARTCyp Reactivity-Based Site of Metabolism Prediction for Major Drug-Metabolizing Cytochrome P450 Enzymes  

E-print Network

Information ABSTRACT: Cytochrome P450 (CYP) 3A4, 2D6, 2C9, 2C19, and 1A2 are the most important drug of CYP3A4 and CYP2D6 substrates with an accuracy that rivals the best and more computationally demanding-drug interactions and metabolism-dependent toxicity issues.3 Among the CYP enzymes, CYP3A4, 2D6, 2C9, 2C19, and 1A2

140

Functional genomics and SNP analysis of human genes encoding proline metabolic enzymes  

PubMed Central

Proline metabolism in mammals involves two other amino acids, glutamate and ornithine, and five enzymatic activities, ?1-pyrroline-5-carboxylate (P5C) reductase (P5CR), proline oxidase, P5C dehydrogenase, P5C synthase and ornithine-?-aminotransferase (OAT). With the exception of OAT, which catalyzes a reversible reaction, the other 4 enzymes are unidirectional, suggesting that proline metabolism is purpose-driven, tightly regulated, and compartmentalized. In addition, this tri-amino-acid system also links with three other pivotal metabolic systems, namely the TCA cycle, urea cycle, and pentose phosphate pathway. Abnormalities in proline metabolism are relevant in several diseases: six monogenic inborn errors involving metabolism and/or transport of proline and its immediate metabolites have been described. Recent advances in the Human Genome Project, in silico database mining techniques, and research in dissecting the molecular basis of proline metabolism prompted us to utilize functional genomic approaches to analyze human genes which encode proline metabolic enzymes in the context of gene structure, regulation of gene expression, mRNA variants, protein isoforms, and single nucleotide polymorphisms. PMID:18506409

Williams, D. Bart; Zhaorigetu, Siqin; Khalil, Shadi; Wan, Guanghua; Valle, David

2009-01-01

141

Comparative investigation of the xenobiotic metabolizing arylamine N-acetyltransferase enzyme family among fungi  

Technology Transfer Automated Retrieval System (TEKTRAN)

Arylamine N-acetyltransferases (NATs) are xenobiotic metabolizing enzymes well-characterized in several bacteria and higher eukaryotes. The role of NATs in fungal biology has only recently been investigated. The NAT1 gene of Gibberella moniliformis was the first NAT cloned and characterized from fun...

142

Pharmacogenomics of drug-metabolizing enzymes and drug transporters in chemotherapy.  

PubMed

There is wide variability in the response of individuals to standard doses of drug therapy. This is an important problem in clinical practice, where it can lead to therapeutic failures or adverse drug events. Polymorphisms in genes coding for metabolizing enzymes and drug transporters can affect drug efficacy and toxicity. Pharmacogenomics aims to identify individuals predisposed to high risk of toxicity and low response from standard doses of anticancer drugs. This chapter focuses on the clinical significance of polymorphisms in drug-metabolizing enzymes and drug transporters in influencing efficacy and toxicity of anticancer therapy. The most important examples to demonstrate the influence of pharmacogenomics on anticancer therapy are thiopurine methyltransferase (TPMT), UGT (uridine diphosphate glucuronosyltransferase) 1A1*28, and DPD (dihydropyrimidine dehydrogenase) *2A, respectively, for 6-mercaptopurine, irinotecan, and 5-fluorouracil therapy. However, in most other anticancer therapies no clear association has been found for polymorphisms in drug-metabolizing enzymes and drug transporters and pharmacokinetics or pharmacodynamics of anticancer drugs. Evaluation of different regimens and tumor types showed that polymorphisms can have different, sometimes even contradictory, pharmacokinetic and pharmacodynamic effects in different tumors in response to different drugs. The clinical application of pharmacogenomics in cancer treatment therefore requires more detailed information regarding the different polymorphisms in drug-metabolizing enzymes and drug transporters. A greater understanding of complexities in pharmacogenomics is needed before individualized therapy can be applied on a routine basis. PMID:18370231

Bosch, Tessa M

2008-01-01

143

Acetyl-coenzyme A carboxylase: crucial metabolic enzyme and attractive target for drug discovery  

E-print Network

Review Acetyl-coenzyme A carboxylase: crucial metabolic enzyme and attractive target for drug revision 2 April 2005; accepted 19 April 2005 Online First 17 June 2005 Abstract. Acetyl-coenzyme. Introduction Acetyl-coenzyme A carboxylase (ACC) (EC 6.4.1.2) cat- alyzes the carboxylation of acetyl

Tong, Liang

144

Hepatic Xenobiotic Metabolizing Enzyme Gene Expression Through the Life Stages of the Mouse  

EPA Science Inventory

BACKGROUND: Differences in responses to environmental chemicals and drugs between life stages are likely due in part to differences in the expression of xenobiotic metabolizing enzymes and transporters (XMETs). No comprehensive analysis of the mRNA expression of XMETs has been ca...

145

Coordinated Changes in Xenobiotic Metabolizing Enzyme Gene Expression in Aging Male Rats  

EPA Science Inventory

In order to gain better insight on aging and susceptibility, we characterized the expression of xenobiotic metabolizing enzymes (XMEs) from the livers of rats to evaluate the change in capacity to respond to xenobiotics across the adult lifespan. Gene expression profiles for XMEs...

146

IDENTIFICATION OF CHANGES IN XENOBIOTIC METABOLISM ENZYME EXPRESSION DURING AGING USING COMPREHENSIVE TRANSCRIPT PROFILING  

EPA Science Inventory

Aging leads to changes in the expression of enzymes and transporters important in the metabolism and fate of xenobiotics in liver, kidney and intestine. Most notable are the changes in a number of CYP and xenobiotic transporter genes regulated by the nuclear receptors PXR, CAR an...

147

The involvement of extracellular enzymes in the metabolism of Bdellovibrio  

Microsoft Academic Search

Summary  \\u000a \\u000a \\u000a \\u000a 1. \\u000a \\u000a Three host-independent (H-I)Bdellovibrio species are stimulated in growth by addition to the medium of cell-free host extracts. A mixture of fifteen labeled amino\\u000a acids was used to follow incorporation of counts into trichloroacetic acid (TCA) insoluble material. No label was recovered\\u000a in the nucleic acids.\\u000a \\u000a \\u000a \\u000a \\u000a 2. \\u000a \\u000a LabeledEscherichia coli RNA, protein, andAeromonas DNA preparations were hydrolyzed byBdellovibrio extracellular enzymes.

H. Mark Engelking; Ramon J. Seidler

1974-01-01

148

Regulation of drug-metabolizing cytochrome P450 enzymes by glucocorticoids.  

PubMed

The regulation of drug-metabolizing cytochrome P450 enzymes (CYP) is a complex process involving multiple mechanisms. Among them, transcriptional regulation through ligand-activated nuclear receptors is the crucial mechanism involved in hormone-controlled and xenobiotic-induced expression of drug-metabolizing CYPs. In this article, we focus, in detail, on the role of the glucocorticoid receptor (GR) in the transcriptional regulation of human drug-metabolizing CYP enzymes and the mechanisms of the regulation. There are at least three distinct transcriptional mechanisms by which GR controls the expression of CYPs: 1) direct binding of GR to a specific gene-promoter sequence called the glucocorticoid responsive element (GRE); 2) indirect binding of GR in the form of a multiprotein complex to gene promoters without a direct contact between GR and promoter DNA; and 3) up- or downregulation of other CYP transcriptional regulators or nuclear receptors (i.e., transcriptional regulatory cross-talk). However, due to the general effect of glucocorticoids on numerous cellular pathways and functions, the net transcriptional effect of glucocorticoids on drug-metabolizing enzymes is usually a combination of several mechanisms. Since synthetic glucocorticoids are widely prescribed in human pharmacotherapy for the treatment of many diseases, comprehensive understanding of the transcriptional regulation of drug-metabolizing CYPs via GR with respect to glucocorticoid therapy or glucocorticoid hormonal status will aid in the development of efficient individualized pharmacotherapy without drug-drug interactions. PMID:20482443

Dvorak, Zdenek; Pavek, Peotr

2010-11-01

149

Homeostatic control of xeno- and endobiotics in the drug-metabolizing enzyme system.  

PubMed

Drug-metabolizing Phase I and II enzyme families, drug transporters (Phase III) and their ligand-activated transcription factors probably evolved as a system involved in homeostatic control of lipophilic endobiotics and detoxification of xenobiotics. The review is focused on CYP, UGT enzymes and the Ah receptor as transcription factor. The hypothesis of a system is supported by (i) coordinate regulation of subsets of these enzyme families and transporters by transcription factors including the AhR, and (ii) feedback loops between endobiotic AhR agonists and substrates of major catabolic target genes/proteins; for example, 6-formylindolo[3,2-b]carbazole as substrate of CYP1A1, and bilirubin, as substrate of UGT1A1. In the latter case the AhR is one of multiple transcription factors contributing to bilirubin homeostasis. Interestingly, xenobiotics including dietary phytochemicals, products of microbiota, ubiquitous environmental pollutants such as benzo[a]pyrene may also have shaped this system in intestinal epithelia during millions of years of evolution. Most lipophilic drugs are metabolized by the same system since drug-metabolizing enzymes are broad substrate spectrum enzymes. Better understanding of this system may lead to generation of drugs with desirable therapeutic properties. PMID:24837423

Bock, Karl Walter

2014-07-01

150

Regional variation in muscle metabolic enzymes in individual American shad (Alosa sapidissima)  

USGS Publications Warehouse

Evaluation of the activity of metabolic enzymes is often used to asses metabolic capacity at the tissue level, but the amount of regional variability within a tissue in an individual fish of a given species is frequently unknown. The activities of four enzymes (citrate synthase (CS), phosphofructokinase, lactate dehydrogenase (LDH), and ??-hydroxyacyl coenzyme A dehydrogenase (HOAD) were assayed in red and white muscle at 10 sites along the body of adult American shad (Alosa sapidissima). Red and white muscle HOAD and white muscle CS and LDH varied significantly, generally increasing posteriorly. Maximal variation occurs in red muscle HOAD (~450%) and white muscle LDH (~60%) activity. Differences between the sexes also vary with sampling location. This study suggests that the variability in enzyme activity may be linked to functional differences in the muscle at different locations, and also provides guidelines for sample collection in this species.

Leonard, J.B.K.

1999-01-01

151

Odorant Metabolism Catalyzed by Olfactory Mucosal Enzymes Influences Peripheral Olfactory Responses in Rats  

PubMed Central

A large set of xenobiotic-metabolizing enzymes (XMEs), such as the cytochrome P450 monooxygenases (CYPs), esterases and transferases, are highly expressed in mammalian olfactory mucosa (OM). These enzymes are known to catalyze the biotransformation of exogenous compounds to facilitate elimination. However, the functions of these enzymes in the olfactory epithelium are not clearly understood. In addition to protecting against inhaled toxic compounds, these enzymes could also metabolize odorant molecules, and thus modify their stimulating properties or inactivate them. In the present study, we investigated the in vitro biotransformation of odorant molecules in the rat OM and assessed the impact of this metabolism on peripheral olfactory responses. Rat OM was found to efficiently metabolize quinoline, coumarin and isoamyl acetate. Quinoline and coumarin are metabolized by CYPs whereas isoamyl acetate is hydrolyzed by carboxylesterases. Electro-olfactogram (EOG) recordings revealed that the hydroxylated metabolites derived from these odorants elicited lower olfactory response amplitudes than the parent molecules. We also observed that glucurono-conjugated derivatives induced no olfactory signal. Furthermore, we demonstrated that the local application of a CYP inhibitor on rat olfactory epithelium increased EOG responses elicited by quinoline and coumarin. Similarly, the application of a carboxylesterase inhibitor increased the EOG response elicited by isoamyl acetate. This increase in EOG amplitude provoked by XME inhibitors is likely due to enhanced olfactory sensory neuron activation in response to odorant accumulation. Taken together, these findings strongly suggest that biotransformation of odorant molecules by enzymes localized to the olfactory mucosa may change the odorant’s stimulating properties and may facilitate the clearance of odorants to avoid receptor saturation. PMID:23555703

Thiebaud, Nicolas; Veloso Da Silva, Stéphanie; Jakob, Ingrid; Sicard, Gilles; Chevalier, Joëlle; Ménétrier, Franck; Berdeaux, Olivier; Artur, Yves; Heydel, Jean-Marie; Le Bon, Anne-Marie

2013-01-01

152

Metabolic Pathways of Inhaled Glucocorticoids by the CYP3A Enzymes  

PubMed Central

Asthma is one of the most prevalent diseases in the world, for which the mainstay treatment has been inhaled glucocorticoids (GCs). Despite the widespread use of these drugs, approximately 30% of asthma sufferers exhibit some degree of steroid insensitivity or are refractory to inhaled GCs. One hypothesis to explain this phenomenon is interpatient variability in the clearance of these compounds. The objective of this research is to determine how metabolism of GCs by the CYP3A family of enzymes could affect their effectiveness in asthmatic patients. In this work, the metabolism of four frequently prescribed inhaled GCs, triamcinolone acetonide, flunisolide, budesonide, and fluticasone propionate, by the CYP3A family of enzymes was studied to identify differences in their rates of clearance and to identify their metabolites. Both interenzyme and interdrug variability in rates of metabolism and metabolic fate were observed. CYP3A4 was the most efficient metabolic catalyst for all the compounds, and CYP3A7 had the slowest rates. CYP3A5, which is particularly relevant to GC metabolism in the lungs, was also shown to efficiently metabolize triamcinolone acetonide, budesonide, and fluticasone propionate. In contrast, flunisolide was only metabolized via CYP3A4, with no significant turnover by CYP3A5 or CYP3A7. Common metabolites included 6?-hydroxylation and ?6-dehydrogenation for triamcinolone acetonide, budesonide, and flunisolide. The structure of ?6-flunisolide was unambiguously established by NMR analysis. Metabolism also occurred on the D-ring substituents, including the 21-carboxy metabolites for triamcinolone acetonide and flunisolide. The novel metabolite 21-nortriamcinolone acetonide was also identified by liquid chromatography–mass spectrometry and NMR analysis. PMID:23143891

Moore, Chad D.; Roberts, Jessica K.; Orton, Christopher R.; Murai, Takahiro; Fidler, Trevor P.; Reilly, Christopher A.; Ward, Robert M.

2013-01-01

153

Incorporation of enzyme concentrations into FBA and identification of optimal metabolic pathways  

PubMed Central

Background In the present article, we propose a method for determining optimal metabolic pathways in terms of the level of concentration of the enzymes catalyzing various reactions in the entire metabolic network. The method, first of all, generates data on reaction fluxes in a pathway based on steady state condition. A set of constraints is formulated incorporating weighting coefficients corresponding to concentration of enzymes catalyzing reactions in the pathway. Finally, the rate of yield of the target metabolite, starting with a given substrate, is maximized in order to identify an optimal pathway through these weighting coefficients. Results The effectiveness of the present method is demonstrated on two synthetic systems existing in the literature, two pentose phosphate, two glycolytic pathways, core carbon metabolism and a large network of carotenoid biosynthesis pathway of various organisms belonging to different phylogeny. A comparative study with the existing extreme pathway analysis also forms a part of this investigation. Biological relevance and validation of the results are provided. Finally, the impact of the method on metabolic engineering is explained with a few examples. Conclusions The method may be viewed as determining an optimal set of enzymes that is required to get an optimal metabolic pathway. Although it is a simple one, it has been able to identify a carotenoid biosynthesis pathway and the optimal pathway of core carbon metabolic network that is closer to some earlier investigations than that obtained by the extreme pathway analysis. Moreover, the present method has identified correctly optimal pathways for pentose phosphate and glycolytic pathways. It has been mentioned using some examples how the method can suitably be used in the context of metabolic engineering. PMID:18634554

De, Rajat K; Das, Mouli; Mukhopadhyay, Subhasis

2008-01-01

154

Role of high-fat diet in regulation of gene expression of drug metabolizing enzymes and transporters  

PubMed Central

Aim Our aim is to investigate the molecular mechanism of regulation of gene expression of drug metabolizing enzymes (DMEs) and transporters in diet-induced obesity. Main methods Adult male CD1 mice were fed diet containing 60% kcal fat (HFD) or 10% kcal fat (LFD) for 14 weeks. RNA levels of hepatic DMEs, transporters and their regulatory nuclear receptors (NRs) were analyzed by real-time PCR. Activation of cell-signaling components (JNK and NF-?B) and pro-inflammatory cytokines (IL-1?, IL-6 and TNF?) were measured in the liver. Finally, the pharmacodynamics of drugs metabolized by DMEs was measured to determine the clinical relevance of our findings. Key findings RNA levels of the hepatic phase I (Cyp3a11, Cyp2b10, Cyp2a4) and phase II (Ugt1a1, Sult1a1, Sultn) enzymes were reduced ~30-60% in HFD compared to LFD mice. RNA levels of Cyp2e1, Cyp1a2 and the drug transporters, multidrug resistance proteins, (Mrp)2, Mrp3 and multidrug resistant gene (Mdr)1b were unaltered in HFD mice. Gene expression of the NRs, PXR and CAR and nuclear protein levels of RXR? was reduced in HFD mice. Cytokines, JNK and NF-?B were induced in HFD mice. Thus reduction in hepatic gene expression in obesity may be modulated by cross-talk between NRs and inflammation-induced cell-signaling. Sleep time of Midazolam (Cyp3a substrate) was prolonged in HFD mice, while Zoxazolamine (Cyp1a2 and Cyp2e1 substrate)-induced sleep time was unaltered. Significance This study demonstrates that gene-specific reductions in DMEs can affect specific drugs metabolized by these enzymes, thus providing a rationale to monitor the effectiveness of drug therapy in obese individuals. PMID:21620874

Ghose, Romi; Omoluabi, Ozozoma; Gandhi, Adarsh; Shah, Pranav; Strohacker, Kelley; Carpenter, Katie C; McFarlin, Brian; Guo, Tao

2011-01-01

155

Enzyme Basis for pH Regulation of Citrate and Pyruvate Metabolism by Leuconostoc oenos  

PubMed Central

Citrate and pyruvate metabolism by nongrowing cells of Leuconostoc oenos was investigated. (sup13)C nuclear magnetic resonance (NMR) spectroscopy was used to elucidate the pathway of citrate breakdown and to probe citrate or pyruvate utilization, noninvasively, in living cell suspensions. The utilization of isotopically enriched substrates allowed us to account for the end products derived from the metabolism of endogenous reserves. The effect of environmental parameters, e.g., pH, gas atmosphere, and presence of malate, on the end products of citrate utilization was studied. Approximately 10% of the citrate supplied was converted to aspartate which remained inside the cells. A metabolic shift with pH was observed, with acetoin production being favored at pH 4, whereas lactate and acetate production increased significantly at higher pH values. The information obtained with NMR was complemented with studies on the relevant enzyme activities in the metabolic pathway of citrate breakdown. The intracellular pH of the cells was strongly dependent on the external pH; this result, together with the determination of the pH profile of the enzymic activities, allowed us to establish the basis for pH regulation; lactate dehydrogenase activity was optimal at pH 7, whereas the acetoin-forming enzymes displayed maximal activities below pH 5. Citrate utilization was also monitored in dilute cell suspensions for comparison with NMR experiments performed with dense suspensions. PMID:16534990

Ramos, A.; Lolkema, J. S.; Konings, W. N.; Santos, H.

1995-01-01

156

UCLA scientists discover how key enzyme involved in aging, cancer assembles  

Cancer.gov

UCLA biochemists have mapped the structure of a key protein–RNA complex that is required for the assembly of telomerase, an enzyme important in both cancer and aging. The researchers found that a region at the end of the p65 protein that includes a flexible tail is responsible for bending telomerase's RNA backbone in order to create a scaffold for the assembly of other protein building blocks. Understanding this protein, which is found in a type of single-celled organism that lives in fresh water ponds, may help researchers predict the function of similar proteins in humans and other organisms. UCLA is home to the Jonsson Comprehensive Cancer Center.

157

A profile of Drosophila species' enzymes assayed by electrophoresis. I. Number of alleles, heterozygosities, and linkage disequilibrium in glucose-metabolizing systems and some other enzymes  

Microsoft Academic Search

Seven isolated large populations of Drosophila belonging to five different species were examined by starch gel electrophoresis for allozyme variation. Six to eleven enzyme loci in the glucose-metabolizing system (group I) and six to eight enzyme loci (group II) which were not directly involved in the above-mentioned system were assayed. The parameters estimated were the average number of alleles per

Ken-ichi Kojima; John Gillespie; Yoshiko N. Tobari

1970-01-01

158

Expression in human prostate of drug- and carcinogen-metabolizing enzymes: association with prostate cancer risk.  

PubMed Central

The role of two common polymorphisms of enzymes involved in the metabolism of drugs and carcinogens was studied in relation to prostate cancer. The gene encoding one of these enzymes (NAT2) is located in an area where frequent allelic loss occurs in prostate cancer. Mutations at the genes CYP2D6 and NAT2 were analysed by allele-specific polymerase chain reaction and restriction mapping in DNA from 94 subjects with prostate cancer and 160 male healthy control subjects. Eleven prostate specimens were analysed for genotype and enzymatic activities NAT2, CYP2D6 and CYP3A by using the enzyme-specific substrates sulphamethazine and dextromethorphan. Enzyme activities with substrate specificities corresponding to NAT2, CYP2D6 and CYP3A are present in human prostate tissue, with mean +/-s.d. activities of 4.8+/-4.4 pmol min(-1) mg(-1) protein, 156+/-91 and 112+/-72 nmol min(-1) mg(-1) protein respectively. The Km values for the prostate CYP2D6 and CYP3A enzyme activities corresponded to that of liver CYP2D6 and CYP3A activities, and the CYP2D6 enzyme activity is related to the CYP2D6 genotype. The N-acetyltransferase, in contrast, had a higher Km than NAT2 and was independent of the NAT2 genotype. The CYP2D6 and CYP3A enzymes, and an N-acetyltransferase activity that is independent of the regulation of the NAT2 gene, are expressed in human prostate tissue. The presence of carcinogen-metabolizing enzymes in human prostate with a high interindividual variability may be involved in the regulation of local levels of carcinogens and mutagens and may underlie interindividual differences in cancer susceptibility. Images Figure 1 PMID:9823980

Agúndez, J. A.; Martínez, C.; Olivera, M.; Gallardo, L.; Ladero, J. M.; Rosado, C.; Prados, J.; Rodriguez-Molina, J.; Resel, L.; Benítez, J.

1998-01-01

159

Controls on the Temperature Sensitivity of Soil Enzymes: A Key Driver of In Situ Enzyme Activity Rates  

Microsoft Academic Search

\\u000a Enzyme activities are commonly measured in lab assays at a single standard temperature, which does not provide any information\\u000a on their temperature sensitivity. Temperature is one of the primary controls on enzyme activities, yet few studies have explored\\u000a how temperature drives enzyme activities in the environment. The temperature sensitivity of enzyme activity is controlled\\u000a by the structure and conformation of

Matthew Wallenstein; Steven D. Allison; Jessica Ernakovich; J. Megan Steinweg; Robert Sinsabaugh

160

Comparative metabolism as a key driver of wildlife species sensitivity to human and veterinary pharmaceuticals.  

PubMed

Human and veterinary drug development addresses absorption, distribution, metabolism, elimination and toxicology (ADMET) of the Active Pharmaceutical Ingredient (API) in the target species. Metabolism is an important factor in controlling circulating plasma and target tissue API concentrations and in generating metabolites which are more easily eliminated in bile, faeces and urine. The essential purpose of xenobiotic metabolism is to convert lipid-soluble, non-polar and non-excretable chemicals into water soluble, polar molecules that are readily excreted. Xenobiotic metabolism is classified into Phase I enzymatic reactions (which add or expose reactive functional groups on xenobiotic molecules), Phase II reactions (resulting in xenobiotic conjugation with large water-soluble, polar molecules) and Phase III cellular efflux transport processes. The human-fish plasma model provides a useful approach to understanding the pharmacokinetics of APIs (e.g. diclofenac, ibuprofen and propranolol) in freshwater fish, where gill and liver metabolism of APIs have been shown to be of importance. By contrast, wildlife species with low metabolic competency may exhibit zero-order metabolic (pharmacokinetic) profiles and thus high API toxicity, as in the case of diclofenac and the dramatic decline of vulture populations across the Indian subcontinent. A similar threat looms for African Cape Griffon vultures exposed to ketoprofen and meloxicam, recent studies indicating toxicity relates to zero-order metabolism (suggesting P450 Phase I enzyme system or Phase II glucuronidation deficiencies). While all aspects of ADMET are important in toxicity evaluations, these observations demonstrate the importance of methods for predicting API comparative metabolism as a central part of environmental risk assessment. PMID:25405970

Hutchinson, Thomas H; Madden, Judith C; Naidoo, Vinny; Walker, Colin H

2014-11-19

161

Localization of enzymes relating to C4 organic acid metabolisms in the marine diatom, Thalassiosira pseudonana.  

PubMed

In the genome of the marine diatom-Thalassiosira pseudonana, there are several putative genes encoding enzymes potentially constitute a classical C4 type biochemical CO2-concentrating mechanism. Two genes encode a carboxylation enzyme phosphoenolpyruvate carboxylase (PEPC)1 and PEPC2; and another two encode decarboxylation enzymes, NAD(+)-dependent malic enzyme (NAD-ME) and phosphoenolpyruvate carboxykinase (PEPCK). These genes were tagged by the enhanced-green fluorescence protein, egfp, ligated in the transformation vector, and transformed into the cells of T. pseudonana for localization of GFP fusion products. The PEPC1:GFP fusion was localized at the matrix of the periplastidal compartment, while the PEPC2:GFP fusion was localized at the mitochondria. The NAD-ME:GFP fusion was localized in the cytosol and the PEPCK:GFP fusion at the mitochondria. The transcripts level of NAD-ME was extremely low, and PEPCK transcript was significantly induced under the dark, suggesting that PEPCK is involved in the dark metabolism such as respiration and amino acid metabolism in the mitochondria. Treatments of low-CO2grown T. pseudonana cells with inhibitors for PEPCK and PEPC efficiently dissipated the maximum rate of photosynthesis while these treatments did not affect high-affinity photosynthesis. These data strongly suggest that classical C4 enzymes play little role in the CCM in T. pseudonana. PMID:24414292

Tanaka, Rie; Kikutani, Sae; Mahardika, Anggara; Matsuda, Yusuke

2014-09-01

162

PLP-dependent enzymes as entry and exit gates of sphingolipid metabolism  

PubMed Central

Sphingolipids are membrane constituents as well as signaling molecules involved in many essential cellular processes. Serine palmitoyltransferase (SPT) and sphingosine-1-phosphate lyase (SPL), both PLP (pyridoxal 5?-phosphate)-dependent enzymes, function as entry and exit gates of the sphingolipid metabolism. SPT catalyzes the condensation of serine and a fatty acid into 3-keto-dihydrosphingosine, whereas SPL degrades sphingosine-1-phosphate (S1P) into phosphoethanolamine and a long-chain aldehyde. The recently solved X-ray structures of prokaryotic homologs of SPT and SPL combined with functional studies provide insight into the structure–function relationship of the two enzymes. Despite carrying out different reactions, the two enzymes reveal striking similarities in the overall fold, topology, and residues crucial for activity. Unlike their eukaryotic counterparts, bacterial SPT and SPL lack a transmembrane helix, making them targets of choice for biochemical characterization because the use of detergents can be avoided. Both human enzymes are linked to severe diseases or disorders and might therefore serve as targets for the development of therapeutics aiming at the modulation of their activity. This review gives an overview of the sphingolipid metabolism and of the available biochemical studies of prokaryotic SPT and SPL, and discusses the major similarities and differences to the corresponding eukaryotic enzymes. PMID:21710479

Bourquin, Florence; Capitani, Guido; Grütter, Markus Gerhard

2011-01-01

163

Stereochemical course, isotope effects, and enzyme inhibitor studies of glaucine metabolism in fungi  

SciTech Connect

The microbial transformation of the aporphine alkaloid glaucine by the fungi Fusarium solani (ATCC 12823) and Aspergillus flavipes (ATCC 1030) proceeds with complete substrate stereoselectivity. The fungus F. solani metabolizes only S-(+)-glaucine (1) to dehydroglaucine (3), and A. flavipes metabolizes only R-(-)-glaucine (2) to dehydroglaucine. This facile microbiological reaction is useful in the destructive resolution of racemic mixtures of glaucine, and may provide a model for producing pure enantiomers (either R or S) of other aporphines from racemic mixtures. In order to extend the reaction to other aporphines and related alkaloids, the overall stereochemical course and enzyme(s) involved in the reaction, and the substrate requirements of the enzyme were investigated. The overall stereochemical course of the transformation was examined using C-7 methyl-blocked analogs of glaucine, cis- and trans-7-methylglaucine, as substrates for the fungi. Isolation and examination of residual substrates from semi-preparative scale incubations by MS, PMR, PMR with a chiral shift reagent, OR and ORD indicated that the transformation was enantioselective in the case of A. flavipes. However, only a 10% enrichment of 6aR,7R-cis-7-methylglaucine was observed in F. solani cultures. The oxidation of glaucine can be envisioned as proceeding through one of several mechanisms, each involving a different enzyme system. Deuterium isotope, induction and enzyme inhibitor experiments helped to distinguish between the three mechanisms.

Kerr, K.M.

1986-01-01

164

Functional genomics and SNP analysis of human genes encoding proline metabolic enzymes  

Microsoft Academic Search

Proline metabolism in mammals involves two other amino acids, glutamate and ornithine, and five enzymatic activities, ?1-pyrroline-5-carboxylate (P5C) reductase (P5CR), proline oxidase, P5C dehydrogenase, P5C synthase and ornithine-?-aminotransferase\\u000a (OAT). With the exception of OAT, which catalyzes a reversible reaction, the other four enzymes are unidirectional, suggesting\\u000a that proline metabolism is purpose-driven, tightly regulated, and compartmentalized. In addition, this tri-amino-acid system

Chien-an A. Hu; D. Bart Williams; Siqin Zhaorigetu; Shadi Khalil; Guanghua Wan; David Valle

2008-01-01

165

Effects of nutrition and temperature on metabolic enzyme activities in larval and juvenile red drum, Sciaenops ocellatus , and lane snapper, Lutjanus synagris  

Microsoft Academic Search

The metabolic enzyme activities were determined in larvae of red drum, Sciaenops ocellatus, and lane snapper, Lutjanus synagris, to determine the effect of temperature and nutrition on metabolic enzyme activities and to evaluate if metabolic enzyme activities are useful in assessing the feeding condition of larval fish. During experiments conducted during the spring of 1990, lactate dehydrogenase (LDH) activities in

M. E. Clarke; C. Calvi; M. Domeier; M. Edmonds; P. J. Walsh

1992-01-01

166

Fructose: A Key Factor in the Development of Metabolic Syndrome and Hypertension  

PubMed Central

Diabetes mellitus and the metabolic syndrome are becoming leading causes of death in the world. Identifying the etiology of diabetes is key to prevention. Despite the similarity in their structures, fructose and glucose are metabolized in different ways. Uric acid, a byproduct of uncontrolled fructose metabolism is known risk factor for hypertension. In the liver, fructose bypasses the two highly regulated steps in glycolysis, glucokinase and phosphofructokinase, both of which are inhibited by increasing concentrations of their byproducts. Fructose is metabolized by fructokinase (KHK). KHK has no negative feedback system, and ATP is used for phosphorylation. This results in intracellular phosphate depletion and the rapid generation of uric acid due to activation of AMP deaminase. Uric acid, a byproduct of this reaction, has been linked to endothelial dysfunction, insulin resistance, and hypertension. We present possible mechanisms by which fructose causes insulin resistance and suggest actions based on this association that have therapeutic implications. PMID:23762544

Kim, Dong Hyun

2013-01-01

167

Mutations in hereditary phosphoglucomutase 1 deficiency map to key regions of enzyme structure and function.  

PubMed

Recent studies have identified phosphoglucomutase 1 (PGM1) deficiency as an inherited metabolic disorder in humans. PGM1 deficiency is classified as both a muscle glycogenosis (type XIV) and a congenital disorder of glycosylation of types I and II. Affected patients show multiple disease phenotypes, reflecting the central role of the enzyme in glucose homeostasis, where it catalyzes the interconversion of glucose 1-phosphate and glucose 6-phosphate. The influence of PGM1 deficiency on protein glycosylation patterns is also widespread, affecting both biosynthesis and processing of glycans and their precursors. To date, 21 different mutations involved in PGM1 deficiency have been identified, including 13 missense mutations resulting in single amino acid changes. Growing clinical interest in PGM1 deficiency prompts a review of the molecular context of these mutations in the three-dimensional structure of the protein. Here the known crystal structure of PGM from rabbit (97 % sequence identity to human) is used to analyze the mutations associated with disease and find that many map to regions with clear significance to enzyme function. In particular, amino acids in and around the active site cleft are frequently involved, including regions responsible for catalysis, binding of the metal ion required for activity, and interactions with the phosphosugar substrate. Several of the known mutations, however, are distant from the active site and appear to manifest their effects indirectly. An understanding of how the different mutations that cause PGM1 deficiency affect enzyme structure and function is foundational to providing clinical prognosis and the development of effective treatment strategies. PMID:25168163

Beamer, Lesa J

2015-03-01

168

A comparative study of some more important experimental animal peroxide metabolism enzymes.  

PubMed

1. We have studied and compared the peroxide metabolism enzymes (SOD, P and C) of the main organs of fresh-water mollusc, chicken, mouse, guinea-pig, rabbit, cat and dog. 2. The liver exhibited the highest SOD activity. The enzymatic activities of the organ homogenates of the guinea-pig stand out in comparison with the values for the other homogenates examined. 3. The liver, kidney and total brain homogenates of the chicken and the vertebrates display no, or only a very low P activity. The highest P activities were measured in the haemolysates. 4. The C and SOD values exhibit a certain parallelism. 5. The peroxide metabolism enzyme activities calculated by utilizing the protein measurements permit the establishment of a more realistic enzymatic activity. PMID:233784

Matkovics, B; Novák, R; Hanh, H D; Szabó, L; Varga, S I

1977-01-01

169

Reactions and enzymes in the metabolism of drugs and other xenobiotics.  

PubMed

In this article, we offer an overview of the compared quantitative importance of biotransformation reactions in the metabolism of drugs and other xenobiotics, based on a meta-analysis of current research interests. Also, we assess the relative significance the enzyme (super)families or categories catalysing these reactions. We put the facts unveiled by the analysis into a drug discovery context and draw some implications. The results confirm the primary role of cytochrome P450-catalysed oxidations and UDP-glucuronosyl-catalysed glucuronidations, but they also document the marked significance of several other reactions. Thus, there is a need for several drug discovery scientists to better grasp the variety of drug metabolism reactions and enzymes and their consequences. PMID:22305937

Testa, Bernard; Pedretti, Alessandro; Vistoli, Giulio

2012-06-01

170

Effect of trifluoperazine on certain arterial wall lipid-metabolizing enzymes inducing atherosclerosis in rhesus monkeys.  

PubMed

The effect of trifluoperazine (TFP) was investigated on arterial wall lipid-metabolizing enzymes like acyl-CoA:cholesterol acyltransferase (ACAT) and cholesterol ester hydrolase (CEH) in rhesus monkeys. The activity was determined in aortic wall homogenates obtained from rhesus monkeys fed an atherogenic diet coupled with intramuscular injections of adrenaline and TFP. Although TFP had no significant effect on serum cholesterol and triglycerides, it decreased significantly the formation of atherosclerotic lesions by decreasing the esterification of cholesterol, by inhibiting ACAT and enhancing its utilization by activating CEH. Hence, the preventive effect of TFP on the development of atherosclerosis in rhesus monkeys is mediated through its ability to influence the activities of arterial wall lipid-metabolizing enzymes like ACAT and CEH. PMID:9270979

Mohindroo, A; Ahluwalia, P

1997-08-01

171

Behavior of Activities of Thymidine Metabolizing Enzymes in Human Leukemia-LymphomaCells1  

Microsoft Academic Search

The behavior of the activities of thymidine metabolizing enzymes, dihydrothymine dehydrogenase (EC 1.3.1.2) and thymidine phosphoryl- ase (EC 2.4.2.4) for thymidine degradation, thymidine kinase (EC 2.7.1.75) and thymidylate synthase (EC 2.1.1.45) for DNA synthesis, was elucidated in cytosolic extracts from normal human lymphocytes and 13 human leukemia-lymphoma cell lines. In the normal human lympho cytes, the activities of dihydrothymine dehydrogenase,

Taiichi Shiotani; Yasuko Hashimoto; Terukazu Tanaka; Shozo Irino

172

Genes, Enzymes, and Regulation of para-Cresol Metabolism in Geobacter metallireducens? †  

PubMed Central

In aerobic and facultatively anaerobic bacteria, the degradation of para-cresol (p-cresol) involves the initial hydroxylation to p-hydroxybenzyl alcohol by water catalyzed by the soluble, periplasmatic flavocytochrome p-cresol methylhydroxylase (PCMH; ?2?2 composition). In denitrifying bacteria the further metabolism proceeds via oxidation to p-hydroxybenzoate, the formation of p-hydroxybenzoyl-coenzyme A (CoA), and the subsequent dehydroxylation of the latter to benzoyl-CoA by reduction. In contrast, the strictly anaerobic Desulfobacterium cetonicum degrades p-cresol by addition to fumarate, yielding p-hydroxybenzylsuccinate. In this work, in vitro enzyme activity measurements revealed that the obligately anaerobic Geobacter metallireducens uses the p-cresol degradation pathway of denitrifying bacteria. Surprisingly, PCMH, which is supposed to catalyze both p-cresol hydroxylation and p-hydroxybenzyl alcohol oxidation to the corresponding aldehyde, was located in the membrane fraction. The ? subunit of the enzyme was present in two isoforms, suggesting an ????2 composition. We propose that the unusual asymmetric architecture and the membrane association of PCMH might be important for alternative electron transfer routes to either cytochrome c (in the case of p-cresol oxidation) or to menaquinone (in the case of p-hydroxybenzyl alcohol oxidation). Unusual properties of further enzymes of p-cresol metabolism, p-hydroxybenzoate-CoA ligase, and p-hydroxybenzoyl-CoA reductase were identified and are discussed. A proteomic approach identified a gene cluster comprising most of the putative structural genes for enzymes involved in p-cresol metabolism (pcm genes). Reverse transcription-PCR studies revealed a different regulation of transcription of pcm genes and the corresponding enzyme activities, suggesting the presence of posttranscriptional regulatory elements. PMID:17449613

Peters, Franziska; Heintz, Dimitri; Johannes, Jörg; van Dorsselaer, Alain; Boll, Matthias

2007-01-01

173

Stem sugar accumulation in sweet sorghum - activity and expression of sucrose metabolizing enzymes and sucrose transporters.  

PubMed

Sugar metabolism was studied in sweet sorghum (SSV74) that is known to store sugars in the mature internodes and which is reported to give grain yields twice that of a grain sorghum variety (SPV1616). Comparison of sugar accumulation in these two varieties was carried out at three stages of growth and in the upper and lower internodes. In spite of large differences in the level of sugar accumulation, osmolarity of the sap did not vary as significantly in the two varieties. Significant contribution of variety, stage and internode position was seen for the variation observed in sugar content. Though the activities of sugar metabolizing enzymes namely sucrose synthase (in the synthesis and cleavage directions), sucrose phosphate synthase and invertase (cytoplasmic and vacuolar) also varied in a stage- and internode-specific manner in the two varieties, these enzymes did not contribute significantly to the variation observed in sugar content. Transcriptional expression of one sucrose synthase (SUC1), two sucrose phosphate synthase (SPS2 and SPS3) and a vacuolar invertase (INV3) gene were lower in sweet sorghum as compared to grain sorghum. Sweet sorghum also showed lower expression of two sucrose transporters (SUT1 and SUT4), which correlated to higher sugar accumulation in this variety. Differential expression of the sugar metabolizing enzymes and sucrose transporters in sweet and grain sorghum suggest a role for signaling molecules and transcription factors in regulating sugar accumulation observed in the mature internodes of sweet sorghum, which needs to be investigated. PMID:22325624

Qazi, Hilal Ahmad; Paranjpe, Sharayu; Bhargava, Sujata

2012-04-15

174

Pharmacokinetic interactions between herbal medicines and prescribed drugs: focus on drug metabolic enzymes and transporters.  

PubMed

Herbal medicines have been widely used for thousands of years, and now are gaining continued popularity worldwide as a complementary or alternative treatment for a variety of diseases, rehabilitation and health care. Since herbal medicines contain more than one pharmacologically active ingredient and are commonly used with many prescribed drugs, there are potential herb-drug interactions. A variety of reported herb-drug interactions are of pharmacokinetic origin, arising from the effects of herbal medicines on metabolic enzymes and/or transporters. Such an alteration in metabolism or transport can result in changes in absorption, distribution, metabolism, and excretion (e.g., induction or inhibition of metabolic enzymes, and modulation of uptake and efflux transporters), leading to changed pharmacokinetics of the concomitantly prescribed drugs. Pharmacokinetic herb-drug interactions have more clinical significance as pharmacokinetic parameters such as the area under the plasma concentration-time curve (AUC), the maximum plasma concentration (Cmax) or the elimination half-life (t1/2) of the concomitant drug alter. This review summarizes the mechanism underlying herb-drug interactions and the approaches to identify the interactions, and discusses pharmacokinetic interactions of eight widely used herbal medicines (Ginkgo biloba, ginseng, garlic, black cohosh, Echinacea, milk thistle, kava, and St. John's wort) with conventional drugs, using various in vitro, animal in vivo, and clinical studies. The increasing understanding of pharmacokinetic herb-drug interactions will make health care professionals and patients pay more attention to the potential interactions. PMID:25705905

Meng, Qiang; Liu, Kexin

2014-01-01

175

Intermediary Metabolism in Clostridium acetobutylicum: Levels of Enzymes Involved in the Formation of Acetate and Butyrate  

PubMed Central

The levels of seven intermediary enzymes involved in acetate and butyrate formation from acetyl coenzyme A in the saccharolytic anaerobe Clostridium acetobutylicum were investigated as a function of time in solvent-producing batch fermentations. Phosphate acetyltransferase and acetate kinase, which are known to form acetate from acetyl coenzyme A, both showed a decrease in specific activity when the organism reached the solvent formation stage. The three consecutive enzymes thiolase, ?-hydroxybutyrylcoenzyme A dehydrogenase, and crotonase exhibited a coordinate expression and a maximal activity after growth had ceased. Only low levels of butyryl coenzyme A dehydrogenase activity were found. Phosphate butyryltransferase activity rapidly decreased after 20 h from 5 to 11 U/mg of protein to below the detection limit (1 mU/mg). Butyrate no longer can be formed, and the metabolic flux may be diverted to butanol. Butyrate kinase showed a 2.5- to 10-fold increase in specific activity after phosphate butyryltransferase activity no longer could be detected. These results suggest that the uptake of acetate and butyrate during solvent formation can not proceed via a complete reversal of the phosphate transferase and kinase reactions. The activities of all enzymes investigated as a function of time in vitro are much higher than the metabolic fluxes through them in vivo. This indicates that none of the maximal activities of the enzymes assayed is rate limiting in C. acetobutylicum. PMID:16346566

Hartmanis, Maris G. N.; Gatenbeck, Sten

1984-01-01

176

Cytoplasmic enzyme activities involved in energy and amino acid metabolism in pathological human renal cortex.  

PubMed

Enzyme levels of lactate dehydrogenase (LDH), alpha-hydroxybutyrate dehydrogenase (HBDH), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were measured in the cytosol of renal cortex samples from either normal and pathologic kidney tissue. The mean enzyme activity values, expressed in Units per gram of cytosolic protein decreased in the following order: normal cortex (LDH = 4,299 +/- 654; AST = 522 +/- 101; ALT = 197 +/- 44). chronic pyelonephritis (LDH = 2,360 +/- 876; AST = 297 +/- 117; ALT = 90 +/- 48), hydronephrosis (LDH = 2,208 +/- 1,264; AST = 279 +/- 165; ALT = 82 +/- 61), pyonephrosis (LDH = 1,410 +/- 596; AST = 158 +/- 69; ALT = 23.4 +/- 16.4) and renal tuberculosis (LDH = 1,149 +/- 481; AST = 93 +/- 34; ALT = 5.6 +/- 2.8). The decrease in the enzyme activities paralleled tissue damage and it was shown to affect cellular functionality in relation with energy and amino acid metabolism. PMID:3244890

González-Buitrago, J M; Corrales, J J; Pastor, I

1988-12-01

177

Expression profiling reveals Spot 42 small RNA as a key regulator in the central metabolism of Aliivibrio salmonicida  

PubMed Central

Background Spot 42 was discovered in Escherichia coli nearly 40 years ago as an abundant, small and unstable RNA. Its biological role has remained obscure until recently, and is today implicated in having broader roles in the central and secondary metabolism. Spot 42 is encoded by the spf gene. The gene is ubiquitous in the Vibrionaceae family of gamma-proteobacteria. One member of this family, Aliivibrio salmonicida, causes cold-water vibriosis in farmed Atlantic salmon. Its genome encodes Spot 42 with 84% identity to E. coli Spot 42. Results We generated a A. salmonicida spf deletion mutant. We then used microarray and Northern blot analyses to monitor global effects on the transcriptome in order to provide insights into the biological roles of Spot 42 in this bacterium. In the presence of glucose, we found a surprisingly large number of ? 2X differentially expressed genes, and several major cellular processes were affected. A gene encoding a pirin-like protein showed an on/off expression pattern in the presence/absence of Spot 42, which suggests that Spot 42 plays a key regulatory role in the central metabolism by regulating the switch between fermentation and respiration. Interestingly, we discovered an sRNA named VSsrna24, which is encoded immediately downstream of spf. This new sRNA has an expression pattern opposite to that of Spot 42, and its expression is repressed by glucose. Conclusions We hypothesize that Spot 42 plays a key role in the central metabolism, in part by regulating the pyruvat dehydrogenase enzyme complex via pirin. PMID:22272603

2012-01-01

178

Alteration of the Expression of Pesticide-Metabolizing Enzymes in Pregnant Mice: Potential Role in the Increased Vulnerability of the Developing Brain  

PubMed Central

Studies on therapeutic drug disposition in humans have shown significant alterations as the result of pregnancy. However, it is not known whether pesticide metabolic capacity changes throughout pregnancy, which could affect exposure of the developing brain. We sought to determine the effect of pregnancy on the expression of hepatic enzymes involved in the metabolism of pesticides. Livers were collected from virgin and pregnant C57BL/6 mice at gestational days (GD)7, GD11, GD14, GD17, and postpartum days (PD)1, PD15, and PD30. Relative mRNA expression of several enzymes involved in the metabolism of pesticides, including hepatic cytochromes (Cyp) P450s, carboxylesterases (Ces), and paraoxonase 1 (Pon1), were assessed in mice during gestation and the postpartum period. Compared with virgin mice, alterations in the expression occurred at multiple time points, with the largest changes observed on GD14. At this time point, the expression of most of the Cyps involved in pesticide metabolism in the liver (Cyp1a2, Cyp2d22, Cyp2c37, Cyp2c50, Cyp2c54, and Cyp3a11) were downregulated by 30% or more. Expression of various Ces isoforms and Pon1 were also decreased along with Pon1 activity. These data demonstrate significant alterations in the expression of key enzymes that detoxify pesticides during pregnancy, which could alter exposure of developing animals to these chemicals. PMID:23223497

Fortin, Marie C.; Aleksunes, Lauren M.

2013-01-01

179

Penn Medicine researchers find that a metabolic enzyme stops progression of most common type of kidney cancer  

Cancer.gov

Researchers found that an enzyme called FBP1 -- essential for regulating metabolism -- binds to a transcription factor in the nucleus of certain kidney cells and restrains energy production in the cell body.

180

Model of 2,3-bisphosphoglycerate metabolism in the human erythrocyte based on detailed enzyme kinetic equations: equations and parameter refinement.  

PubMed Central

Over the last 25 years, several mathematical models of erythrocyte metabolism have been developed. Although these models have identified the key features in the regulation and control of erythrocyte metabolism, many important aspects remain unexplained. In particular, none of these models have satisfactorily accounted for 2,3-bisphosphoglycerate (2,3-BPG) metabolism. 2,3-BPG is an important modulator of haemoglobin oxygen affinity, and hence an understanding of the regulation of 2,3-BPG concentration is important for understanding blood oxygen transport. A detailed, comprehensive, and hence realistic mathematical model of erythrocyte metabolism is presented that can explain the regulation and control of 2,3-BPG concentration and turnover. The model is restricted to the core metabolic pathways, namely glycolysis, the 2,3-BPG shunt and the pentose phosphate pathway (PPP), and includes membrane transport of metabolites, the binding of metabolites to haemoglobin and Mg(2+), as well as pH effects on key enzymic reactions and binding processes. The model is necessarily complex, since it is intended to describe the regulation and control of 2,3-BPG metabolism under a wide variety of physiological and experimental conditions. In addition, since H(+) and blood oxygen tension are important external effectors of 2,3-BPG concentration, it was important that the model take into account the large array of kinetic and binding phenomena that result from changes in these effectors. Through an iterative loop of experimental and simulation analysis many values of enzyme-kinetic parameters of the model were refined to yield close conformity between model simulations and 'real' experimental data. This iterative process enabled a single set of parameters to be found which described well the metabolic behaviour of the erythrocyte under a wide variety of conditions. PMID:10477269

Mulquiney, P J; Kuchel, P W

1999-01-01

181

Catalase is a key enzyme in seed recovery from ageing during priming.  

PubMed

Ageing induces seed deterioration expressed as the loss of seed vigour and/or viability. Priming treatment, which consists in soaking of seeds in a solution of low water potential, has been shown to reinvigorate aged seeds. We investigate the importance of catalase in oxidation protection during accelerated ageing and repair during subsequent priming treatment of sunflower (Helianthus annuus L.) seeds. Seeds equilibrated to 0.29g H2Og(-1) dry matter (DM) were aged at 35°C for different durations and then primed by incubation for 7 days at 15°C in a solution of polyethylene glycol 8000 at -2MPa. Accelerated ageing affected seed germination and priming treatment reversed partially the ageing effect. The inhibition of catalase by the addition of aminotriazol during priming treatment reduced seed repair indicating that catalase plays a key role in protection and repair systems during ageing. Ageing was associated with H2O2 accumulation as showed by biochemical quantification and CeCl3 staining. Catalase was reduced at the level of gene expression, protein content and affinity. Interestingly, priming induced catalase synthesis by activating expression and translation of the enzyme. Immunocytolocalization of catalase showed that the enzyme co-localized with H2O2 in the cytosol. These results clearly indicate that priming induce the synthesis of catalase which is involved in seed recovery during priming. PMID:21763542

Kibinza, Serge; Bazin, Jérémie; Bailly, Christophe; Farrant, Jill M; Corbineau, Françoise; El-Maarouf-Bouteau, Hayat

2011-09-01

182

Gestational Age-Dependent Changes in Gene Expression of Metabolic Enzymes and Transporters in Pregnant Mice  

PubMed Central

Pregnancy-induced changes in drug pharmacokinetics can be explained by changes in expression of drug-metabolizing enzymes and transporters and/or normal physiology. In this study, we determined gestational age-dependent expression profiles for all metabolic enzyme and transporter genes in the maternal liver, kidney, small intestine, and placenta of pregnant mice by microarray analysis. We specifically examined the expression of genes important for xenobiotic, bile acid, and steroid hormone metabolism and disposition, namely, cytochrome P450s (Cyp), UDP-glucuronosyltranserases (Ugt), sulfotransferases (Sult), and ATP-binding cassette (Abc), solute carrier (Slc), and solute carrier organic anion (Slco) transporters. Few Ugt and Sult genes were affected by pregnancy. Cyp17a1 expression in the maternal liver increased 3- to 10-fold during pregnancy, which was the largest observed change in the maternal tissues. Cyp1a2, most Cyp2 isoforms, Cyp3a11, and Cyp3a13 expression in the liver decreased on gestation days (gd) 15 and 19 compared with nonpregnant controls (gd 0). In contrast, Cyp2d40, Cyp3a16, Cyp3a41a, Cyp3a41b, and Cyp3a44 in the liver were induced throughout pregnancy. In the placenta, Cyp expression on gd 10 and 15 was upregulated compared with gd 19. Notable changes were also observed in Abc and Slc transporters. Abcc3 expression in the liver and Abcb1a, Abcc4, and Slco4c1 expression in the kidney were downregulated on gd 15 and 19. In the placenta, Slc22a3 (Oct3) expression on gd 10 was 90% lower than that on gd 15 and 19. This study demonstrates important gestational age-dependent expression of metabolic enzyme and transporter genes, which may have mechanistic relevance to drug disposition in human pregnancy. PMID:23175668

Shuster, Diana L.; Bammler, Theo K.; Beyer, Richard P.; MacDonald, James W.; Tsai, Jesse M.; Farin, Frederico M.; Hebert, Mary F.; Thummel, Kenneth E.

2013-01-01

183

Carbohydrate Metabolism in Archaea: Current Insights into Unusual Enzymes and Pathways and Their Regulation  

PubMed Central

SUMMARY The metabolism of Archaea, the third domain of life, resembles in its complexity those of Bacteria and lower Eukarya. However, this metabolic complexity in Archaea is accompanied by the absence of many “classical” pathways, particularly in central carbohydrate metabolism. Instead, Archaea are characterized by the presence of unique, modified variants of classical pathways such as the Embden-Meyerhof-Parnas (EMP) pathway and the Entner-Doudoroff (ED) pathway. The pentose phosphate pathway is only partly present (if at all), and pentose degradation also significantly differs from that known for bacterial model organisms. These modifications are accompanied by the invention of “new,” unusual enzymes which cause fundamental consequences for the underlying regulatory principles, and classical allosteric regulation sites well established in Bacteria and Eukarya are lost. The aim of this review is to present the current understanding of central carbohydrate metabolic pathways and their regulation in Archaea. In order to give an overview of their complexity, pathway modifications are discussed with respect to unusual archaeal biocatalysts, their structural and mechanistic characteristics, and their regulatory properties in comparison to their classic counterparts from Bacteria and Eukarya. Furthermore, an overview focusing on hexose metabolic, i.e., glycolytic as well as gluconeogenic, pathways identified in archaeal model organisms is given. Their energy gain is discussed, and new insights into different levels of regulation that have been observed so far, including the transcript and protein levels (e.g., gene regulation, known transcription regulators, and posttranslational modification via reversible protein phosphorylation), are presented. PMID:24600042

Esser, Dominik; Rauch, Bernadette

2014-01-01

184

Reduction of nuclear encoded enzymes of mitochondrial energy metabolism in cells devoid of mitochondrial DNA  

SciTech Connect

Highlights: Black-Right-Pointing-Pointer We examined OXPHOS and citrate synthase enzyme activities in HEK293 cells devoid of mtDNA. Black-Right-Pointing-Pointer Enzymes partially encoded by mtDNA show reduced activities. Black-Right-Pointing-Pointer Also the entirely nuclear encoded complex II and citrate synthase exhibit reduced activities. Black-Right-Pointing-Pointer Loss of mtDNA induces a feedback mechanism that downregulates complex II and citrate synthase. -- Abstract: Mitochondrial DNA (mtDNA) depletion syndromes are generally associated with reduced activities of oxidative phosphorylation (OXPHOS) enzymes that contain subunits encoded by mtDNA. Conversely, entirely nuclear encoded mitochondrial enzymes in these syndromes, such as the tricarboxylic acid cycle enzyme citrate synthase (CS) and OXPHOS complex II, usually exhibit normal or compensatory enhanced activities. Here we report that a human cell line devoid of mtDNA (HEK293 {rho}{sup 0} cells) has diminished activities of both complex II and CS. This finding indicates the existence of a feedback mechanism in {rho}{sup 0} cells that downregulates the expression of entirely nuclear encoded components of mitochondrial energy metabolism.

Mueller, Edith E., E-mail: ed.mueller@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Mayr, Johannes A., E-mail: h.mayr@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Zimmermann, Franz A., E-mail: f.zimmermann@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Feichtinger, Rene G., E-mail: r.feichtinger@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Stanger, Olaf, E-mail: o.stanger@rbht.nhs.uk [Department of Cardiac Surgery, Paracelsus Medical University, Muellner Hauptstrasse 48, 5020 Salzburg (Austria)] [Department of Cardiac Surgery, Paracelsus Medical University, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Sperl, Wolfgang, E-mail: w.sperl@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria)] [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Kofler, Barbara, E-mail: b.kofler@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria)] [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria)

2012-01-20

185

Enzyme  

MedlinePLUS

Enzymes are complex proteins that cause a specific chemical change in all parts of the body. For ... use them. Blood clotting is another example of enzymes at work. Enzymes are needed for all body ...

186

Reconstruction of Ancestral Metabolic Enzymes Reveals Molecular Mechanisms Underlying Evolutionary Innovation through Gene Duplication  

PubMed Central

Gene duplications are believed to facilitate evolutionary innovation. However, the mechanisms shaping the fate of duplicated genes remain heavily debated because the molecular processes and evolutionary forces involved are difficult to reconstruct. Here, we study a large family of fungal glucosidase genes that underwent several duplication events. We reconstruct all key ancestral enzymes and show that the very first preduplication enzyme was primarily active on maltose-like substrates, with trace activity for isomaltose-like sugars. Structural analysis and activity measurements on resurrected and present-day enzymes suggest that both activities cannot be fully optimized in a single enzyme. However, gene duplications repeatedly spawned daughter genes in which mutations optimized either isomaltase or maltase activity. Interestingly, similar shifts in enzyme activity were reached multiple times via different evolutionary routes. Together, our results provide a detailed picture of the molecular mechanisms that drove divergence of these duplicated enzymes and show that whereas the classic models of dosage, sub-, and neofunctionalization are helpful to conceptualize the implications of gene duplication, the three mechanisms co-occur and intertwine. PMID:23239941

Vanneste, Kevin; van der Zande, Elisa; Voet, Arnout; Maere, Steven; Verstrepen, Kevin J.

2012-01-01

187

Metabolic pathway analysis of enzyme-deficient human red blood cells.  

PubMed

Five enzymopathies (G6PDH, TPI, PGI, DPGM and PGK deficiencies) in the human red blood cells are investigated using a stoichiometric modeling approach, i.e., metabolic pathway analysis. Elementary flux modes (EFMs) corresponding to each enzyme deficiency case are analyzed in terms of functional capabilities. When available, experimental findings reported in literature related to metabolic behavior of the human red blood cells are compared with the results of EFM analysis. Control-effective flux (CEF) calculation, a novel approach which allows quantification and interpretation of determined EFMs, is performed for further analysis of enzymopathies. Glutathione reductase reaction is found to be the most effective reaction in terms of its CEF value in all enzymopathies in parallel with its known essential role for red blood cells. Efficiency profiles of the enzymatic reactions upon the degree of enzyme deficiency are obtained by the help of the CEF approach, as a basis for future experimental studies. CEF analysis, which is found to be promising in the analysis of erythrocyte enzymopathies, has the potential to be used in modeling efforts of human metabolism. PMID:15555758

Cakir, Tunahan; Tacer, Cemil Serhan; Ulgen, Kutlu Ozergin

2004-12-01

188

Structural Phylogenomics Reveals Gradual Evolutionary Replacement of Abiotic Chemistries by Protein Enzymes in Purine Metabolism  

PubMed Central

The origin of metabolism has been linked to abiotic chemistries that existed in our planet at the beginning of life. While plausible chemical pathways have been proposed, including the synthesis of nucleobases, ribose and ribonucleotides, the cooption of these reactions by modern enzymes remains shrouded in mystery. Here we study the emergence of purine metabolism. The ages of protein domains derived from a census of fold family structure in hundreds of genomes were mapped onto enzymes in metabolic diagrams. We find that the origin of the nucleotide interconversion pathway benefited most parsimoniously from the prebiotic formation of adenine nucleosides. In turn, pathways of nucleotide biosynthesis, catabolism and salvage originated ?300 million years later by concerted enzymatic recruitments and gradual replacement of abiotic chemistries. Remarkably, this process led to the emergence of the fully enzymatic biosynthetic pathway ?3 billion years ago, concurrently with the appearance of a functional ribosome. The simultaneous appearance of purine biosynthesis and the ribosome probably fulfilled the expanding matter-energy and processing needs of genomic information. PMID:23516625

Caetano-Anollés, Kelsey; Caetano-Anollés, Gustavo

2013-01-01

189

Metabolism of halogenated alkanes by cytochrome P450 enzymes. Aerobic oxidation versus anaerobic reduction.  

PubMed

The cytochromes P450 are a large class of heme-containing enzymes that catalyze a broad range of chemical reactions in biosystems, mainly through oxygen-atom transfer to substrates. A relatively unknown reaction catalyzed by the P450s, but very important for human health, is the activation of halogenated substrates, which may lead to toxicity problems. However, its catalytic mechanism is currently unknown and, therefore, we performed a detailed computational study. To gain insight into the metabolism of halogenated compounds by P450 enzymes, we have investigated the oxidative and reductive P450-mediated activation of tetra- and trichloromethane as halogenated models with density functional theory (DFT) methods. We propose an oxidative halosylation mechanism for CCl4 under aerobic conditions by Compound?I of P450, which follows the typical Groves-type rebound mechanism. By contrast, the metabolism of CHCl3 occurs preferentially via an initial hydrogen-atom abstraction rather than halosylation. Kinetic isotope effect studies should, therefore, be able to distinguish the mechanistic pathways of CCl4 versus CHCl3 . We find a novel mechanism that is different from the well accepted P450 substrate activation mechanisms reported previously. Moreover, the studies highlight the substrate specific activation pathways by P450 enzymes leading to different products. These reactivity differences are rationalized using Marcus theory equations, which reproduce experimental product distributions. PMID:24501011

Ji, Li; Zhang, Jing; Liu, Weiping; de Visser, Sam P

2014-04-01

190

The effects of space flight on some rat liver enzymes regulating carbohydrate and lipid metabolism  

NASA Technical Reports Server (NTRS)

The effects of space flight conditions on the activities of certain enzymes regulating carbohydrate and lipid metabolism in rat liver are investigated in an attempt to account for the losses in body weight observed during space flight despite preflight caloric consumption. Liver samples were analyzed for the activities of 32 cytosolic and microsomal enzymes as well as hepatic glycogen and individual fatty acid levels for ground control rats and rats flown on board the Cosmos 936 biosatellite under normal space flight conditions and in centrifuges which were sacrificed upon recovery or 25 days after recovery. Significant decreases in the activities of glycogen phosphorylase, alpha-glycerol phosphate acyl transferase, diglyceride acyl transferase, aconitase and 6-phosphogluconate dehydrogenase and an increase in palmitoyl CoA desaturase are found in the flight stationary relative to the flight contrifuged rats upon recovery, with all enzymes showing alterations returning to normal values 25 days postflight. The flight stationary group is also observed to be characterized by more than twice the amount of liver glycogen of the flight centrifuged group as well as a significant increase in the ratio of palmitic to palmitoleic acid. Results thus indicate metabolic changes which may be involved in the mechanism of weight loss during weightlessness, and demonstrate the equivalence of centrifugation during space flight to terrestrial gravity.

Abraham, S.; Lin, C. Y.; Klein, H. P.; Volkmann, C.

1981-01-01

191

Botanicals as medicinal food and their effects on drug metabolizing enzymes.  

PubMed

Botanicals fall under different regulations in different countries and are mostly consumed without the consultation of the healthcare professional. Over the last decade, utilization of herbal therapies has been extensively documented. The findings indicate the possibility of potential herb-drug interactions due to the concomitant administration of herbal extracts and prescription/over-the-counter drugs. Simultaneously, with the increasing public awareness and search for safer herbal remedies, the study on herbal-drug interactions has gained momentum through the study of drug metabolizing enzymes. Cytochrome P450 (CYP) inhibition or induction is probably the most common mechanism for the pharmacokinetic interactions of herbs and drugs. Any inhibition of CYP enzymes by herbal extracts may result in enhanced plasma and tissue concentration of drugs, leading to toxicity, while induction results in reduced drug concentration leading to decreased drug efficacy and treatment failure. Considering the rapidly growing herbal markets, these types of clinical interactions remain under-reported and unclear. With the increasing consumption of herbal extracts along with prescription medicines, the safety of herbs has become a concern. This article reviews the potential for drug interactions by herbal extracts through drug metabolizing enzymes. PMID:21959528

Mukherjee, Pulok K; Ponnusankar, S; Pandit, Subrata; Hazam, Praksh K; Ahmmed, Milan; Mukherjee, Kakali

2011-12-01

192

Integrative analyses of genetic variation in enzyme activities of primary carbohydrate metabolism reveal distinct modes of regulation in Arabidopsis thaliana  

Microsoft Academic Search

Background Plant primary carbohydrate metabolism is complex and flexible, and is regulated at many levels. Changes of transcript levels do not always lead to changes in enzyme activities, and these do not always affect metabolite levels and fluxes. To analyze interactions between these three levels of function, we have performed parallel genetic analyses of 15 enzyme activities involved in primary

Joost JB Keurentjes; Ronan Sulpice; Yves Gibon; Marie-Caroline Steinhauser; Jingyuan Fu; Maarten Koornneef; Mark Stitt; Dick Vreugdenhil

2008-01-01

193

Impact of a community-based diabetes self-management program on key metabolic parameters  

PubMed Central

Objective: Characterize the impact of a pharmacist-led diabetes self-management program on three key metabolic parameters: glycosylated hemoglobin (HbA1c), low-density lipoprotein cholesterol (LDL-C), and mean arterial blood pressure (MAP) among employee health program participants. Methods: A self-insured company in the Kansas City metropolitan area began offering a pharmacist-led diabetes self-management program to eligible company employees and their dependents in 2008. A retrospective pre-post analysis was conducted to determine if the program affected key metabolic parameters in participants by determining mean change after one year of participation. Results: Among 183 program participants, 65 participants met inclusion criteria. All three key metabolic parameters were significantly reduced from baseline to one year of program participation: HbA1c decreased from 8.1 % to 7.3% (p=0.007); LDL-C decreased from 108.3 mg/dL to 96.4 mg/dL (p=0.009); and MAP decreased from 96.1 to 92.3 mm Hg (p=0.005). Conclusions: The pharmacist-led diabetes self-management program demonstrated significant reductions in HbA1c, LDL-C, and MAP from baseline to one year of program participation. Improvements were statistically significant and clinically relevant for each parameter. Previous studies indicate these reductions may cause reduced overall healthcare costs. PMID:25580174

Johnson, Courtney; Ruisinger, Janelle F.; Bates, Jessica; Barnes., Brian J.

2014-01-01

194

Mannose-6-Phosphate Reductase, a Key Enzyme in Photoassimilate Partitioning, Is Abundant and Located in the Cytosol of Photosynthetically Active Cells of Celery (Apium graveolens L.) Source Leaves.  

PubMed Central

Mannitol, a major photosynthetic product and transport carbohydrate in many plants, accounts for approximately 50% of the carbon fixed by celery (Apium graveolens L.) leaves. Previous subfractionation studies of celery leaves indicated that the enzymes for mannitol synthesis were located in the cytosol, but these data are inconsistent with that published for the sites of sugar alcohol synthesis in other families and taxa, including apple (Malus) and a brown alga (Fucus). Using antibodies to a key synthetic enzyme, NADPH-dependent mannose-6-phosphate reductase (M6PR), and immunocytochemical techniques, we have resolved both the inter-cellular and intracellular sites of mannitol synthesis. In leaves, M6PR was found only in cells containing ribulose-1,5-bisphosphate carboxylase/oxygenase. M6PR was almost exclusively cytosolic in these cells, with the nucleus being the only organelle to show labeling. The key step in transport carbohydrate biosynthesis that is catalyzed by M6PR displays no apparent preferential association with vascular tissues or with the bundle sheath. These results show that M6PR and, thus, mannitol synthesis are closely associated with the distribution of photosynthetic carbon metabolism in celery leaves. The principal role of M6PR is, therefore, in the assimilation of carbon being exported from the chloroplast, and it seems unlikely that this enzyme plays even an indirect role in phloem loading of mannitol. PMID:12231825

Everard, J. D.; Franceschi, V. R.; Loescher, W. H.

1993-01-01

195

The Role of Intracellular Signaling in Insulin-mediated Regulation of Drug Metabolizing Enzyme Gene and Protein Expression  

PubMed Central

Endogenous factors, including hormones, growth factors and cytokines, play an important role in the regulation of hepatic drug metabolizing enzyme expression in both physiological and pathophysiological conditions. Alterations of hepatic drug metabolizing enzymes gene and protein expression, observed in diabetes, fasting, obesity, protein-calorie malnutrition and long-term alcohol consumption alters the metabolism of xenobiotics, including procarcinogens, carcinogens, toxicants, and therapeutic agents and may also impact the efficacy and safety of therapeutic agents, as well as result in drug-drug interactions. Although the mechanisms by which xenobiotics regulate drug metabolizing enzymes have been studied intensively, less is known regarding the cellular signaling pathways and components which regulate drug metabolizing enzyme gene and protein expression in response to hormones and cytokines. Recent findings, however, have revealed that several cellular signaling pathways are involved in hormone- and growth factor-mediated regulation of drug metabolizing enzymes. Our laboratory, and others, have demonstrated that insulin and growth factors regulate drug metabolizing enzyme gene and protein expression, including cytochromes P450, glutathione S-transferases and microsomal epoxide hydrolase, through receptors which are members of the large receptor tyrosine kinase family, and by downstream effectors such as phosphatidylinositol 3-kinase, the mitogen activated protein kinase, Akt/protein kinase B, mTOR, and the p70S6 kinase. Here, we review current knowledge of the signaling pathways implicated in regulation of drug metabolizing enzyme gene and protein expression in response to insulin and growth factors, with the goal of increasing our understanding of how chronic disease affects these signaling pathways, components, and ultimately gene expression and translational control. PMID:17097148

Kim, Sang K.; Novak, Raymond F.

2007-01-01

196

Activity of hepatic drug metabolizing enzymes following oxazepam-dosed feed treatment in B6C3F1 mice  

Microsoft Academic Search

Oxazepam has been determined to be a potent hepatocarcinogen in mice. Evidence in the literature indicates that oxazepam is capable of inducing drug metabolizing enzymes in rodents and an association between enzyme induction and carcinogenesis has been proposed for other compounds such as phenobarbital. We examined the pattern of enzyme induction that occurs under bioassay conditions in male B6C3F1 mice.

Robert J. Griffin; Leo T. Burka; Michael L. Cunningham

1995-01-01

197

Identification of Multiple Phosphorylation Sites on Maize Endosperm Starch Branching Enzyme IIb, a Key Enzyme in Amylopectin Biosynthesis  

PubMed Central

Starch branching enzyme IIb (SBEIIb) plays a crucial role in amylopectin biosynthesis in maize endosperm by defining the structural and functional properties of storage starch and is regulated by protein phosphorylation. Native and recombinant maize SBEIIb were used as substrates for amyloplast protein kinases to identify phosphorylation sites on the protein. A multidisciplinary approach involving bioinformatics, site-directed mutagenesis, and mass spectrometry identified three phosphorylation sites at Ser residues: Ser649, Ser286, and Ser297. Two Ca2+-dependent protein kinase activities were partially purified from amyloplasts, termed K1, responsible for Ser649 and Ser286 phosphorylation, and K2, responsible for Ser649 and Ser297 phosphorylation. The Ser286 and Ser297 phosphorylation sites are conserved in all plant branching enzymes and are located at opposite openings of the 8-stranded parallel ?-barrel of the active site, which is involved with substrate binding and catalysis. Molecular dynamics simulation analysis indicates that phospho-Ser297 forms a stable salt bridge with Arg665, part of a conserved Cys-containing domain in plant branching enzymes. Ser649 conservation appears confined to the enzyme in cereals and is not universal, and is presumably associated with functions specific to seed storage. The implications of SBEIIb phosphorylation are considered in terms of the role of the enzyme and the importance of starch biosynthesis for yield and biotechnological application. PMID:24550386

Makhmoudova, Amina; Williams, Declan; Brewer, Dyanne; Massey, Sarah; Patterson, Jenelle; Silva, Anjali; Vassall, Kenrick A.; Liu, Fushan; Subedi, Sanjeena; Harauz, George; Siu, K. W. Michael; Tetlow, Ian J.; Emes, Michael J.

2014-01-01

198

Calcium-regulated nuclear enzymes: potential mediators of phytochrome-induced changes in nuclear metabolism?  

NASA Technical Reports Server (NTRS)

Calcium ions have been proposed to serve as important regulatory elements in stimulus-response coupling for phytochrome responses. An important test of this hypothesis will be to identify specific targets of calcium action that are required for some growth or development process induced by the photoactivated form of phytochrome (Pfr). Initial studies have revealed that there are at least two enzymes in pea nuclei that are stimulated by Pfr in a Ca(2+)-dependent fashion, a calmodulin-regulated nucleoside triphosphatase and a calmodulin-independent but Ca(2+)-dependent protein kinase. The nucleoside triphosphatase appears to be associated with the nuclear envelope, while the protein kinase co-purifies with a nuclear fraction highly enriched for chromatin. This short review summarizes the latest findings on these enzymes and relates them to what is known about Pfr-regulated nuclear metabolism.

Roux, S. J.

1992-01-01

199

In vivo cytochrome P450 drug metabolizing enzyme characterization using surface-enhanced Raman spectroscopy  

NASA Astrophysics Data System (ADS)

The development of a rapid, inexpensive, and accurate in vivo phenotyping methodology for characterizing drug-metabolizing phenotypes with reference to the cytochrome P450 (CYP450) enzymes would be very beneficial. In terms of application, in the wake of the human genome project, considerable interest is focused on the development of new drugs whose uses will be tailored to specific genetic polymorphisms, and on the individualization of dosing regimens that are also tailored to meet individual patient needs depending upon genotype. In this investigation, chemical probes for CYP450 enzymes were characterized and identified with Raman spectroscopy. Furthermore, gold-based metal colloid clusters were utilized to generate surface enhanced Raman spectra for each of the chemical probes. Results will be presented demonstrating the ability of SERS to identify minute quantities of these probes on the order needed for in vivo application.

Li, Yanfang; Bachmann, Kenneth A.; Cameron, Brent D.

2003-07-01

200

Parvoviruses Cause Nuclear Envelope Breakdown by Activating Key Enzymes of Mitosis  

PubMed Central

Disassembly of the nuclear lamina is essential in mitosis and apoptosis requiring multiple coordinated enzymatic activities in nucleus and cytoplasm. Activation and coordination of the different activities is poorly understood and moreover complicated as some factors translocate between cytoplasm and nucleus in preparatory phases. Here we used the ability of parvoviruses to induce nuclear membrane breakdown to understand the triggers of key mitotic enzymes. Nuclear envelope disintegration was shown upon infection, microinjection but also upon their application to permeabilized cells. The latter technique also showed that nuclear envelope disintegration was independent upon soluble cytoplasmic factors. Using time-lapse microscopy, we observed that nuclear disassembly exhibited mitosis-like kinetics and occurred suddenly, implying a catastrophic event irrespective of cell- or type of parvovirus used. Analyzing the order of the processes allowed us to propose a model starting with direct binding of parvoviruses to distinct proteins of the nuclear pore causing structural rearrangement of the parvoviruses. The resulting exposure of domains comprising amphipathic helices was required for nuclear envelope disintegration, which comprised disruption of inner and outer nuclear membrane as shown by electron microscopy. Consistent with Ca++ efflux from the lumen between inner and outer nuclear membrane we found that Ca++ was essential for nuclear disassembly by activating PKC. PKC activation then triggered activation of cdk-2, which became further activated by caspase-3. Collectively our study shows a unique interaction of a virus with the nuclear envelope, provides evidence that a nuclear pool of executing enzymes is sufficient for nuclear disassembly in quiescent cells, and demonstrates that nuclear disassembly can be uncoupled from initial phases of mitosis. PMID:24204256

Porwal, Manvi; Cohen, Sarah; Snoussi, Kenza; Popa-Wagner, Ruth; Anderson, Fenja; Dugot-Senant, Nathalie; Wodrich, Harald; Dinsart, Christiane; Kleinschmidt, Jürgen A.; Panté, Nelly; Kann, Michael

2013-01-01

201

Identification of a Metabolizing Enzyme in Human Kidney by Proteomic Correlation Profiling  

PubMed Central

Molecular identification of endogenous enzymes and biologically active substances from complex biological sources remains a challenging task, and although traditional biochemical purification is sometimes regarded as outdated, it remains one of the most powerful methodologies for this purpose. While biochemical purification usually requires large amounts of starting material and many separation steps, we developed an advanced method named “proteomic correlation profiling” in our previous study. In proteomic correlation profiling, we first fractionated biological material by column chromatography, and then calculated each protein's correlation coefficient between the enzyme activity profile and protein abundance profile determined by proteomics technology toward fractions. Thereafter, we could choose possible candidates for the enzyme among proteins with a high correlation value by domain predictions using informatics tools. Ultimately, this streamlined procedure requires fewer purification steps and reduces starting materials dramatically due to low required purity compared with conventional approaches. To demonstrate the generality of this approach, we have now applied an improved workflow of proteomic correlation profiling to a drug metabolizing enzyme and successfully identified alkaline phosphatase, tissue-nonspecific isozyme (ALPL) as a phosphatase of CS-0777 phosphate (CS-0777-P), a selective sphingosine 1-phosphate receptor 1 modulator with potential benefits in the treatment of autoimmune diseases including multiple sclerosis, from human kidney extract. We identified ALPL as a candidate protein only by the 200-fold purification and only from 1 g of human kidney. The identification of ALPL as CS-0777-P phosphatase was strongly supported by a recombinant protein, and contribution of the enzyme in human kidney extract was validated by immunodepletion and a specific inhibitor. This approach can be applied to any kind of enzyme class and biologically active substance; therefore, we believe that we have provided a fast and practical option by combination of traditional biochemistry and state-of-the-art proteomic technology. PMID:23674616

Sakurai, Hidetaka; Kubota, Kazuishi; Inaba, Shin-ichi; Takanaka, Kaoru; Shinagawa, Akira

2013-01-01

202

Metabolism of (+)-terpinen-4-ol by cytochrome P450 enzymes in human liver microsomes.  

PubMed

We examined the in vitro metabolism of (+)-terpinen-4-ol by human liver microsomes and recombinant enzymes. The biotransformation of (+)-terpinen-4-ol was investigated by gas chromatography-mass spectrometry (GC-MS). (+)-Terpinen-4-ol was found to be oxidized to (+)-(1R,2S,4S)-1,2-epoxy-p-menthan-4-ol, (+)-(1S,2R,4S)-1,2-epoxy-p-menthan-4-ol, and (4S)-p-menth-1-en-4,8-diol by human liver microsomal P450 enzymes. The identities of (+)-terpinen-4-ol metabolites were determined through the relative abundance of mass fragments and retention times on GC-MS. Of 11 recombinant human P450 enzymes tested, CYP1A2, CYP2A6, and CYP3A4 were found to catalyze the oxidation of (+)-terpinen-4-ol. Based on several lines of evidence, CYP2A6 and CYP3A4 were determined to be major enzymes involved in the oxidation of (+)-terpinen-4-ol by human liver microsomes. First, of the 11 recombinant human P450 enzymes tested, CYP1A2, CYP2A6 and CYP3A4 catalyzed oxidation of (+)-terpinen-4-ol. Second, oxidation of (+)-terpinen-4-ol was inhibited by (+)-menthofuran and ketoconazole, inhibitors known to be specific for these enzymes. Finally, there was a good correlation between CYP2A6 and CYP3A4 activities and (+)-terpinen-4-ol oxidation activities in the 10 human liver microsomes. PMID:22188805

Haigou, Risa; Miyazawa, Mitsuo

2012-01-01

203

Structure of glycerol-3-phosphate dehydrogenase, an essential monotopic membrane enzyme involved in respiration and metabolism  

SciTech Connect

Sn-glycerol-3-phosphate dehydrogenase (GlpD) is an essential membrane enzyme, functioning at the central junction of respiration, glycolysis, and phospholipid biosynthesis. Its critical role is indicated by the multitiered regulatory mechanisms that stringently controls its expression and function. Once expressed, GlpD activity is regulated through lipid-enzyme interactions in Escherichia coli. Here, we report seven previously undescribed structures of the fully active E. coli GlpD, up to 1.75 {angstrom} resolution. In addition to elucidating the structure of the native enzyme, we have determined the structures of GlpD complexed with substrate analogues phosphoenolpyruvate, glyceric acid 2-phosphate, glyceraldehyde-3-phosphate, and product, dihydroxyacetone phosphate. These structural results reveal conformational states of the enzyme, delineating the residues involved in substrate binding and catalysis at the glycerol-3-phosphate site. Two probable mechanisms for catalyzing the dehydrogenation of glycerol-3-phosphate are envisioned, based on the conformational states of the complexes. To further correlate catalytic dehydrogenation to respiration, we have additionally determined the structures of GlpD bound with ubiquinone analogues menadione and 2-n-heptyl-4-hydroxyquinoline N-oxide, identifying a hydrophobic plateau that is likely the ubiquinone-binding site. These structures illuminate probable mechanisms of catalysis and suggest how GlpD shuttles electrons into the respiratory pathway. Glycerol metabolism has been implicated in insulin signaling and perturbations in glycerol uptake and catabolism are linked to obesity in humans. Homologs of GlpD are found in practically all organisms, from prokaryotes to humans, with >45% consensus protein sequences, signifying that these structural results on the prokaryotic enzyme may be readily applied to the eukaryotic GlpD enzymes.

Yeh, Joanne I.; Chinte, Unmesh; Du, Shoucheng (Pitt)

2008-04-02

204

[Hypotensive, organoprotective, and metabolic effects of Angiotensin converting enzyme inhibitor moexipril in women with postmenopausal syndrome].  

PubMed

Hypotensive, organoprotective, and metabolic effects of angiotensin converting enzyme inhibitor moexipril (7.5-15 mg/day for 16 weeks) with or without combination with hydrochlorothiazide was studied in 34 women (mean age 59.6+/-1.6 years) with postmenopausal metabolic syndrome and hypertension. Thirty four women had dyslipidemia, 22 -- disturbances of carbohydrate metabolism, 18 -- obesity (mean body mass index 31.1+/-0.8 kg/m(2)). Treatment was associated with lowering of office systolic(-20.1%) and diastolic (-17.4%) blood pressure (BP). Target BP (140/90 mm Hg) was achieved in 27 patients. There also occurred significant lowering of mean 24 hour, diurnal, and nocturnal systolic and diastolic BP (p<0.05), significant changes of values of systolic and diastolic BP time indexes, normalization of which was observed both during day and night hours. Significant lowering of total cholesterol (-11.6%, p<0.05), low density lipoprotein cholesterol (-16.3%, p<0.02), and in patients with obesity of triglycerides (-27%, p<0.02) was revealed at the background of treatment with moexipril. In a group as a whole we observed significant lowering of excretion of albumins and b2-microglobulin; most pronounced antiproteinuretic effect was noted in patients with high microproteinuria and obesity. Vasodilating function of vessels improved in all patients with postmenopausal metabolic syndrome, mainly at the account of increment of endothelium dependent vasodilation and normalization of index of vasodilatation. PMID:16858353

Leonova, M V; Demidova, M A; Tarasov, A V; Belousov, Iu B

2006-01-01

205

Modeling of sensing potency of cytoskeletal systems decorated with metabolic enzymes.  

PubMed

The highly dynamic cytoskeleton interacts with enzymes and other proteins that are involved in metabolic or signaling pathways. These interactions can influence the structural and functional characteristics of the partners at the microscopic level of individual proteins and polymers. In this work the functional consequences of such interactions have been studied at the macroscopic level in order to evaluate the integrative and regulatory roles of the metabolic pathways associated with the microtubule cytoskeleton. Here we present mathematical models of the interactions between a hypothetical metabolic pathway and microtubule assembly, and explore for the first time the functional consequences of these interactions in distinct situations. The models include kinetic constants of the individual steps and testable, relevant parameters which allow the quantification of the coupled processes at the microscopic and macroscopic levels. For example our kinetic model for the self-assembly of microtubules reproduces the alteration of the time-dependent turbidity caused by pyruvate kinase binding. Our data reveal the power of a mechanistic description of a filamentous system to explain how cells sense the state of metabolic and other pathways. PMID:25451961

Oláh, Judit; Norris, Vic; Ovádi, Judit

2015-01-21

206

Multiple roles for lipins\\/phosphatidate phosphatase enzymes in lipid metabolism  

Microsoft Academic Search

Phosphatidate phosphatase-1 (PAP1) enzymes have a key role in glycerolipid synthesis through the con- version of phosphatidate to diacylglycerol, the immediate precursor of triacylglycerol, phosphatidylcholine, and phos- phatidylethanolamine. PAP1 activity in mammals is deter- mined by the lipin family of proteins, lipin-1, lipin-2, and lipin-3, which each have distinct tissue expression patterns and appear to have unique physiological functions. In

Karen Reue; David N. Brindley

207

Novel Mutation Causing Derepression of Several Enzymes of Sulfur Metabolism in Neurospora crassa  

PubMed Central

A group of enzymes of sulfur metabolism (arylsulfatase, cholinesulfatase, and a number of others) are normally repressed in Neurospora crassa by an abundant supply of a “favored” sulfur source such as methionine or inorganic sulfate. A mutant called sconc was isolated in which the formation of each of these enzymes is largely or completely nonrepressible. The structural genes for three of these enzymes have been mapped; sconc is not linked to any of them. It is also not linked to cys-3, another gene which is involved in control of the same group of enzymes. Two alleles of the structural gene for arylsulfatase [ars+ and ars(UFC-220)] produce electrophoretically distinguishable forms of arylsulfatase. Heterokaryons with the constitution sconc ars+ + scon+ars(UFC-220) were prepared. These heterokaryons produce both forms of arylsulfatase under conditions of sulfur limitation, but produce only the wild-type (ars+) form under conditions of sulfur abundance. When the alleles of ars and scon are in the opposite relationship, only the ars(UFC-220) form of arylsulfatase can be detected under conditions of sulfur abundance. Thus the effect of the sconc mutation seems to be limited to its own nucleus. The implications of these findings are discussed. Images PMID:4257980

Burton, Earl G.; Metzenberg, Robert L.

1972-01-01

208

Inflammation-induced phenoconversion of polymorphic drug metabolizing enzymes: hypothesis with implications for personalized medicine.  

PubMed

Phenoconversion transiently converts genotypic extensive metabolizers (EMs) into phenotypic poor metabolizers (PMs) of drugs, potentially with corresponding changes in clinical response. This phenomenon, typically resulting from coadministration of medications that inhibit certain drug metabolizing enzymes (DMEs), is especially well documented for enzymes of the cytochrome P450 family. Nonclinical evidence gathered over the last two decades also strongly implicates elevated levels of some proinflammatory cytokines, released during inflammation, in down-regulation of drug metabolism, especially by certain DMEs of the P450 family, thereby potentially causing transient phenoconversion. Clinically, phenoconversion of NAT2, CYP2C19, and CYP2D6 has been documented in inflammatory conditions associated with elevated cytokines, such as human immunodeficiency virus infection, cancer, and liver disease. The potential of other inflammatory conditions to cause phenoconversion has not been studied but experimental and anecdotal clinical evidence supports infection-induced down-regulation of CYP1A2, CYP3A4, and CYP2C9 as well. Collectively, the evidence supports a hypothesis that certain inflammatory conditions associated with elevated proinflammatory cytokines may cause phenoconversion of certain DMEs. Since inflammatory conditions associated with elevated levels of proinflammatory cytokines are highly prevalent, phenoconversion of genotypic EM patients into transient phenotypic PMs may be more frequent than appreciated. Since drug pharmacokinetics, and therefore the clinical response, is influenced by DME phenotype rather than genotype per se, phenoconversion (whatever its cause) can have a significant impact on the analysis and interpretation of genotype-focused clinical outcome association studies. There is a risk that focusing on genotype alone may miss important associations between clinical outcomes and DME phenotypes, thus compromising future prospects of personalized medicine. PMID:25519488

Shah, Rashmi R; Smith, Robert L

2015-03-01

209

[Enzyme deficiencies in glycolysis and nucleotide metabolism of red blood cells in nonspherocytic hemolytic anemia (author's transl)].  

PubMed

The detection of enzyme deficiencies in glycolytic and nucleotide metabolism of human red blood cells has enriched the pathophysiological knowledge on the origin of nonspherocytic hemolytic anemias (NSHA). So far for 11 of 13 glycolytic enzymes deficiencies have been described which are connected with alterations of biochemical enzymatic properties. The most frequent enzyme deficiencies are those of GPI and PK. By performance of special electrophoretic techniques genetic studies allow the demonstration of homozygote and double heterozygote defect carriers. Up to now only adenylate kinase and pyrimidine 5' nucleotidase deficiencies have been detected as genetically determined in altered nucleotide metabolism. The metabolic alterations of several enzymopathies have been characterized so well, that the pathophysiological relations between enzyme deficiency and NSHA probably have been found to be a sufficient explanation. PMID:184346

Waller, H D; Benöhr, H C

1976-09-01

210

Identification of metabolic pathways and enzyme systems involved in the in vitro human hepatic metabolism of dronedarone, a potent new oral antiarrhythmic drug  

PubMed Central

The in vitro metabolism of dronedarone and its major metabolites has been studied in human liver microsomes and cryopreserved hepatocytes in primary culture through the use of specific or total cytochrome P450 (CYP) and monoamine oxidase (MAO) inhibitors. The identification of the main metabolites and enzymes participating in their metabolism was also elucidated by using rhCYP, rhMAO, flavin monooxygenases (rhFMO) and UDP-glucuronosyltransferases (rhUGT) and liquid chromatography/tandem mass spectrometry (LC/MS-MS) analysis. Dronedarone was extensively metabolized in human hepatocytes with a metabolic clearance being almost completely inhibited (98 ± 2%) by 1-aminobenzotriazole. Ketoconazole also inhibited dronedarone metabolism by 89 ± 7%, demonstrating the crucial role of CYP3A in its metabolism. CYP3A isoforms mostly contributed to N-debutylation while hydroxylation on the butyl-benzofuran moiety was catalyzed by CYP2D6. However, hydroxylation on the dibutylamine moiety did not appear to be CYP-dependent. N-debutyl-dronedarone was less rapidly metabolized than dronedarone, the major metabolic pathway being catalyzed by MAO-A to form propanoic acid-dronedarone and phenol-dronedarone. Propanoic acid-dronedarone was metabolized at a similar rate to that of N-debutyl-dronedarone and was predominantly hydroxylated by CYP2C8 and CYP1A1. Phenol-dronedarone was extensively glucuronidated while C-dealkyl-dronedarone was metabolized at a slow rate. The evaluation of the systemic clearance of each metabolic process together with the identification of both the major metabolites and predominant enzyme systems and isoforms involved in the formation and subsequent metabolism of these metabolites has enhanced the overall understanding of metabolism of dronedarone in humans. PMID:25505590

Klieber, Sylvie; Arabeyre-Fabre, Catherine; Moliner, Patricia; Marti, Eric; Mandray, Martine; Ngo, Robert; Ollier, Céline; Brun, Priscilla; Fabre, Gérard

2014-01-01

211

Identification of metabolic pathways and enzyme systems involved in the in vitro human hepatic metabolism of dronedarone, a potent new oral antiarrhythmic drug.  

PubMed

The in vitro metabolism of dronedarone and its major metabolites has been studied in human liver microsomes and cryopreserved hepatocytes in primary culture through the use of specific or total cytochrome P450 (CYP) and monoamine oxidase (MAO) inhibitors. The identification of the main metabolites and enzymes participating in their metabolism was also elucidated by using rhCYP, rhMAO, flavin monooxygenases (rhFMO) and UDP-glucuronosyltransferases (rhUGT) and liquid chromatography/tandem mass spectrometry (LC/MS-MS) analysis. Dronedarone was extensively metabolized in human hepatocytes with a metabolic clearance being almost completely inhibited (98 ± 2%) by 1-aminobenzotriazole. Ketoconazole also inhibited dronedarone metabolism by 89 ± 7%, demonstrating the crucial role of CYP3A in its metabolism. CYP3A isoforms mostly contributed to N-debutylation while hydroxylation on the butyl-benzofuran moiety was catalyzed by CYP2D6. However, hydroxylation on the dibutylamine moiety did not appear to be CYP-dependent. N-debutyl-dronedarone was less rapidly metabolized than dronedarone, the major metabolic pathway being catalyzed by MAO-A to form propanoic acid-dronedarone and phenol-dronedarone. Propanoic acid-dronedarone was metabolized at a similar rate to that of N-debutyl-dronedarone and was predominantly hydroxylated by CYP2C8 and CYP1A1. Phenol-dronedarone was extensively glucuronidated while C-dealkyl-dronedarone was metabolized at a slow rate. The evaluation of the systemic clearance of each metabolic process together with the identification of both the major metabolites and predominant enzyme systems and isoforms involved in the formation and subsequent metabolism of these metabolites has enhanced the overall understanding of metabolism of dronedarone in humans. PMID:25505590

Klieber, Sylvie; Arabeyre-Fabre, Catherine; Moliner, Patricia; Marti, Eric; Mandray, Martine; Ngo, Robert; Ollier, Céline; Brun, Priscilla; Fabre, Gérard

2014-06-01

212

Effects of Naturally Occurring Coumarins on Hepatic Drug Metabolizing Enzymes in Mice  

PubMed Central

Cytochromes P450 (P450s) and glutathione S-transferases (GSTs) constitute two important enzyme families involved in carcinogen metabolism. Generally, P450s play activation or detoxifying roles while GSTs act primarily as detoxifying enzymes. We previously demonstrated that oral administration of the linear furanocoumarins, isopimpinellin and imperatorin, modulated P450 and GST activities in various tissues of mice. The purpose of the present study was to compare a broader range of naturally occurring coumarins (simple coumarins, and furanocoumarins of the linear and angular type) for their abilities to modulate hepatic drug metabolizing enzymes when administered orally to mice. We now report that all of the different coumarins tested (coumarin, limettin, auraptene, angelicin, bergamottin, imperatorin and isopimpinellin) induced hepatic GST activities, whereas the linear furanocoumarins possessed the greatest abilities to induce hepatic P450 activities, in particular P450 2B and 3A. In both cases, this corresponded to an increase in protein expression of the enzymes. Induction of P4502B10, 3A11, and 2C9 by xenobiotics often are a result of activation of the pregnane X receptor (PXR) and/or constitutive androstane receptor (CAR). Using a pregnane X receptor reporter system, our results demonstrated that isopimpinellin activated both PXR and its human ortholog SXR by recruiting coactivator SRC-1 in transfected cells. In CAR transfection assays, isopimpinellin counteracted the inhibitory effect of androstanol on full length mCAR, a Gal4-mCAR ligand binding domain fusion, and restored coactivator binding. Orally administered isopimpinellin induced hepatic mRNA expression of Cyp2b10,Cyp3a1, GSTa in CAR(+/+) wild-type mice. In contrast, the induction of Cyp2b10 mRNA by isopimpinellin was attenuated in the CAR(?/?) mice, suggesting that isopimpinellin induces Cyp2b10 via the CAR receptor. Overall, the current data indicate that naturally occurring coumarins have diverse activities in terms of inducing various xenobiotic metabolizing enzymes based on their chemical structure. PMID:18692084

Kleiner, Heather E.; Xia, Xiojun; Sonoda, Junichiro; Zhang, Jun; Pontius, Elizabeth; Abey, Jane; Evans, Ronald M.; Moore, David D.; DiGiovanni., John

2008-01-01

213

Effects of naturally occurring coumarins on hepatic drug-metabolizing enzymes inmice  

SciTech Connect

Cytochromes P450 (P450s) and glutathione S-transferases (GSTs) constitute two important enzyme families involved in carcinogen metabolism. Generally, P450s play activation or detoxifying roles while GSTs act primarily as detoxifying enzymes. We previously demonstrated that oral administration of the linear furanocoumarins, isopimpinellin and imperatorin, modulated P450 and GST activities in various tissues of mice. The purpose of the present study was to compare a broader range of naturally occurring coumarins (simple coumarins, and furanocoumarins of the linear and angular type) for their abilities to modulate hepatic drug-metabolizing enzymes when administered orally to mice. We now report that all of the different coumarins tested (coumarin, limettin, auraptene, angelicin, bergamottin, imperatorin and isopimpinellin) induced hepatic GST activities, whereas the linear furanocoumarins possessed the greatest abilities to induce hepatic P450 activities, in particular P450 2B and 3A. In both cases, this corresponded to an increase in protein expression of the enzymes. Induction of P4502B10, 3A11, and 2C9 by xenobiotics often is a result of activation of the pregnane X receptor (PXR) and/or constitutive androstane receptor (CAR). Using a pregnane X receptor reporter system, our results demonstrated that isopimpinellin activated both PXR and its human ortholog SXR by recruiting coactivator SRC-1 in transfected cells. In CAR transfection assays, isopimpinellin counteracted the inhibitory effect of androstanol on full-length mCAR, a Gal4-mCAR ligand-binding domain fusion, and restored coactivator binding. Orally administered isopimpinellin induced hepatic mRNA expression of Cyp2b10, Cyp3a11, and GSTa in CAR(+/+) wild-type mice. In contrast, the induction of Cyp2b10 mRNA by isopimpinellin was attenuated in the CAR(-/-) mice, suggesting that isopimpinellin induces Cyp2b10 via the CAR receptor. Overall, the current data indicate that naturally occurring coumarins have diverse activities in terms of inducing various xenobiotic metabolizing enzymes based on their chemical structure.

Kleiner, Heather E. [Department of Carcinogenesis, University of Texas M.D. Anderson Cancer Center, Science Park-Research Division, Park Road 1-C, Smithville, TX 78957 (United States)], E-mail: hklein@lsuhsc.edu; Xia, Xiaojun; Sonoda, Junichiro [Howard Hughes Medical Institute, Gene Expression Laboratory, Salk Institute for Biological Sciences, La Jolla, CA 92037 (United States); Zhang, Jun [Department of Molecular and Cellular Biology, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030 (United States); Pontius, Elizabeth; Abey, Jane [Department of Carcinogenesis, University of Texas M.D. Anderson Cancer Center, Science Park-Research Division, Park Road 1-C, Smithville, TX 78957 (United States); Evans, Ronald M. [Howard Hughes Medical Institute, Gene Expression Laboratory, Salk Institute for Biological Sciences, La Jolla, CA 92037 (United States); Moore, David D. [Department of Molecular and Cellular Biology, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030 (United States); DiGiovanni, John [Department of Carcinogenesis, University of Texas M.D. Anderson Cancer Center, Science Park-Research Division, Park Road 1-C, Smithville, TX 78957 (United States)

2008-10-15

214

PAPER www.rsc.org/loc | Lab on a Chip Programmable assembly of a metabolic pathway enzyme in a pre-packaged  

E-print Network

PAPER www.rsc.org/loc | Lab on a Chip Programmable assembly of a metabolic pathway enzyme in a pre report a biofunctionalization strategy for the assembly of catalytically active enzymes within and temporally defined sites. The enzyme of a bacterial metabolic pathway, S-adenosylhomocysteine nucleosidase

Rubloff, Gary W.

215

Disaccharide-Mediated Regulation of Sucrose:Fructan-6-Fructosyltransferase, a Key Enzyme of Fructan Synthesis in Barley Leaves1  

PubMed Central

Previous work has indicated that sugar sensing may be important in the regulation of fructan biosynthesis in grasses. We used primary leaves of barley (Hordeum vulgare cv Baraka) to study the mechanisms involved. Excised leaf blades were supplied in the dark with various carbohydrates. Fructan pool sizes and two key enzymes of fructan biosynthesis, sucrose (Suc):Suc-1-fructosyltransferase (1-SST; EC 2.4.1.99) and Suc:fructan-6-fructosyltransferase (6-SFT; EC 2.4.1.10) were analyzed. Upon supply of Suc, fructan pool sizes increased markedly. Within 24 h, 1-SST activity was stimulated by a factor of three and 6-SFT-activity by a factor of more than 20, compared with control leaves supplemented with mannitol (Mit). At the same time, the level of mRNA encoding 6-SFT increased conspicuously. These effects were increased in the presence of the invertase inhibitor 2,5-dideoxy-2,5-imino-d-mannitol. Compared with equimolar solutions of Suc, glucose (Glu) and fructose stimulated 6-SFT activity to a lesser extent. Remarkably, trehalose (Tre; Glc-?-1 and 1-?-Glc) had stimulatory effects on 6-SFT activity and, to a somewhat lesser extent, on 6-SFT mRNA, even in the presence of validoxylamine A, a potent trehalase inhibitor. Tre by itself, however, in the presence or absence of validoxylamine A, did not stimulate fructan accumulation. Monosaccharides phosphorylated by hexokinase but not or weakly metabolized, such as mannose (Man) or 2-deoxy-Glc, had no stimulatory effects on fructan synthesis. When fructose or Man were supplied together with Tre, fructan and starch biosynthesis were strongly stimulated. Concomitantly, phospho-Man isomerase (EC 5.3.1.8) activity was detected. These results indicate that the regulation of fructan synthesis in barley leaves occurs independently of hexokinase and is probably based on the sensing of Suc, and also that the structurally related disaccharide Tre can replace Suc as a regulatory compound. PMID:10806243

Müller, Joachim; Aeschbacher, Roger A.; Sprenger, Norbert; Boller, Thomas; Wiemken, Andres

2000-01-01

216

Activity of enzymes of carbon metabolism during the induction of Crassulacean acid metabolism in Mesembryanthemum crystallinum L.  

PubMed

The maximum extractable activities of twenty-one photosynthetic and glycolytic enzymes were measured in mature leaves of Mesembryanthemum crystallinum plants, grown under a 12 h light 12 h dark photoperiod, exhibiting photosynthetic characteristics of either a C3 or a Crassulacean acid metabolism (CAM) plant. Following the change from C3 photosynthesis to CAM in response to an increase in the salinity of in the rooting medium from 100 mM to 400 mM NaCl, the activity of phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) increased about 45-fold and the activities of NADP malic enzyme (EC 1.1.1.40) and NAD malic enzyme (EC 1.1.1.38) increased about 4- to 10-fold. Pyruvate, Pi dikinase (EC 2.7.9.1) was not detected in the non-CAM tissue but was present in the CAM tissue; PEP carboxykinase (EC 4.1.1.32) was detected in neither tissue. The induction of CAM was also accompanied by large increases in the activities of the glycolytic enzymes enolase (EC 4.2.1.11), phosphoglyceromutase (EC 2.7.5.3), phosphoglycerate kinase (EC 2.7.2.3), NAD glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), and glucosephosphate isomerase (EC 2.6.1.2). There were 1.5- to 2-fold increases in the activities of NAD malate dehydrogenase (EC 1.1.1.37), alanine and aspartate aminotransferases (EC 2.6.1.2 and 2.6.1.1 respectively) and NADP glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13). The activities of ribulose-1,5-bisphosphate (RuBP) carboxylase (EC 4.1.1.39), fructose-1,6-bisphosphatase (EC 3.1.3.11), phosphofructokinase (EC 2.7.1.11), hexokinase (EC 2.7.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) remained relatively constant. NADP malate dehydrogenase (EC 1.1.1.82) activity exhibited two pH optima in the non-CAM tissue, one at pH 6.0 and a second at pH 8.0. The activity at pH 8.0 increased as CAM was induced. With the exceptions of hexokinase and glucose-6-phosphate dehydrogenase, the activities of all enzymes examined in extracts from M. crystallinum exhibiting CAM were equal to, or greater than, those required to sustain the maximum rates of carbon flow during acidification and deacidification observed in vivo. There was no day-night variation in the maximum extractable activities of phosphoenolpyruvate carboxylase, NADP malic enzyme, NAD malic enzyme, fructose-1,6-bisphosphatase and NADP malate dehydrogenase in leaves of M. crystallinum undergoing CAM. PMID:24271620

Holtum, J A; Winter, K

1982-06-01

217

Metabolic syndrome and mean platelet volume variation in patients with chest pain and negative cardiac enzymes  

PubMed Central

Introduction. The mean platelet volume (MPV) is an easily measurable parameter directly correlated with platelet aggregation function, proven to be increased in acute coronary syndromes, but also in the presence of cardiovascular risk factors such as the metabolic syndrome, dyslipidemia, diabetes mellitus, arterial hypertension. Objective. This study intended to assess the role of the metabolic syndrome in MPV variation in patients presenting with chest pain. Materials and Methods. We retrospectively analyzed data from 122 patients with chest pain and negative cardiac enzymes admitted consecutively to our clinic from September 1st 2011 to January 30th 2012. Our group included 27 (22.13%) patients with stable angina (SA), 74 (60.65%) patients with unstable angina (UA) and 21 (17.22%) patients with non-coronary chest pain. Results. Patients with UA had a higher mean value of the MPV 9.31 ± 1.19 fL compared to patients with SA 8.72 ± 1.14 fL (p=0.0279) and patients with non-coronary chest pain 8.85 ± 0.90 L (p=0.0908). All the patients with metabolic syndrome had increased MPVs, regardless of the etiology of chest pain. Patients with non-coronary chest pain presented significantly higher MPVs if associated with metabolic syndrome or arterial hypertension. Conclusions. Patients with cardiovascular risk factors, especially complex ones like the metabolic syndrome had an increased MPV, as did the patients with UA whether or not associated with the risk factors. In patients without such comorbidities, the MPV could be useful in distinguishing unstable angina from non-coronary chest pain. PMID:23904875

Nechita, AC; Delcea, C; Enache, V; Ploesteanu, RL; Cazacu, C; Andronescu, AM; Stroi, AM; Stamate, CS

2013-01-01

218

Dual Targeting of Antioxidant and Metabolic Enzymes to the Mitochondrion and the Apicoplast of Toxoplasma gondii  

PubMed Central

Toxoplasma gondii is an aerobic protozoan parasite that possesses mitochondrial antioxidant enzymes to safely dispose of oxygen radicals generated by cellular respiration and metabolism. As with most Apicomplexans, it also harbors a chloroplast-like organelle, the apicoplast, which hosts various biosynthetic pathways and requires antioxidant protection. Most apicoplast-resident proteins are encoded in the nuclear genome and are targeted to the organelle via a bipartite N-terminal targeting sequence. We show here that two antioxidant enzymes—a superoxide dismutase (TgSOD2) and a thioredoxin-dependent peroxidase (TgTPX1/2)—and an aconitase are dually targeted to both the apicoplast and the mitochondrion of T. gondii. In the case of TgSOD2, our results indicate that a single gene product is bimodally targeted due to an inconspicuous variation within the putative signal peptide of the organellar protein, which significantly alters its subcellular localization. Dual organellar targeting of proteins might occur frequently in Apicomplexans to serve important biological functions such as antioxidant protection and carbon metabolism. PMID:17784785

Kwok, Lai-Yu; Sheiner, Lilach; Schepers, Rebecca; Soldati, Thierry; Soldati-Favre, Dominique

2007-01-01

219

Natural allelic variations of xenobiotic-metabolizing enzymes affect sexual dimorphism in Oryzias latipes.  

PubMed

Sexual dimorphisms, which are phenotypic differences between males and females, are driven by sexual selection. Interestingly, sexually selected traits show geographical variations within species despite strong directional selective pressures. This paradox has eluded many evolutionary biologists for some time, and several models have been proposed (e.g. 'indicator model' and 'trade-off model'). However, disentangling which of these theories explains empirical patterns remains difficult, because genetic polymorphisms that cause variation in sexual differences are still unknown. In this study, we show that polymorphisms in cytochrome P450 (CYP) 1B1, which encodes a xenobiotic-metabolizing enzyme, are associated with geographical differences in sexual dimorphism in the anal fin morphology of medaka fish (Oryzias latipes). Biochemical assays and genetic cross experiments show that high- and low-activity CYP1B1 alleles enhanced and declined sex differences in anal fin shapes, respectively. Behavioural and phylogenetic analyses suggest maintenance of the high-activity allele by sexual selection, whereas the low-activity allele possibly has experienced positive selection due to by-product effects of CYP1B1 in inferred ancestral populations. The present data can elucidate evolutionary mechanisms behind genetic variations in sexual dimorphism and indicate trade-off interactions between two distinct mechanisms acting on the two alleles with pleiotropic effects of xenobiotic-metabolizing enzymes. PMID:25377463

Katsumura, Takafumi; Oda, Shoji; Nakagome, Shigeki; Hanihara, Tsunehiko; Kataoka, Hiroshi; Mitani, Hiroshi; Kawamura, Shoji; Oota, Hiroki

2014-12-22

220

Role of aldo-keto reductase family 1 (AKR1) enzymes in human steroid metabolism.  

PubMed

Human aldo-keto reductases AKR1C1-AKR1C4 and AKR1D1 play essential roles in the metabolism of all steroid hormones, the biosynthesis of neurosteroids and bile acids, the metabolism of conjugated steroids, and synthetic therapeutic steroids. These enzymes catalyze NADPH dependent reductions at the C3, C5, C17 and C20 positions on the steroid nucleus and side-chain. AKR1C1-AKR1C4 act as 3-keto, 17-keto and 20-ketosteroid reductases to varying extents, while AKR1D1 acts as the sole ?(4)-3-ketosteroid-5?-reductase (steroid 5?-reductase) in humans. AKR1 enzymes control the concentrations of active ligands for nuclear receptors and control their ligand occupancy and trans-activation, they also regulate the amount of neurosteroids that can modulate the activity of GABAA and NMDA receptors. As such they are involved in the pre-receptor regulation of nuclear and membrane bound receptors. Altered expression of individual AKR1C genes is related to development of prostate, breast, and endometrial cancer. Mutations in AKR1C1 and AKR1C4 are responsible for sexual development dysgenesis and mutations in AKR1D1 are causative in bile-acid deficiency. PMID:24189185

Rižner, Tea Lanišnik; Penning, Trevor M

2014-01-01

221

Cytochrome P4502C9: an enzyme of major importance in human drug metabolism  

PubMed Central

Accumulating evidence indicates that CYP2C9 ranks amongst the most important drug metabolizing enzymes in humans. Substrates for CYP2C9 include fluoxetine, losartan, phenytoin, tolbutamide, torsemide, S-warfarin, and numerous NSAIDs. CYP2C9 activity in vivo is inducible by rifampicin. Evidence suggests that CYP2C9 substrates may also be induced variably by carbamazepine, ethanol and phenobarbitone. Apart from the mutual competitive inhibition which may occur between alternate substrates, numerous other drugs have been shown to inhibit CYP2C9 activity in vivo and/or in vitro. Clinically significant inhibition may occur with coadministration of amiodarone, fluconazole, phenylbutazone, sulphinpyrazone, sulphaphenazole and certain other sulphonamides. Polymorphisms in the coding region of the CYP2C9 gene produce variants at amino acid residues 144 (Arg144Cys) and 359 (Ile359Leu) of the CYP2C9 protein. Individuals homozygous for Leu359 have markedly diminished metabolic capacities for most CYP2C9 substrates, although the frequency of this allele is relatively low. Consistent with the modulation of enzyme activity by genetic and other factors, wide interindividual variability occurs in the elimination and/or dosage requirements of prototypic CYP2C9 substrates. Individualisation of dose is essential for those CYP2C9 substrates with a narrow therapeutic index. PMID:9663807

Miners, John O; Birkett, Donald J

1998-01-01

222

Effect of aging on drug-metabolizing enzymes important in acetaminophen elimination.  

PubMed

The effects of aging on selected drug-metabolizing enzyme activities, the pattern of phenol and bile salt sulfotransferase isoenzymes and the pharmacokinetics of acetaminophen were examined in male Fischer 344 rats at ages 5, 14 and 25 months. Aging decreased sulfotransferase activity toward acetaminophen while activity toward glycolithocholate increased with age. Glucuronosyltransferase activity toward estrone increased with age, while activity toward testosterone, morphine and naphthol remained constant. Glutathione-S-transferase (1-chloro-2,4-dinitrobenzene) activity was also unchanged through the various age groups. Cytochrome P-450 content and monooxygenase activity (p-nitroanisole demethylation) activity decreased with advancing age. Overall, the age-related in vitro changes in enzyme activities approached or equaled values measured in 5-month-old female Fischer 344 rats. Moreover, age-related alterations in total phenol sulfotransferase activity and the isozyme pattern paralleled changes in the in vivo elimination kinetics and metabolic fate of acetaminophen. The fraction of drug excreted as the sulfate conjugate and the partial clearance to acetaminophen sulfate decreased with increasing age. Conversely, the fraction excreted as the glucuronide and the partial clearance to acetaminophen glucuronide increased with increasing age. There was no effect of aging on the total clearance of acetaminophen. The gender-related differences in the pattern of sulfotransferase isozyme activity toward phenolic and bile salt acceptors disappeared with age. Age-related changes in sulfation and perhaps glucuronidation in male rats appear to feminize hepatic biotransformation and may arise due to altered gonadal hormone status. PMID:3083094

Galinsky, R E; Kane, R E; Franklin, M R

1986-04-01

223

Role of Aldo-Keto Reductase Family 1 (AKR1) Enzymes in Human Steroid Metabolism  

PubMed Central

Human aldo-keto reductases AKR1C1-AKR1C4 and AKR1D1 play essential roles in the metabolism of all steroid hormones, the biosynthesis of neurosteroids and bile acids, the metabolism of conjugated steroids, and synthetic therapeutic steroids. These enzymes catalyze NADPH dependent reductions at the C3, C5, C17 and C20 positions on the steroid nucleus and side-chain. AKR1C1-AKR1C4 act as 3-keto, 17-keto and 20-ketosteroid reductases to varying extents, while AKR1D1 acts as the sole ?4-3-ketosteroid-5?-reductase (steroid 5?-reductase) in humans. AKR1 enzymes control the concentrations of active ligands for nuclear receptors and control their ligand occupancy and trans-activation, they also regulate the amount of neurosteroids that can modulate the activity of GABAA and NMDA receptors. As such they are involved in the pre-receptor regulation of nuclear and membrane bound receptors. Altered expression of individual AKR1C genes is related to development of prostate, breast, and endometrial cancer. Mutations in AKR1C1 and AKR1C4 are responsible for sexual development dysgenesis and mutations in AKR1D1 are causative in bile-acid deficiency. PMID:24189185

Rižner, Tea Lanišnik; Penning, Trevor M.

2013-01-01

224

Metabolic engineering of Saccharomyces cerevisiae: a key cell factory platform for future biorefineries.  

PubMed

Metabolic engineering is the enabling science of development of efficient cell factories for the production of fuels, chemicals, pharmaceuticals, and food ingredients through microbial fermentations. The yeast Saccharomyces cerevisiae is a key cell factory already used for the production of a wide range of industrial products, and here we review ongoing work, particularly in industry, on using this organism for the production of butanol, which can be used as biofuel, and isoprenoids, which can find a wide range of applications including as pharmaceuticals and as biodiesel. We also look into how engineering of yeast can lead to improved uptake of sugars that are present in biomass hydrolyzates, and hereby allow for utilization of biomass as feedstock in the production of fuels and chemicals employing S. cerevisiae. Finally, we discuss the perspectives of how technologies from systems biology and synthetic biology can be used to advance metabolic engineering of yeast. PMID:22388689

Hong, Kuk-Ki; Nielsen, Jens

2012-08-01

225

The activity of drug-metabolizing enzymes and the biotransformation of selected anthelmintics in the model tapeworm Hymenolepis diminuta.  

PubMed

The drug-metabolizing enzymes of some helminths can deactivate anthelmintics and therefore partially protect helminths against these drugs' toxic effect. The aim of our study was to assess the activity of the main drug-metabolizing enzymes and evaluate the metabolism of selected anthelmintics (albendazole, flubendazole, mebendazole) in the rat tapeworm Hymenolepis diminuta, a species often used as a model tapeworm. In vitro and ex vivo experiments were performed. Metabolites of the anthelmintics were detected and identified by HPLC with spectrofluorometric or mass-spectrometric detection. The enzymes of H. diminuta are able to reduce the carbonyl group of flubendazole, mebendazole and several other xenobiotics. Although the activity of a number of oxidation enzymes was determined, no oxidative metabolites of albendazole were detected. Regarding conjugation enzymes, a high activity of glutathione S-transferase was observed. A methyl derivative of reduced flubendazole was the only conjugation metabolite identified in ex vivo incubations of H. diminuta with anthelmintics. The results revealed that H. diminuta metabolized flubendazole and mebendazole, but not albendazole. The biotransformation pathways found in H. diminuta differ from those described in Moniezia expanza and suggest the interspecies differences in drug metabolism not only among classes of helminths, but even among tapeworms. PMID:22309895

Bártíková, Hana; Vok?ál, Ivan; Skálová, Lenka; Kubí?ek, Vladimír; Firbasová, Jana; Briestenský, David; Lamka, Ji?í; Szotáková, Barbora

2012-05-01

226

Antioxidative Properties and Inhibition of Key Enzymes Relevant to Type-2 Diabetes and Hypertension by Essential Oils from Black Pepper  

PubMed Central

The antioxidant properties and effect of essential oil of black pepper (Piper guineense) seeds on ?-amylase, ?-glucosidase (key enzymes linked to type-2 diabetes), and angiotensin-I converting enzyme (ACE) (key enzyme linked to hypertension) were assessed. The essential oil was obtained by hydrodistillation and dried with anhydrous Na2SO4, and the phenolic content, radical [1,1-diphenyl-2 picrylhydrazyl (DPPH), 2,2?-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS) and nitric oxide (NO)] scavenging abilities as well as the ferric reducing antioxidant property (FRAP) and Fe2+-chelating ability of the essential oil were investigated. Furthermore, the effect on ?-amylase, ?-glucosidase, and ACE enzyme activities was also investigated. The characterization of the constituents was done using GC. The essential oil scavenged DPPH?, NO?, and ABTS? and chelated Fe2+. ?-Pinene, ?-pinene, cis-ocimene, myrcene, allo-ocimene, and 1,8-cineole were among the constituents identified by GC. The essential oil inhibited ?-amylase, ?-glucosidase, and ACE enzyme activities in concentration-dependent manners, though exhibiting a stronger inhibition of ?-glucosidase than ?-amylase activities. Conclusively, the phenolic content, antioxidant activity, and inhibition of ?-amylase, ?-glucosidase, and angiotensin-1 converting enzyme activities by the essential oil extract of black pepper could be part of the mechanism by which the essential oil could manage and/or prevent type-2 diabetes and hypertension. PMID:24348547

Oboh, Ganiyu; Ademosun, Ayokunle O.; Odubanjo, Oluwatoyin V.; Akinbola, Ifeoluwa A.

2013-01-01

227

Effect of urethane, dimethylnitrosamine, paraquat, and butylated hydroxytoluene on the activities of glycolytic key enzymes in mouse lung  

SciTech Connect

Effects of carcinogens and noncarcinogenic pulmonary toxicants on the activities of glycolytic key enzymes in the mouse lung were investigated. The carcinogens urethane (URTH) and dimethylnitrosamine (DMN) permanently enhanced, and the noncarcinogenic pulmonary toxicants paraquat (PAR) and butylated hydroxytoluene (BHT) temporarily, enhanced the activities of hexokinase (HK), phosphofructokinase (PFK), and pyruvate kinase (PK) in the lungs of mice.

Arany, I.; Rady, P.; Bojan, I.; Kertai, P.

1981-12-01

228

Evaluation of Indian sacred tree Crataeva magna (Lour.) DC. for antioxidant activity and inhibition of key enzymes relevant to hyperglycemia.  

PubMed

In the present investigation, leaf and stem bark of Crataeva magna are evaluated for their antioxidant activity and inhibition of key enzymes relevant to hyperglycemia. Both the parts exhibited significant antioxidant and anti-?-glucosidase activity. The results will lead in favor of the use of this plant as a potential additive/nutraceutical antioxidant compound. PMID:22196938

Loganayaki, Nataraj; Manian, Sellamuthu

2012-03-01

229

Correlating Structure and Function of Drug-Metabolizing Enzymes: Progress and Ongoing Challenges  

PubMed Central

This report summarizes a symposium sponsored by the American Society for Pharmacology and Experimental Therapeutics at Experimental Biology held April 20-24 in Boston, MA. Presentations discussed the status of cytochrome P450 (P450) knowledge, emphasizing advances and challenges in relating structure with function and in applying this information to drug design. First, at least one structure of most major human drug-metabolizing P450 enzymes is known. However, the flexibility of these active sites can limit the predictive value of one structure for other ligands. A second limitation is our coarse-grain understanding of P450 interactions with membranes, other P450 enzymes, NADPH–cytochrome P450 reductase, and cytochrome b5. Recent work has examined differential P450 interactions with reductase in mixed P450 systems and P450:P450 complexes in reconstituted systems and cells, suggesting another level of functional control. In addition, protein nuclear magnetic resonance is a new approach to probe these protein/protein interactions, identifying interacting b5 and P450 surfaces, showing that b5 and reductase binding are mutually exclusive, and demonstrating ligand modulation of CYP17A1/b5 interactions. One desired outcome is the application of such information to control drug metabolism and/or design selective P450 inhibitors. A final presentation highlighted development of a CYP3A4 inhibitor that slows clearance of human immunodeficiency virus drugs otherwise rapidly metabolized by CYP3A4. Although understanding P450 structure/function relationships is an ongoing challenge, translational advances will benefit from continued integration of existing and new biophysical approaches. PMID:24130370

Johnson, Eric F.; Connick, J. Patrick; Reed, James R.; Backes, Wayne L.; Desai, Manoj C.; Xu, Lianhong; Estrada, D. Fernando; Laurence, Jennifer S.

2014-01-01

230

Correlation of Homocysteine Metabolic Enzymes Gene Polymorphism and Mild Cognitive Impairment in the Xinjiang Uygur Population  

PubMed Central

Background The aim of this study was to investigate the genetic polymorphisms in the homocysteine (HCY) metabolic enzymes in the Xinjiang Uygur population who have mild cognitive impairment (MCI). Material/Methods Based on the epidemiological investigation, 129 cases of diagnosed Uygur MCI patients and a matched control group with 131 cases were enrolled for analyzing the association between the polymorphisms in the HCY metabolism related genes (C677T, A1298C, and G1968A polymorphisms in MTHFR, as well as the A2756G polymorphism in MS) and MCI by using the SNaPshot method. We then determined the homocysteine level in patients. Results In Xinjiang Uygur subjects, the A1298C polymorphisms in MTHFR and the A2756G polymorphisms in the MS gene in the MCI group were different from those in the control group. However, the C677T and G1968A polymorphisms in the MTHFR gene in MCI patients were not different from those in the control group. Multivariate logistic regression showed that, in addition to the well-known risk factors, such as low education level, high cholesterol level, high level of low-density lipoprotein, and high homocysteine levels, the A>G mutation in the MS gene at the rs1805087 locus was another independent risk factor for MCI in the Uyghur MCI population. The risk of MCI in G allele carriers was 2.265 times higher than that in matched control individuals (95% CI: 1.205~4.256, P<0.05). Conclusions The genetic polymorphism of HCY metabolizing enzymes is correlated to the occurrence of MCI in the Xinjiang Uygur population. The A2756G polymorphism in the MS gene could be an independent risk factor for MCI in the Xinjiang Uygur population. PMID:25625218

Luo, Mei; Ji, Huihui; Zhou, Xiaohui; Liang, Jie; Zou, Ting

2015-01-01

231

Dorsomedial hindbrain catecholamine regulation of hypothalamic astrocyte glycogen metabolic enzyme protein expression: Impact of estradiol.  

PubMed

The brain astrocyte glycogen reservoir is a vital energy reserve and, in the cerebral cortex, subject among other factors to noradrenergic control. The ovarian steroid estradiol potently stimulates nerve cell aerobic respiration, but its role in glial glycogen metabolism during energy homeostasis or mismatched substrate supply/demand is unclear. This study examined the premise that estradiol regulates hypothalamic astrocyte glycogen metabolic enzyme protein expression during normo- and hypoglycemia in vivo through dorsomedial hindbrain catecholamine (CA)-dependent mechanisms. Individual astrocytes identified in situ by glial fibrillary acidic protein immunolabeling were laser-microdissected from the ventromedial hypothalamic (VMH), arcuate hypothalamic (ARH), and paraventricular hypothalamic (PVH) nuclei and the lateral hypothalamic area (LHA) of estradiol (E)- or oil (O)-implanted ovariectomized (OVX) rats after insulin or vehicle injection, and pooled within each site. Stimulation [VMH, LHA] or suppression [PVH, ARH] of basal glycogen synthase (GS) protein expression by E was reversed in the former three sites by caudal fourth ventricular pretreatment with the CA neurotoxin 6-hydroxydopamine (6-OHDA). E diminished glycogen phosphorylase (GP) protein profiles by CA-dependent [VMH, PVH] or -independent mechanisms [LHA]. Insulin-induced hypoglycemia (IIH) increased GS expression in the PVH in OVX+E, but reduced this protein in the PVH, ARH, and LHA in OVX+O. Moreover, IIH augmented GP expression in the VMH, LHA, and ARH in OVX+E and in the ARH in OVX+O, responses that normalized by 6-OHDA. Results demonstrate site-specific effects of E on astrocyte glycogen metabolic enzyme expression in the female rat hypothalamus, and identify locations where dorsomedial hindbrain CA input is required for such action. Evidence that E correspondingly increases and reduces basal GS and GP in the VMH and LHA, but augments the latter protein during IIH suggests that E regulates glycogen content and turnover in these structures during glucose sufficiency and shortage. PMID:25701713

Tamrakar, P; Shrestha, P K; Briski, K P

2015-04-30

232

Genetically Modeled Mice with Mutations in Mitochondrial Metabolic Enzymes for the Study of Cancer  

PubMed Central

Mitochondrial dysfunction has long been implicated in progression of cancer. As a paradigm, the “Warburg effect,” which by means of a switch toward anaerobic metabolism enables cancer cells to proliferate in oxygen limiting conditions, is well established. Besides this metabolic transformation of tumors, it has been discovered that mutations in genes encoding mitochondrial proteins are the etiological factors in different types of cancer. This confers to mitochondrial dysfunction a causative role, rather than resultant, in tumor genesis beyond its role in tumor progression and development. Mitochondrial proteins encoded by tumor-suppressor genes are part of the succinate-dehydrogenase, the fumarate-hydratase, and the mitochondrial isocitrate-dehydrogenase enzymes, all of them participating in the Krebs cycle. The spectrum of tumors associated with mutations in these genes is becoming larger and varies between each enzyme. Several mechanisms of tumorigenesis have been proposed for the different enzymatic defects, most of them based on studies using cellular and animal models. Regarding the molecular pathways implicated in the oncogenic transformation, one of the first accepted theories was based on the constitutive expression of the hypoxia-inducible factor 1? (Hif1?) at normal oxygen tension, a theory referred to as “pseudo-hypoxic drive.” This mechanism has been linked to the three types of mutations, thus suggesting a central role in cancer. However, other alternative molecular processes, such as oxidative stress or altered chromatin remodeling, have been also proposed to play an onco-pathogenic role. In the recent years, the role of oncometabolites, a new concept emerged from biochemical studies upon these tumors, has acquired relevance as responsible for tumor formation. Nevertheless, the actual contribution of each of these mechanisms has not been definitively established. In this review, we summarize the results obtained from mouse strains genetically modified in the three different enzymes. PMID:25126540

Piruat, José I.; Millán-Uclés, África

2014-01-01

233

The effects of aflatoxin B1 on transporters and steroid metabolizing enzymes in JEG-3 cells.  

PubMed

Effects of 96 h aflatoxin B1 (AFB1) exposure at concentrations from 0.2 ?M to 6 ?M on the mRNA and protein expression levels of the following transporters ABCB1/B4, ABCC1, ABCC2, ABCG2, OAT4 and the mRNA expression of steroid-metabolizing enzymes CYP1A1, CYP19A1, HSD3B1 and HSD17B1, and conjugating enzyme family UGT1A were evaluated in trophoblastic JEG-3 cells. Statistically significant dose-dependent five-fold increases in the expression levels with ABCC2 and OAT4 were recorded at 2 and 6?M AFB1. Protein expression of ABCG2 was decreased dose-dependently with 0.2-6 ?M AFB1. With the other transporters, only a trend of increased expression was observed. Analogously, a three-fold increase in the expressions of CYP19A1, HSD3B1, HSD17B1 and UGT1A-family were observed at 0.3 ?M AFB1. When an inhibitor of CYP19A1, finrozole, was dosed simultaneously with AFB1, no increases in the transcripts of transporters or steroid hydroxylases or CYP19A1 were observed. This delayed increase in the expression levels - only after 96h incubations - may indicate that the response is due to a secondary metabolite of AFB1 or other secondary controlling cascades rather than the parent compound itself. In conclusion, AFB1 affected the placental steroid synthesizing, metabolizing and conjugating enzymes as well as the expression levels of several transporter proteins in JEG-3 cells. These alterations may lead to anomalies in the foetoplacental hormonal homeostasis. PMID:23402939

Huuskonen, Pasi; Myllynen, Päivi; Storvik, Markus; Pasanen, Markku

2013-04-26

234

Effect of honokiol on the induction of drug-metabolizing enzymes in human hepatocytes  

PubMed Central

Honokiol, 2-(4-hydroxy-3-prop-2-enyl-phenyl)-4-prop-2-enyl-phenol, an active component of Magnolia officinalis and Magnolia grandiflora, exerts various pharmacological activities such as antitumorigenic, antioxidative, anti-inflammatory, neurotrophic, and antithrombotic effects. To investigate whether honokiol acts as a perpetrator in drug interactions, messenger ribonucleic acid (mRNA) levels of phase I and II drug-metabolizing enzymes, including cytochrome P450 (CYP), UDP-glucuronosyltransferase (UGT), and sulfotransferase 2A1 (SULT2A1), were analyzed by real-time reverse transcription polymerase chain reaction following 48-hour honokiol exposure in three independent cryopreserved human hepatocyte cultures. Honokiol treatment at the highest concentration tested (50 ?M) increased the CYP2B6 mRNA level and CYP2B6-catalyzed bupropion hydroxylase activity more than two-fold in three different hepatocyte cultures, indicating that honokiol induces CYP2B6 at higher concentrations. However, honokiol treatment (0.5–50 ?M) did not significantly alter the mRNA levels of phase I enzymes (CYP1A2, CYP3A4, CYP2C8, CYP2C9, and CYP2C19) or phase II enzymes (UGT1A1, UGT1A4, UGT1A9, UGT2B7, and SULT2A1) in cryopreserved human hepatocyte cultures. CYP1A2-catalyzed phenacetin O-deethylase and CYP3A4-catalyzed midazolam 1?-hydroxylase activities were not affected by 48-hour honokiol treatment in cryopreserved human hepatocytes. These results indicate that honokiol is a weak CYP2B6 inducer and is unlikely to increase the metabolism of concomitant CYP2B6 substrates and cause pharmacokinetic-based drug interactions in humans. PMID:25395831

Cho, Yong-Yeon; Jeong, Hyeon-Uk; Kim, Jeong-Han; Lee, Hye Suk

2014-01-01

235

Effect of a PCB-based transformer oil on testicular steroidogenesis and xenobiotic-metabolizing enzymes.  

PubMed

Pyralene is a PCB-based transformer oil with a unique PCB congener profile when compared to other mixtures. We studied the influence of Pyralene on testicular steroidogenesis and the status of xenobiotic-metabolizing enzymes in the testis and liver of rats during oral exposure (10 and 50 mg/kg body weight, p.o. daily for 1 week) and a 3-week post-treatment recovery period. As expected, Pyralene induced a rapid and sustained increase in mRNA transcripts for CYP1A1 and CYP2B1 in hepatocytes that was associated with a dramatic increase in ethoxyresorufin-O-deethylase (EROD) and pentoxyresorufin-O-deethylase (PROD) activities. Testicular androgenesis and the conversion of progesterone to testosterone in testicular microsomes were bidirectionally affected. An increase in these parameters was observed 24h after the initial administration of Pyralene, followed by inhibition that lasted until the fourth post-treatment day. Expression PCR analysis revealed a significant decrease in 17beta-hydroxysteroid dehydrogenase (17betaHSD) transcript abundance at 48 h after Pyralene administration. In contrast, transcripts for several other steroidogenic enzymes and for testicular CYP1A1, CYP1B1, and CYP2B1 were unaffected under the same conditions. These results in the rat indicate that a sub-chronic exposure to Pyralene disrupted testicular steroidogenesis and suggest the mechanism may involve direct action on the regulation of specific steroidogenic enzymes such as 17betaHSD. PMID:16439096

Andric, Nebojsa L; Kostic, Tatjana S; Zoric, Sonja N; Stanic, Bojana D; Andric, Silvana A; Kovacevic, Radmila Z

2006-07-01

236

The evolution of pyrroline-5-carboxylate synthase in plants: a key enzyme in proline synthesis  

Microsoft Academic Search

Many plants synthesize and accumulate proline in response to osmotic stress conditions. A central enzyme in the proline biosynthesis\\u000a is the bifunctional enzyme ?1-pyrroline-5-carboxylate synthase (P5CS) that includes two functional catalytic domains: the ?-glutamyl kinase and the glutamic-?-semialdehyde\\u000a dehydrogenase. This enzyme catalyzes the first two steps of the proline biosynthetic pathway and plays a central role in the\\u000a regulation of

Andreia Carina Turchetto-Zolet; Marcia Margis-Pinheiro; Rogerio Margis

2009-01-01

237

Garlic Oil Attenuated Nitrosodiethylamine-Induced Hepatocarcinogenesis by Modulating the Metabolic Activation and Detoxification Enzymes  

PubMed Central

Nitrosodiethylamine (NDEA) is a potent carcinogen widely existing in the environment. Our previous study has demonstrated that garlic oil (GO) could prevent NDEA-induced hepatocarcinogenesis in rats, but the underlying mechanisms are not fully understood. It has been well documented that the metabolic activation may play important roles in NDEA-induced hepatocarcinogenesis. Therefore, we designed the current study to explore the potential mechanisms by investigating the changes of hepatic phase ? enzymes (including cytochrome P450 enzyme (CYP) 2E1, CYP1A2 and CYP1A1) and phase ? enzymes (including glutathione S transferases (GSTs) and UDP- Glucuronosyltransferases (UGTs)) by using enzymatic methods, real-time PCR, and western blotting analysis. We found that NDEA treatment resulted in significant decreases of the activities of CYP2E1, CYP1A2, GST alpha, GST mu, UGTs and increases of the activities of CYP1A1 and GST pi. Furthermore, the mRNA and protein levels of CYP2E1, CYP1A2, GST alpha, GST mu and UGT1A6 in the liver of NDEA-treated rats were significantly decreased compared with those of the control group rats, while the mRNA and protein levels of CYP1A1 and GST pi were dramatically increased. Interestingly, all these adverse effects induced by NDEA were simultaneously and significantly suppressed by GO co-treatment. These data suggest that the protective effects of GO against NDEA-induced hepatocarcinogenesis might be, at least partially, attributed to the modulation of phase I and phase II enzymes. PMID:23494807

Zhang, Cui-Li; Zeng, Tao; Zhao, Xiu-Lan; Xie, Ke-Qin

2013-01-01

238

Photosynthesis in Rhodospirillum rubrum. III. Metabolic Control of Reductive Pentose Phosphate and Tricarboxylic Acid Cycle Enzymes 1  

PubMed Central

Enzymes of the reductive pentose phosphate cycle including ribulose-diphosphate carboxylase, ribulose-5-phosphate kinase, ribose-5-phosphate isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and alkaline fructose-1,6-diphos-phatase were shown to be present in autotrophically grown Rhodospirillum rubrum. Enzyme levels were measured in this organism grown photo- and dark heterotrophically as well. Several, but not all, of these enzymes appeared to be under metabolic control, mediated by exogenous carbon and nitrogen compounds. Light had no effect on the presence or levels of any of these enzymes in this photosynthetic bacterium. The enzymes of the tricarboxylic acid cycle and enolase were shown to be present in R. rubrum cultured aerobically, autotrophically, or photoheterotrophically, both in cultures evolving hydrogen and under conditions where hydrogen evolution is not observed. Light had no clearly demonstrable effect on the presence or levels of any of these enzymes. PMID:6042359

Anderson, Louise; Fuller, R. C.

1967-01-01

239

A network-based approach for predicting key enzymes explaining metabolite abundance alterations in a disease phenotype  

PubMed Central

Background The study of metabolism has attracted much attention during the last years due to its relevance in various diseases. The advance in metabolomics platforms allows us to detect an increasing number of metabolites in abnormal high/low concentration in a disease phenotype. Finding a mechanistic interpretation for these alterations is important to understand pathophysiological processes, however it is not an easy task. The availability of genome scale metabolic networks and Systems Biology techniques open new avenues to address this question. Results In this article we present a novel mathematical framework to find enzymes whose malfunction explains the accumulation/depletion of a given metabolite in a disease phenotype. Our approach is based on a recently introduced pathway concept termed Carbon Flux Paths (CFPs), which extends classical topological definition by including network stoichiometry. Using CFPs, we determine the Connectivity Curve of an altered metabolite, which allows us to quantify changes in its pathway structure when a certain enzyme is removed. The influence of enzyme removal is then ranked and used to explain the accumulation/depletion of such metabolite. For illustration, we center our study in the accumulation of two metabolites (L-Cystine and Homocysteine) found in high concentration in the brain of patients with mental disorders. Our results were discussed based on literature and found a good agreement with previously reported mechanisms. In addition, we hypothesize a novel role of several enzymes for the accumulation of these metabolites, which opens new strategies to understand the metabolic processes underlying these diseases. Conclusions With personalized medicine on the horizon, metabolomic platforms are providing us with a vast amount of experimental data for a number of complex diseases. Our approach provides a novel apparatus to rationally investigate and understand metabolite alterations under disease phenotypes. This work contributes to the development of Systems Medicine, whose objective is to answer clinical questions based on theoretical methods and high-throughput “omics” data. PMID:23870038

2013-01-01

240

Changes in nitrogen metabolism and antioxidant enzyme activities of maize tassel in black soils region of northeast China  

PubMed Central

Two varieties of maize (Zea mays L.) grown in fields in black soils of northeast China were tested to study the dynamic changes of nitrogen metabolism and antioxidant enzyme activity in tassels of maize. Results showed that antioxidant enzyme activity in tassels of maize increased first and then decreased with the growing of maize, and reached peak value at shedding period. Pattern of proline was consistent with antioxidant enzyme activity, showing that osmotic adjustment could protect many enzymes, which are important for cell metabolism. Continuous reduction of soluble protein content along with the growing of maize was observed in the study, which indicated that quantitative material and energy were provided for pollen formation. Besides, another major cause was that a large proportion of nitrogen was used for the composition of structural protein. Nitrate nitrogen concentrations of tassels were more variable than ammonium nitrogen, which showed that nitrate nitrogen was the favored nitrogen source for maize. PMID:25324855

Xu, Hongwen; Lu, Yan; Xie, Zhiming; Song, Fengbin

2014-01-01

241

Variation in the Activity of Some Enzymes of Photorespiratory Metabolism in C4 Grasses  

PubMed Central

• Background and Aims Photorespiration occurs in C4 plants, although rates are small compared with C3 plants. The amount of glycine decarboxylase in the bundle sheath (BS) varies among C4 grasses and is positively correlated with the granal index (ratio of the length of appressed thylakoid membranes to the total length of all thylakoid membranes) of the BS chloroplasts: C4 grasses with high granal index contained more glycine decarboxylase per unit leaf area than those with low granal index, probably reflecting the differences in O2 production from photosystem II and the potential photorespiratory capacity. Thus, it is hypothesized that the activities of peroxisomal enzymes involved in photorespiration are also correlated with the granal development. • Methods The granal development in BS chloroplasts was investigated and activities of the photorespiratory enzymes assayed in 28 C4 grasses and seven C3 grasses. • Key Results The NADP–malic enzyme grasses were divided into two groups: one with low granal index and the other with relatively high granal index in the BS chloroplasts. Both the NAD–malic enzyme and phosphoenolpyruvate carboxykinase grasses had high granal index in the BS chloroplasts. No statistically significant differences were found in activity of hydroxypyruvate reductase between the C3 and C4 grasses, or between the C4 subtypes. The activity of glycolate oxidase and catalase were smaller in the C4 grasses than in the C3 grasses. Among the C4 subtypes, glycolate oxidase activities were significantly smaller in the NADP–malic enzyme grasses with low granal index in the BS chloroplasts, compared with in the C4 grasses with substantial grana in the BS chloroplasts. • Conclusions There is interspecies variation in glycolate oxidase activity associated with the granal development in the BS chloroplasts and the O2 production from photosystem II, which suggests different potential photorespiration capacities among C4 grasses. PMID:16100226

UENO, OSAMU; YOSHIMURA, YASUYUKI; SENTOKU, NAOKI

2005-01-01

242

Efficient production of optically pure l-lactic acid from food waste at ambient temperature by regulating key enzyme activity.  

PubMed

Bio-production of optically pure l-lactic acid from food waste has attracted much interest as it can treat organic wastes with simultaneous recovery of valuable by-products. However, the yield of l-lactic acid was very low and no optically pure l-lactic acid was produced in the literature due to (1) the lower activity of enzymes involved in hydrolysis and l-lactic acid generation, and (2) the participation of other enzymes related to d-lactic acid and acetic and propionic acids production. In this paper, a new strategy was reported for effective production of optically pure l-lactic acid from food waste at ambient temperature, i.e. via regulating key enzyme activity by sewage sludge supplement and intermittent alkaline fermentation. It was found that not only optically pure l-lactic acid was produced, but the yield was enhanced by 2.89-fold. The mechanism study showed that the activities of enzymes relevant to food waste hydrolysis and lactic acid production were enhanced, and the key enzymes related to volatile fatty acids and d-lactic acid generations were severally decreased or inhibited. Also, the microbes responsible for l-lactic acid production were selectively proliferated. Finally, the pilot-scale continuous experiment was conducted to testify the feasibility of this new technique. PMID:25528545

Li, Xiang; Chen, Yinguang; Zhao, Shu; Chen, Hong; Zheng, Xiong; Luo, Jinyang; Liu, Yanan

2015-03-01

243

Berberine bridge enzyme, a key branch-point enzyme in benzylisoquinoline alkaloid biosynthesis, contains a vacuolar sorting determinant.  

PubMed

In opium poppy (Papaver somniferum L.), (S)-reticuline is the last common intermediate in sanguinarine and morphine biosynthesis. Sanguinarine accumulates in the vacuole of cultured opium poppy cells in response to treatment with fungal elicitors. The first committed step in sanguinarine biosynthesis is catalyzed by the berberine bridge enzyme (BBE), which converts (S)-reticuline to (S)-scoulerine. An N-terminal signal peptide and novel vacuolar sorting determinant were identified and characterized in BBE. In vitro translation of BBE mRNA in the presence of canine pancreatic microsomes produced a glycosylated, proteolysis-resistant protein, confirming the existence of a signal peptide. Transcripts encoding a BBE N-terminal deletion series fused to beta-glucuronidase or green fluorescent protein (GFP) were also translated in the presence of canine microsomes, and introduced into cultured opium poppy cells via microprojectile bombardment. The signal peptide was restricted to the first 25 amino acids and shown to initially target BBE to the endoplasmic reticulum. Fusion of 50 N-terminal residues from BBE to GFP resulted in the localization of the reporter to the vacuole. GFP was also sorted to the vacuole when fused to a heterologous N-terminal signal peptide followed by BBE amino acids 26-50. The BBE vacuolar sorting determinant was further localized between residues 26 and 41 by deletion analysis. The final subcellular destination of BBE is consistent with the vacuolar sequestration of sanguinarine. However, the vacuolar pH is below the functional range for BBE, suggesting that the enzyme is active only prior to its entry into the vacuole. PMID:11722125

Bird, D A; Facchini, P J

2001-10-01

244

Enzyme Informatics  

PubMed Central

Over the last 50 years, sequencing, structural biology and bioinformatics have completely revolutionised biomolecular science, with millions of sequences and tens of thousands of three dimensional structures becoming available. The bioinformatics of enzymes is well served by, mostly free, online databases. BRENDA describes the chemistry, substrate specificity, kinetics, preparation and biological sources of enzymes, while KEGG is valuable for understanding enzymes and metabolic pathways. EzCatDB, SFLD and MACiE are key repositories for data on the chemical mechanisms by which enzymes operate. At the current rate of genome sequencing and manual annotation, human curation will never finish the functional annotation of the ever-expanding list of known enzymes. Hence there is an increasing need for automated annotation, though it is not yet widespread for enzyme data. In contrast, functional ontologies such as the Gene Ontology already profit from automation. Despite our growing understanding of enzyme structure and dynamics, we are only beginning to be able to design novel enzymes. One can now begin to trace the functional evolution of enzymes using phylogenetics. The ability of enzymes to perform secondary functions, albeit relatively inefficiently, gives clues as to how enzyme function evolves. Substrate promiscuity in enzymes is one example of imperfect specificity in protein-ligand interactions. Similarly, most drugs bind to more than one protein target. This may sometimes result in helpful polypharmacology as a drug modulates plural targets, but also often leads to adverse side-effects. Many cheminformatics approaches can be used to model the interactions between druglike molecules and proteins in silico. We can even use quantum chemical techniques like DFT and QM/MM to compute the structural and energetic course of enzyme catalysed chemical reaction mechanisms, including a full description of bond making and breaking. PMID:23116471

Alderson, Rosanna G.; Ferrari, Luna De; Mavridis, Lazaros; McDonagh, James L.; Mitchell, John B. O.; Nath, Neetika

2012-01-01

245

The Stabilisation of Enzymes — a Key Factor in the Practical Application of Biocatalysis  

NASA Astrophysics Data System (ADS)

The present state of the problem of the practical application of biocatalysis is examined and physicochemical approaches whereby the denaturation of enzymes under the influence of elevated temperatures, extreme pH values, and organic solvents can be suppressed are analysed. The general principles of the stabilisation of enzymes are formulated. The bibliography includes 225 references.

Martinek, K.; Berezin, I. B.

1980-05-01

246

Protein Acetylation Microarray Reveals NuA4 Controls Key Metabolic Target Regulating Gluconeogenesis  

PubMed Central

SUMMARY Histone acetyltransferases (HATs) and histone deacetylases (HDACs) conduct many critical functions through nonhistone substrates in metazoans, but only chromatin-associated nonhistone substrates are known in Saccharomyces cerevisiae. Using yeast proteome microarrays, we identified and validated many nonchromatin substrates of the essential nucleosome acetyltransferase of H4 (NuA4) complex. Among these, acetylation sites (Lys 19 and 514) of phosphoenolpyruvate carboxykinase (Pck1p) were determined by tandem mass spectrometry. Acetylation at Lys 514 was crucial for enzymatic activity and the ability of yeast cells to grow on non-fermentable carbon sources. Loss of Pck1p activity blocked the extension of yeast chronological life span caused by water starvation. In human hepatocellular carcinoma (HepG2) cells, human Pck1 acetylation and glucose production was dependent on TIP60, the human homolog of ESA1. Our results demonstrate a novel regulatory function for the NuA4 complex in glucose metabolism and life span by acetylating a critical metabolic enzyme. PMID:19303850

Lin, Yu-yi; Lu, Jin-ying; Zhang, Junmei; Walter, Wendy; Dang, Weiwei; Wan, Jun; Tao, Sheng-Ce; Qian, Jiang; Zhao, Yingming; Boeke, Jef D.; Berger, Shelley L.; Zhu, Heng

2009-01-01

247

Overexpression of metabolic enzymes at the junction of glycolylsis and the TCA cycle in Escherichia coli: physiological effects and application  

E-print Network

R. ngler (Member) James C. Holste (Member) Rayford . Anthony (Head of Department) December 1999 Major Subject: Chemical Engineering ABSTRACT Overexpression of Metabolic Enzymes at the Junction of Glycolysis and the TCA Cycle in Escherichia... AND METHODS . . 1 5 2. 1 2. 2 2. 3 Strains and Plasmids Used Growth Media and Conditions Other Methods. 2. 3. 1 DNA Transductions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2. 3. 2 Enzyme Assays. . 2. 3. 3 SDS-PAGE. , 2. 3. 4...

Spitzer, Richard G.

1999-01-01

248

Elucidating Organ-Specific Metabolic Toxicity Chemistry from Electrochemiluminescent Enzyme/DNA Arrays and Bioreactor Bead-LC-MS/MS†  

PubMed Central

Human toxic responses are very often related to metabolism. Liver metabolism is traditionally studied, but other organs also convert chemicals and drugs to reactive metabolites leading to toxicity. When DNA damage is found, the effects are termed genotoxic. Here we describe a comprehensive new approach to evaluate chemical genotoxicity pathways from metabolites formed in-situ by a broad spectrum of liver, lung, kidney and intestinal enzymes. DNA damage rates are measured with a microfluidic array featuring a 64-nanowell chip to facilitate fabrication of films of DNA, electrochemiluminescent (ECL) detection polymer [Ru(bpy)2(PVP)10]2+ {(PVP = poly(4-vinylpyridine)} and metabolic enzymes. First, multiple enzyme reactions are run on test compounds using the array, then ECL light related to the resulting DNA damage is measured. A companion method next facilitates reaction of target compounds with DNA/enzyme-coated magnetic beads in 96 well plates, after which DNA is hydrolyzed and nucleobase-metabolite adducts are detected by LC-MS/MS. The same organ enzymes are used as in the arrays. Outcomes revealed nucleobase adducts from DNA damage, enzymes responsible for reactive metabolites (e.g. cyt P450s), influence of bioconjugation, relative dynamics of enzymes suites from different organs, and pathways of possible genotoxic chemistry. Correlations between DNA damage rates from the cell-free array and organ-specific cell-based DNA damage were found. Results illustrate the power of the combined DNA/enzyme microarray/LC-MS/MS approach to efficiently explore a broad spectrum of organ-specific metabolic genotoxic pathways for drugs and environmental chemicals.

Wasalathanthri, Dhanuka P.; Li, Dandan; Song, Donghui; Zheng, Zhifang; Choudhary, Dharamainder; Jansson, Ingela; Lu, Xiuling; Schenkman, John B.; Rusling, James F.

2015-01-01

249

Complete Proteomic-Based Enzyme Reaction and Inhibition Kinetics Reveal How Monolignol Biosynthetic Enzyme Families Affect Metabolic Flux and Lignin in Populus trichocarpa[W  

PubMed Central

We established a predictive kinetic metabolic-flux model for the 21 enzymes and 24 metabolites of the monolignol biosynthetic pathway using Populus trichocarpa secondary differentiating xylem. To establish this model, a comprehensive study was performed to obtain the reaction and inhibition kinetic parameters of all 21 enzymes based on functional recombinant proteins. A total of 104 Michaelis-Menten kinetic parameters and 85 inhibition kinetic parameters were derived from these enzymes. Through mass spectrometry, we obtained the absolute quantities of all 21 pathway enzymes in the secondary differentiating xylem. This extensive experimental data set, generated from a single tissue specialized in wood formation, was used to construct the predictive kinetic metabolic-flux model to provide a comprehensive mathematical description of the monolignol biosynthetic pathway. The model was validated using experimental data from transgenic P. trichocarpa plants. The model predicts how pathway enzymes affect lignin content and composition, explains a long-standing paradox regarding the regulation of monolignol subunit ratios in lignin, and reveals novel mechanisms involved in the regulation of lignin biosynthesis. This model provides an explanation of the effects of genetic and transgenic perturbations of the monolignol biosynthetic pathway in flowering plants. PMID:24619611

Wang, Jack P.; Naik, Punith P.; Chen, Hsi-Chuan; Shi, Rui; Lin, Chien-Yuan; Liu, Jie; Shuford, Christopher M.; Li, Quanzi; Sun, Ying-Hsuan; Tunlaya-Anukit, Sermsawat; Williams, Cranos M.; Muddiman, David C.; Ducoste, Joel J.; Sederoff, Ronald R.; Chiang, Vincent L.

2014-01-01

250

Complete proteomic-based enzyme reaction and inhibition kinetics reveal how monolignol biosynthetic enzyme families affect metabolic flux and lignin in Populus trichocarpa.  

PubMed

We established a predictive kinetic metabolic-flux model for the 21 enzymes and 24 metabolites of the monolignol biosynthetic pathway using Populus trichocarpa secondary differentiating xylem. To establish this model, a comprehensive study was performed to obtain the reaction and inhibition kinetic parameters of all 21 enzymes based on functional recombinant proteins. A total of 104 Michaelis-Menten kinetic parameters and 85 inhibition kinetic parameters were derived from these enzymes. Through mass spectrometry, we obtained the absolute quantities of all 21 pathway enzymes in the secondary differentiating xylem. This extensive experimental data set, generated from a single tissue specialized in wood formation, was used to construct the predictive kinetic metabolic-flux model to provide a comprehensive mathematical description of the monolignol biosynthetic pathway. The model was validated using experimental data from transgenic P. trichocarpa plants. The model predicts how pathway enzymes affect lignin content and composition, explains a long-standing paradox regarding the regulation of monolignol subunit ratios in lignin, and reveals novel mechanisms involved in the regulation of lignin biosynthesis. This model provides an explanation of the effects of genetic and transgenic perturbations of the monolignol biosynthetic pathway in flowering plants. PMID:24619611

Wang, Jack P; Naik, Punith P; Chen, Hsi-Chuan; Shi, Rui; Lin, Chien-Yuan; Liu, Jie; Shuford, Christopher M; Li, Quanzi; Sun, Ying-Hsuan; Tunlaya-Anukit, Sermsawat; Williams, Cranos M; Muddiman, David C; Ducoste, Joel J; Sederoff, Ronald R; Chiang, Vincent L

2014-03-01

251

Evidence that a Linear Megaplasmid Encodes Enzymes of Aliphatic Alkene and Epoxide Metabolism and Coenzyme M (2-Mercaptoethanesulfonate) Biosynthesis in Xanthobacter Strain Py2  

PubMed Central

The bacterial metabolism of propylene proceeds by epoxidation to epoxypropane followed by a sequence of three reactions resulting in epoxide ring opening and carboxylation to form acetoacetate. Coenzyme M (2-mercaptoethanesulfonic acid) (CoM) plays a central role in epoxide carboxylation by serving as the nucleophile for epoxide ring opening and the carrier of the C3 unit that is ultimately carboxylated to acetoacetate, releasing CoM. In the present work, a 320-kb linear megaplasmid has been identified in the gram-negative bacterium Xanthobacter strain Py2, which contains the genes encoding the key enzymes of propylene oxidation and epoxide carboxylation. Repeated subculturing of Xanthobacter strain Py2 under nonselective conditions, i.e., with glucose or acetate as the carbon source in the absence of propylene, resulted in the loss of the propylene-positive phenotype. The propylene-negative phenotype correlated with the loss of the 320-kb linear megaplasmid, loss of induction and expression of alkene monooxgenase and epoxide carboxylation enzyme activities, and the loss of CoM biosynthetic capability. Sequence analysis of a hypothetical protein (XecG), encoded by a gene located downstream of the genes for the four enzymes of epoxide carboxylation, revealed a high degree of sequence identity with proteins of as-yet unassigned functions in the methanogenic archaea Methanobacterium thermoautotrophicum and Methanococcus jannaschii and in Bacillus subtilis. The M. jannaschii homolog of XecG, MJ0255, is located next to a gene, MJ0256, that has been shown to encode a key enzyme of CoM biosynthesis (M. Graupner, H. Xu, and R. H. White, J. Bacteriol. 182: 4862–4867, 2000). We propose that the propylene-positive phenotype of Xanthobacter strain Py2 is dependent on the selective maintenance of a linear megaplasmid containing the genes for the key enzymes of alkene oxidation, epoxide carboxylation, and CoM biosynthesis. PMID:11244054

Krum, Jonathan G.; Ensign, Scott A.

2001-01-01

252

Modeling the role of covalent enzyme modification in Escherichia coli nitrogen metabolism  

PubMed Central

In the bacterium Escherichia coli, the enzyme glutamine synthetase (GS) converts ammonium into the amino acid glutamine. GS is principally active when the cell is experiencing nitrogen limitation, and its activity is regulated by a bicyclic covalent modification cascade. The advantages of this bicyclic-cascade architecture are poorly understood. We analyze a simple model of the GS cascade in comparison to other regulatory schemes and conclude that the bicyclic cascade is suboptimal for maintaining metabolic homeostasis of the free glutamine pool. Instead, we argue that the lag inherent in the covalent modification of GS slows the response to an ammonium shock and thereby allows GS to transiently detoxify the cell, while maintaining homeostasis over longer times. PMID:20057006

Kidd, Philip B

2013-01-01

253

Changes in metabolic enzymes, cortisol and glucose concentrations of Beluga (Huso huso) exposed to dietary methylmercury.  

PubMed

In this paper, effects of dietary methylmercury (MeHg) on several blood biochemical parameters including GLU (glucose), LDH (lactate dehydrogenase), AST (aspartate aminotransferase), ALT (alanine aminotransferase), ALP (alkaline phosphatase) and cortisol were investigated in the Beluga sturgeon (Huso huso). Beluga juveniles were fed for 32 days on four diets containing MeHg (control: 0.04 mg kg?ą; low: 0.76 mg kg?ą; medium: 7.88 mg kg?ą; and high 16.22 mg kg?ą treatment). Significant increases (P < 0.05) were observed in all biochemical parameters, except ALP levels, which decreased significantly (P < 0.05) compared to the control group with either dose- or time-dependent effects. These results suggest that long-term dietary MeHg exposure may affect metabolic enzyme activity and glucose levels in Belugas. These findings provide useful information for environmental and fishery officials to apply in future decisions for managing fish resources in Caspian Sea. PMID:21110088

Gharaei, Ahmad; Ghaffari, Mostafa; Keyvanshokooh, Saeed; Akrami, Reza

2011-09-01

254

Gene Expression Variability in Human Hepatic Drug Metabolizing Enzymes and Transporters  

PubMed Central

Interindividual variability in the expression of drug-metabolizing enzymes and transporters (DMETs) in human liver may contribute to interindividual differences in drug efficacy and adverse reactions. Published studies that analyzed variability in the expression of DMET genes were limited by sample sizes and the number of genes profiled. We systematically analyzed the expression of 374 DMETs from a microarray data set consisting of gene expression profiles derived from 427 human liver samples. The standard deviation of interindividual expression for DMET genes was much higher than that for non-DMET genes. The 20 DMET genes with the largest variability in the expression provided examples of the interindividual variation. Gene expression data were also analyzed using network analysis methods, which delineates the similarities of biological functionalities and regulation mechanisms for these highly variable DMET genes. Expression variability of human hepatic DMET genes may affect drug-gene interactions and disease susceptibility, with concomitant clinical implications. PMID:23637747

Yang, Lun; Price, Elvin T.; Chang, Ching-Wei; Li, Yan; Huang, Ying; Guo, Li-Wu; Guo, Yongli; Kaput, Jim; Shi, Leming; Ning, Baitang

2013-01-01

255

Enzymatic synthesis of RNAs capped with nucleotide analogues reveals the molecular basis for substrate selectivity of RNA capping enzyme: impacts on RNA metabolism.  

PubMed

RNA cap binding proteins have evolved to specifically bind to the N7-methyl guanosine cap structure found at the 5' ends of eukaryotic mRNAs. The specificity of RNA capping enzymes towards GTP for the synthesis of this structure is therefore crucial for mRNA metabolism. The fact that ribavirin triphosphate was described as a substrate of a viral RNA capping enzyme, raised the possibility that RNAs capped with nucleotide analogues could be generated in cellulo. Owing to the fact that this prospect potentially has wide pharmacological implications, we decided to investigate whether the active site of the model Paramecium bursaria Chlorella virus-1 RNA capping enzyme was flexible enough to accommodate various purine analogues. Using this approach, we identified several key structural determinants at each step of the RNA capping reaction and generated RNAs harboring various different cap analogues. Moreover, we monitored the binding affinity of these novel capped RNAs to the eIF4E protein and evaluated their translational properties in cellulo. Overall, this study establishes a molecular rationale for the specific selection of GTP over other NTPs by RNA capping enzyme It also demonstrates that RNAs can be enzymatically capped with certain purine nucleotide analogs, and it also describes the impacts of modified RNA caps on specific steps involved in mRNA metabolism. For instance, our results indicate that the N7-methyl group of the classical N7-methyl guanosine cap is not always indispensable for binding to eIF4E and subsequently for translation when compensatory modifications are present on the capped residue. Overall, these findings have important implications for our understanding of the molecular determinants involved in both RNA capping and RNA metabolism. PMID:24086504

Issur, Moheshwarnath; Bougie, Isabelle; Despins, Simon; Bisaillon, Martin

2013-01-01

256

Enzymatic Synthesis of RNAs Capped with Nucleotide Analogues Reveals the Molecular Basis for Substrate Selectivity of RNA Capping Enzyme: Impacts on RNA Metabolism  

PubMed Central

RNA cap binding proteins have evolved to specifically bind to the N7-methyl guanosine cap structure found at the 5’ ends of eukaryotic mRNAs. The specificity of RNA capping enzymes towards GTP for the synthesis of this structure is therefore crucial for mRNA metabolism. The fact that ribavirin triphosphate was described as a substrate of a viral RNA capping enzyme, raised the possibility that RNAs capped with nucleotide analogues could be generated in cellulo. Owing to the fact that this prospect potentially has wide pharmacological implications, we decided to investigate whether the active site of the model Parameciumbursaria Chlorella virus-1 RNA capping enzyme was flexible enough to accommodate various purine analogues. Using this approach, we identified several key structural determinants at each step of the RNA capping reaction and generated RNAs harboring various different cap analogues. Moreover, we monitored the binding affinity of these novel capped RNAs to the eIF4E protein and evaluated their translational properties in cellulo. Overall, this study establishes a molecular rationale for the specific selection of GTP over other NTPs by RNA capping enzyme It also demonstrates that RNAs can be enzymatically capped with certain purine nucleotide analogs, and it also describes the impacts of modified RNA caps on specific steps involved in mRNA metabolism. For instance, our results indicate that the N7-methyl group of the classical N7-methyl guanosine cap is not always indispensable for binding to eIF4E and subsequently for translation when compensatory modifications are present on the capped residue. Overall, these findings have important implications for our understanding of the molecular determinants involved in both RNA capping and RNA metabolism. PMID:24086504

Issur, Moheshwarnath; Bougie, Isabelle; Despins, Simon; Bisaillon, Martin

2013-01-01

257

In vitro drug metabolism by human carboxylesterase 1: focus on angiotensin-converting enzyme inhibitors.  

PubMed

Carboxylesterase 1 (CES1) is the major hydrolase in human liver. The enzyme is involved in the metabolism of several important therapeutic agents, drugs of abuse, and endogenous compounds. However, no studies have described the role of human CES1 in the activation of two commonly prescribed angiotensin-converting enzyme inhibitors: enalapril and ramipril. Here, we studied recombinant human CES1- and CES2-mediated hydrolytic activation of the prodrug esters enalapril and ramipril, compared with the activation of the known substrate trandolapril. Enalapril, ramipril, and trandolapril were readily hydrolyzed by CES1, but not by CES2. Ramipril and trandolapril exhibited Michaelis-Menten kinetics, while enalapril demonstrated substrate inhibition kinetics. Intrinsic clearances were 1.061, 0.360, and 0.02 ml/min/mg protein for ramipril, trandolapril, and enalapril, respectively. Additionally, we screened a panel of therapeutic drugs and drugs of abuse to assess their inhibition of the hydrolysis of p-nitrophenyl acetate by recombinant CES1 and human liver microsomes. The screening assay confirmed several known inhibitors of CES1 and identified two previously unreported inhibitors: the dihydropyridine calcium antagonist, isradipine, and the immunosuppressive agent, tacrolimus. CES1 plays a role in the metabolism of several drugs used in the treatment of common conditions, including hypertension, congestive heart failure, and diabetes mellitus; thus, there is a potential for clinically relevant drug-drug interactions. The findings in the present study may contribute to the prediction of such interactions in humans, thus opening up possibilities for safer drug treatments. PMID:24141856

Thomsen, Ragnar; Rasmussen, Henrik B; Linnet, Kristian

2014-01-01

258

Association of cyclophosphamide drug metabolizing enzyme polymorphisms and chemotherapy related ovarian failure in breast cancer survivors  

PubMed Central

STRUCTURED ABSTRACT Objective To determine if genetic variation in chemotherapy metabolism are associated with risk of ovarian failure in breast cancer patients after adjuvant chemotherapy. Design Prospective cohort study. Setting Comprehensive cancer center. Patients Early stage breast cancer patients who were premenopausal at cancer diagnosis and treatment. Interventions None. Main outcomes measures Chemotherapy related ovarian failure (CROF) Results 127 breast cancer subjects who were premenopausal at cancer diagnosis and underwent cyclophosphamide-based chemotherapy were genotyped for 9 single nucleotide polymorphisms (SNPs) in enzymes involved in cyclophosphamide activation (CYP3A4, CYP2B6, CYP3A5) and detoxification (GSTA1, GSTM1, GSTP1, GSTT1). Median age at chemotherapy was 43.2 years. Median years of follow up since chemotherapy were 5.2 years. For the entire cohort, there was no significant association between CROF and SNPs. However, the association between CROF and SNPs was modified by age at chemotherapy. In subjects younger than 45 at chemotherapy, CYP3A4*1B variants had significantly longer time to CROF than CYP3A4*1A homozygotes in an adjusted multivariable Cox model (HR 0.25 [95% CI 0.07–0.9]). Age and tamoxifen use were also independently associated with CROF. Conclusions A common SNP in a cyclophosphamide drug metabolizing enzyme appears to be related to ovarian failure after cyclophosphamide-based chemotherapy in young women with breast cancer. Larger prospective studies to validate these results should be directed toward women less than 45 years of age at chemotherapy. PMID:19376514

Su, H. Irene; Sammel, Mary D.; Velders, Luke; Horn, Michelle; Stankiewicz, Corrie; Matro, Jennifer; Gracia, Clarisa R.; Green, Jamie; DeMichele, Angela

2010-01-01

259

Protective effect of p-methoxycinnamic acid, an active phenolic acid against 1,2-dimethylhydrazine-induced colon carcinogenesis: modulating biotransforming bacterial enzymes and xenobiotic metabolizing enzymes.  

PubMed

Objective of the study is to evaluate the modifying potential of p-methoxycinnamic acid (p-MCA), an active rice bran phenolic acid on biotransforming bacterial enzymes and xenobiotic metabolizing enzymes in 1,2-dimethylhydrazine-induced rat colon carcinogenesis. 48 male albino wistar rats were divided into six groups. Group1 (control) received modified pellet diet and 0.1 % carboxymethylcellulose; group2 received modified pellet diet along with p-MCA (80 mg/kg b.wt. p.o.) everyday for 16 weeks; groups 3-6 received 1,2-dimethylhydrazine (DMH) (20 mg/kg b.wt.) subcutaneous injection once a week for the first 4 weeks, while groups 4-6 received p-MCA at three different doses of 20, 40 and 80 mg/kg b.wt. p.o. everyday for 16 weeks. A significant increase in carcinogen-activating enzymes (cytochrome P450, cytochrome b5, cytochrome P4502E1, NADH-cytochrome-b5-reductase and NADPH-cytochrome-P450 reductase) with concomitant decrease in phaseII enzymes, DT-Diaphorase, glutathione S-transferase, UDP-glucuronyl-transferase and gamma glutamyltransferase were observed in group3 compared to control. DMH treatment significantly increased the activities of feacal and colonic bacterial enzymes (?-glucosidase, ?-galactosidase, ?-glucuronidase, nitroreductase, sulphatase and mucinase). p-MCA supplementation (40 mg/kg b.wt) to carcinogen exposed rats inhibited these enzymes, which were near those of control rats. The formation of dysplastic aberrant crypt foci in the colon and the histopathological observations of the liver also supports our biochemical findings. p-MCA (40 mg/kg b.wt.) offers remarkable modulating efficacy of biotransforming bacterial and xenobiotic metabolizing enzymes in colon carcinogenesis. PMID:24908112

Gunasekaran, Sivagami; Venkatachalam, Karthikkumar; Jeyavel, Kabalimoorthy; Namasivayam, Nalini

2014-09-01

260

Epigenetic regulation of uterine biology by transcription factor KLF11 via posttranslational histone deacetylation of cytochrome p450 metabolic enzymes.  

PubMed

Endocrine regulation of uterine biology is critical for embryo receptivity and human reproduction. Uterine endometrium depends on extrinsic sex steroid input and hence likely has mechanisms that enable adaptation to hormonal variation. Emerging evidence suggests that sex steroid bioavailability in the endometrium is determined by adjusting their metabolic rate and fate via regulation of cytochrome (CYP) p450 enzymes. The CYP enzymes are targeted by ubiquitously expressed Sp/Krüppel-like (Sp/KLF) transcription factors. Specifically, KLF11 is highly expressed in reproductive tissues, regulates an array of endocrine/metabolic pathways via epigenetic histone-based mechanisms and, when aberrantly expressed, is associated with diabetes and reproductive tract diseases, such as leiomyoma and endometriosis. Using KLF11 as a model to investigate epigenetic regulation of endometrial first-pass metabolism, we evaluated the expression of a comprehensive array of metabolic enzymes in Ishikawa cells. KLF11 repressed most endometrial CYP enzymes. To characterize KLF11-recruited epigenetic regulatory mechanisms, we focused on the estrogen-metabolizing enzyme CYP3A4. KLF11 expression declined in secretory phase endometrial epithelium associated with increased CYP3A4 expression. Additionally, KLF11 bound to CYP3A4 promoter GC elements and thereby repressed promoter, message, protein as well as enzymatic function. This repression was epigenetically mediated, because KLF11 colocalized with and recruited the corepressor SIN3A/histone deacetylase resulting in selective deacetylation of the CYP3A4 promoter. Repression was reversed by a mutation in KLF11 that abrogated cofactor recruitment and binding. This repression was also pharmacologically reversible with an histone deacetylase inhibitor. Pharmacological alteration of endometrial metabolism could have long-term translational implications on human reproduction and uterine disease. PMID:25076120

Zheng, Ye; Tabbaa, Zaid M; Khan, Zaraq; Schoolmeester, John K; El-Nashar, Sherif; Famuyide, Abimbola; Keeney, Gary L; Daftary, Gaurang S

2014-11-01

261

Rice debranching enzyme isoamylase3 facilitates starch metabolism and affects plastid morphogenesis.  

PubMed

Debranching enzymes, which hydrolyze ?-1 and 6-glucosidic linkages in ?-polyglucans, play a dual role in the synthesis and degradation of starch in plants. A transposon-inserted rice mutant of isoamylase3 (isa3) contained an increased amount of starch in the leaf blade at the end of the night, indicating that ISA3 plays a role in the degradation of transitory starch during the night. An epitope-tagged ISA3 expressed in Escherichia coli exhibited hydrolytic activity on ?-limit dextrin and amylopectin. We investigated whether ISA3 plays a role in amyloplast development and starch metabolism in the developing endosperm. ISA3-green fluorescent protein (GFP) fusion protein expressed under the control of the rice ISA3 promoter was targeted to the amyloplast stroma in the endosperm. Overexpression of ISA3 in the sugary1 mutant, which is deficient in ISA1 activity, did not convert water-soluble phytoglycogen to starch granules, indicating that ISA1 and ISA3 are not functionally redundant. Both overexpression and loss of function of ISA3 in the endosperm generated pleomorphic amyloplasts and starch granules. Furthermore, chloroplasts in the leaf blade of isa3 seedlings were large and pleomorphic. These results suggest that ISA3 facilitates starch metabolism and affects morphological characteristics of plastids in rice. PMID:21551159

Yun, Min-Soo; Umemoto, Takayuki; Kawagoe, Yasushi

2011-06-01

262

Two chitin metabolic enzyme genes from Hyriopsis cumingii: cloning, characterization, and potential functions.  

PubMed

Chitin, the second most important natural polymer in the world, and its N-deacetylated derivative chitosan are found in a wide variety of organisms. These versatile biopolymers are associated with a broad range of biological functions. This article is the first to report the potential functions of 2 chitin metabolic enzyme genes from Hyriopsis cumingii. A chitinase-3 gene (Chi-3) and a chitin deacetylase gene (Cda) were cloned from H. cumingii and characterized. Semi-quantitative reverse transcription polymerase chain reaction analysis revealed that the Cda gene was expressed in blood, mantle, liver, stomach, kidney, intestine, gill, and foot, whereas Chi-3 was also expressed in those tissues but not in blood. The tissue-specific expression of H. cumingii Chi-3 indicated that other Chi genes may be involved in the H. cumingii immune system. Real-time quantitative polymerase chain reaction analysis showed that the expression of Chi-3 was significantly (P < 0.05) upregulated 12 h after shell damage, suggesting that Chi-3 might hydrolyze superfluous chitin after shell recovery and play a role in shell formation. Conversely, Cda expression did not change significantly (P > 0.05) to maintain a certain degree of acetylation in chitin/chitosan. This study enriches the basic research on chitin metabolic genes and lays foundations for further research of shell regeneration in mussels. PMID:23096918

Wang, G-L; Xu, B; Bai, Z-Y; Li, J-L

2012-01-01

263

Alteration of Fatty-Acid-Metabolizing Enzymes Affects Mitochondrial Form and Function in Hereditary Spastic Paraplegia  

PubMed Central

Hereditary spastic paraplegia (HSP) is considered one of the most heterogeneous groups of neurological disorders, both clinically and genetically. The disease comprises pure and complex forms that clinically include slowly progressive lower-limb spasticity resulting from degeneration of the corticospinal tract. At least 48 loci accounting for these diseases have been mapped to date, and mutations have been identified in 22 genes, most of which play a role in intracellular trafficking. Here, we identified mutations in two functionally related genes (DDHD1 and CYP2U1) in individuals with autosomal-recessive forms of HSP by using either the classical positional cloning or a combination of whole-genome linkage mapping and next-generation sequencing. Interestingly, three subjects with CYP2U1 mutations presented with a thin corpus callosum, white-matter abnormalities, and/or calcification of the basal ganglia. These genes code for two enzymes involved in fatty-acid metabolism, and we have demonstrated in human cells that the HSP pathophysiology includes alteration of mitochondrial architecture and bioenergetics with increased oxidative stress. Our combined results focus attention on lipid metabolism as a critical HSP pathway with a deleterious impact on mitochondrial bioenergetic function. PMID:23176821

Tesson, Christelle; Nawara, Magdalena; Salih, Mustafa A.M.; Rossignol, Rodrigue; Zaki, Maha S.; Al Balwi, Mohammed; Schule, Rebecca; Mignot, Cyril; Obre, Emilie; Bouhouche, Ahmed; Santorelli, Filippo M.; Durand, Christelle M.; Oteyza, Andrés Caballero; El-Hachimi, Khalid H.; Al Drees, Abdulmajeed; Bouslam, Naima; Lamari, Foudil; Elmalik, Salah A.; Kabiraj, Mohammad M.; Seidahmed, Mohammed Z.; Esteves, Typhaine; Gaussen, Marion; Monin, Marie-Lorraine; Gyapay, Gabor; Lechner, Doris; Gonzalez, Michael; Depienne, Christel; Mochel, Fanny; Lavie, Julie; Schols, Ludger; Lacombe, Didier; Yahyaoui, Mohamed; Al Abdulkareem, Ibrahim; Zuchner, Stephan; Yamashita, Atsushi; Benomar, Ali; Goizet, Cyril; Durr, Alexandra; Gleeson, Joseph G.; Darios, Frederic; Brice, Alexis; Stevanin, Giovanni

2012-01-01

264

Polymorphisms in drug-metabolizing enzymes: What is their clinical relevance and why do they exist?  

SciTech Connect

The beautiful report by Sachse in this issue of the journal represents the culmination of 2 decades of increasingly exciting work on the {open_quotes}debrisoquine oxidation polymorphism,{close_quotes} one of dozens of pharmacogenetic or ecogenetic polymorphisms that have been shown to have an important impact on innumerable clinical diseases. Pharmacogenetics is the study of the hereditary basis of the differences in responses to drugs. Ecogenetics is the broader field of interindividual differences in response to all environmental chemical and physical agents (e.g., heavy metals, insecticides, compounds formed during combustion, and UV radiation). It is now clear that each of us has his or her own {open_quotes}individual fingerprint{close_quotes} of unique alleles encoding the so-called drug-metabolizing enzymes (DMEs) and the receptors that regulate these enzymes. In this invited editorial, I first introduce the current thinking in the field of DME (and DME-receptor) research and how DMEs have evolved from animal-plant interactions. I then describe the debrisoquine oxidation polymorphism, as well as two other relevant DME polymorphisms; show the relationship between these polymorphisms and human disease; provide examples of synergistic effects caused by the combination of two DME polymorphisms; and discuss the ethical considerations of such research. Last, I speculate on why these allelic frequencies of the DME genes might exist in human populations in the first place. 35 refs.

Nebert, D.W. [Univ. of Cincinnati Medical Center, OH (United States)

1997-02-01

265

Saturated fatty acid metabolism is key link between cell division, cancer, and senescence in cellular and whole organism aging  

PubMed Central

Cellular senescence is an in vivo and in vitro phenomenon, accompanied by physiological changes including cessation of division and disturbances of organelle structure and function. Review of the literature was undertaken to determine whether there is evidence that whole organism aging and cell senescence share a common initiation pathway. In vivo aged cells of different lineages, including aged T lymphocytes, show high expression of the INK4A-p16 gene. In cell culture when telomeres are shortened past a key length or state, the Arf/Ink gene system (p16/p14 humans, p16/p19 mice) switches on and activates p53, which suppresses further cell division. The p53 gene is a key tumor suppressor and its deletion or mutation allows cancerous growth. The switching on of p53 also causes changes in fatty acid metabolism, especially down-regulation of both fatty acid synthase and stearoyl-CoA (delta-9) desaturase. The co-suppression of these genes together with enhanced uptake of extracellular fatty acids, leads to raised levels of cellular palmitate and induction of either apoptosis or senescence. In senescent cells, the fatty acid composition of the cellular membranes alters and leads to changes in both structure and function of organelles, especially mitochondria. Animal models of accelerated aging exhibit repression of stearoyl-CoA desaturase activity while anti-aging calorie restriction stimulates the same enzyme system. It is concluded that aging in cells and whole organisms share a common initiation pathway and that cellular senescence is protective against cancer. Healthy longevity is likely to be most enhanced by factors that actively suppress excessive cell division. PMID:20431990

2010-01-01

266

The Catalytic Machinery of a Key Enzyme in Amino Acid Biosynthesis  

SciTech Connect

The aspartate pathway of amino acid biosynthesis is essential for all microbial life but is absent in mammals. Characterizing the enzyme-catalyzed reactions in this pathway can identify new protein targets for the development of antibiotics with unique modes of action. The enzyme aspartate {beta}-semialdehyde dehydrogenase (ASADH) catalyzes an early branch point reaction in the aspartate pathway. Kinetic, mutagenic, and structural studies of ASADH from various microbial species have been used to elucidate mechanistic details and to identify essential amino acids involved in substrate binding, catalysis, and enzyme regulation. Important structural and functional differences have been found between ASADHs isolated from these bacterial and fungal organisms, opening the possibility for developing species-specific antimicrobial agents that target this family of enzymes.

Viola, Ronald E.; Faehnle, Christopher R.; Blanco, Julio; Moore, Roger A.; Liu, Xuying; Arachea, Buenafe T.; Pavlovsky, Alexander G. (Toledo); (Yale); (Cold Spring); (NIH)

2013-02-28

267

Molecular, cellular, and tissue impact of depleted uranium on xenobiotic-metabolizing enzymes.  

PubMed

Enzymes that metabolize xenobiotics (XME) are well recognized in experimental models as representative indicators of organ detoxification functions and of exposure to toxicants. As several in vivo studies have shown, uranium can alter XME in the rat liver or kidneys after either acute or chronic exposure. To determine how length or level of exposure affects these changes in XME, we continued our investigation of chronic rat exposure to depleted uranium (DU, uranyl nitrate). The first study examined the effect of duration (1-18 months) of chronic exposure to DU, the second evaluated dose dependence, from a level close to that found in the environment near mining sites (0.2 mg/L) to a supra-environmental dose (120 mg/L, 10 times the highest level naturally found in the environment), and the third was an in vitro assessment of whether DU exposure directly affects XME and, in particular, CYP3A. The experimental in vivo models used here demonstrated that CYP3A is the enzyme modified to the greatest extent: high gene expression changed after 6 and 9 months. The most substantial effects were observed in the liver of rats after 9 months of exposure to 120 mg/L of DU: CYP3A gene and protein expression and enzyme activity all decreased by more than 40 %. Nonetheless, no direct effect of DU by itself was observed after in vitro exposure of rat microsomal preparations, HepG2 cells, or human primary hepatocytes. Overall, these results probably indicate the occurrence of regulatory or adaptive mechanisms that could explain the indirect effect observed in vivo after chronic exposure. PMID:24146111

Gueguen, Yann; Rouas, Caroline; Monin, Audrey; Manens, Line; Stefani, Johanna; Delissen, Olivia; Grison, Stéphane; Dublineau, Isabelle

2014-02-01

268

Molecular Architecture of DesI:  A Key Enzyme in the Biosynthesis of Desosamine † , ‡  

Microsoft Academic Search

Desosamine is a 3-(dimethylamino)-3,4,6-trideoxyhexose found, for example, in such macrolide antibiotics as erthyromycin, azithromycin, and clarithromycin. The efficacies of these macrolide antibiotics are markedly reduced in the absence of desosamine. In the bacterium Streptomyces Venezuelae, six enzymes are required for the production of dTDP-desosamine. The focus of this X-ray crystallographic analysis is the third enzyme in the pathway, a PLP-dependent

E. Sethe Burgie; Hazel M. Holden

2007-01-01

269

In vitro evaluation of the interaction potential of irosustat with drug-metabolizing enzymes.  

PubMed

Irosustat is a first-generation, irreversible, steroid sulfatase inhibitor currently in development for hormone-dependent cancer therapy. To predict clinical drug-drug interactions between irosustat and possible concomitantly administered medications, the inhibition/induction potential of irosustat with the main drug-metabolizing enzymes was investigated in vitro. The interaction of aromatase inhibitors in the in vitro metabolism of irosustat was also studied. Irosustat inhibited CYP1A2 activity in human liver microsomes through the formation of its desulfamoylated degradation product and metabolite 667-coumarin. CYP1A2 inhibition by 667-coumarin was competitive, with a K(i) of 0.77 ?M, a concentration exceeding by only 5-fold the maximal steady-state concentration of 667-coumarin in human plasma with the recommended dose of irosustat. In addition, 667-coumarin metabolites enhanced the inhibition of CYP1A2 activity. Additional clinical interaction studies of irosustat with CYP1A2 substrate drugs are strongly recommended. 667-Coumarin also appeared to be a competitive inhibitor of CYP2C19 (K(i) = 5.8 ?M) in human liver microsomes, and this inhibition increased with assessment in human hepatocytes. Inhibition of CYP2C19 enzyme activity was not caused by repression of CYP2C19 gene expression. Therefore, additional mechanistic experiments or follow-up studies with clinical evaluation are recommended. Irosustat neither inhibited CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2D6, CYP2E1, CYP3A4/5, or UDP-glucuronosyltransferase 1A1, 1A4, or 2B7 activities nor induced CYP1A2, CYP2C9, CYP2C19, or CYP3A4/5 at clinically relevant concentrations. Results from human liver microsomes indicated that no changes in irosustat pharmacokinetics in vivo are expected as a result of inhibition of irosustat metabolism in cases of concomitant medication administration or irosustat-aromatase inhibitor combination therapy with letrozole, anastrozole, or exemestane. PMID:22451700

Ventura, Verňnica; Solŕ, Josep; Peraire, Concepción; Brée, Françoise; Obach, Rosendo

2012-07-01

270

In-Silico Prediction of Key Metabolic Differences between Two Non-Small Cell Lung Cancer Subtypes  

PubMed Central

Metabolism expresses the phenotype of living cells and understanding it is crucial for different applications in biotechnology and health. With the increasing availability of metabolomic, proteomic and, to a larger extent, transcriptomic data, the elucidation of specific metabolic properties in different scenarios and cell types is a key topic in systems biology. Despite the potential of the elementary flux mode (EFM) concept for this purpose, its use has been limited so far, mainly because their computation has been infeasible for genome-scale metabolic networks. In a recent work, we determined a subset of EFMs in human metabolism and proposed a new protocol to integrate gene expression data, spotting key 'characteristic EFMs' in different scenarios. Our approach was successfully applied to identify metabolic differences among several human healthy tissues. In this article, we evaluated the performance of our approach in clinically interesting situation. In particular, we identified key EFMs and metabolites in adenocarcinoma and squamous-cell carcinoma subtypes of non-small cell lung cancers. Results are consistent with previous knowledge of these major subtypes of lung cancer in the medical literature. Therefore, this work constitutes the starting point to establish a new methodology that could lead to distinguish key metabolic processes among different clinical outcomes. PMID:25093336

Rezola, Alberto; Pey, Jon; Rubio, Ángel; Planes, Francisco J.

2014-01-01

271

Plastid Localization of the Key Carotenoid Enzyme Phytoene Synthase Is Altered by Isozyme, Allelic Variation, and Activity[W  

PubMed Central

Plant carotenoids have unique physiological roles related to specific plastid suborganellar locations. Carotenoid metabolic engineering could enhance plant adaptation to climate change and improve food security and nutritional value. However, lack of fundamental knowledge on carotenoid pathway localization limits targeted engineering. Phytoene synthase (PSY), a major rate-controlling carotenoid enzyme, is represented by multiple isozymes residing at unknown plastid sites. In maize (Zea mays), the three isozymes were transiently expressed and found either in plastoglobuli or in stroma and thylakoid membranes. PSY1, with one to two residue modifications of naturally occurring functional variants, exhibited altered localization, associated with distorted plastid shape and formation of a fibril phenotype. Mutating the active site of the enzyme reversed this phenotype. Discovery of differential PSY locations, linked with activity and isozyme type, advances the engineering potential for modifying carotenoid biosynthesis. PMID:23023170

Shumskaya, Maria; Bradbury, Louis M.T.; Monaco, Regina R.; Wurtzel, Eleanore T.

2012-01-01

272

Relationships between environmental organochlorine contaminant residues, plasma corticosterone concentrations, and intermediary metabolic enzyme activities in Great Lakes herring gull embryos.  

PubMed Central

Experiments were conducted to survey and detect differences in plasma corticosterone concentrations and intermediary metabolic enzyme activities in herring gull (Larus argentatus) embryos environmentally exposed to organochlorine contaminants in ovo. Unincubated fertile herring gull eggs were collected from an Atlantic coast control site and various Great Lakes sites in 1997 and artificially incubated in the laboratory. Liver and/or kidney tissues from approximately half of the late-stage embryos were analyzed for the activities of various intermediary metabolic enzymes known to be regulated, at least in part, by corticosteroids. Basal plasma corticosterone concentrations were determined for the remaining embryos. Yolk sacs were collected from each embryo and a subset was analyzed for organochlorine contaminants. Regression analysis of individual yolk sac organochlorine residue concentrations, or 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents (TEQs), with individual basal plasma corticosterone concentrations indicated statistically significant inverse relationships for polychlorinated dibenzo-p-dioxins/polychlorinated dibenzofurans (PCDDs/PCDFs), total polychlorinated biphenyls (PCBs), non-ortho PCBs, and TEQs. Similarly, inverse relationships were observed for the activities of two intermediary metabolic enzymes (phosphoenolpyruvate carboxykinase and malic enzyme) when regressed against PCDDs/PCDFs. Overall, these data suggest that current levels of organochlorine contamination may be affecting the hypothalamo-pituitary-adrenal axis and associated intermediary metabolic pathways in environmentally exposed herring gull embryos in the Great Lakes. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:10064546

Lorenzen, A; Moon, T W; Kennedy, S W; Glen, G A

1999-01-01

273

Effect of Soy Protein on Serum Lipid Profile and Some Lipid-metabolizing Enzymes in Cholesterol Fed Rats  

Microsoft Academic Search

2 Abstract: The effect of soy protein on serum lipid profile and some lipid metabolizing enzymes in rats fed with cholesterol diets was examined in this study. Rats were subjected to feeding trial over a period of six weeks on formulated diets containing: 20% soy protein with 0% cholesterol (group A); 20% soy protein with 5% cholesterol (group B), 20%

E. Chukwu Onyeneke; Olarewaju M. Oluba; Olalekan Adeyemi; George E. Eriyamremu; Samuel I. Ojeaburu; Kayode E. Adebisi; Oyeyemi Adeyemi

2007-01-01

274

Characterization of the Impact of Life Stage on Xenobiotic Metabolizing Enzyme Expression and Gene -Chemical Interactions in the Liver  

EPA Science Inventory

Differences in responses to environmental chemicals and drugs between life stages are likely due in part to differences in the expression of xenobiotic metabolizing enzymes and transporters (XMETs). We have carried out a comprehensive analysis of the mRNA expression of XMETs thro...

275

Archaeal Tetrathionate Hydrolase Goes Viral: Secretion of a Sulfur Metabolism Enzyme in the Form of Virus-Like Particles  

PubMed Central

In the course of screening for virus-host systems in extreme thermal environments, we have isolated a strain of the hyperthermophilic archaeaon Acidianus hospitalis producing unusual filamentous particles with a zipper-like appearance. The particles were shown to represent a secreted form of a genuine cellular enzyme, tetrathionate hydrolase, involved in sulfur metabolism. PMID:22636008

Krupovic, Mart; Peixeiro, Nuno; Bettstetter, Marcus; Rachel, Reinhard

2012-01-01

276

Archaeal tetrathionate hydrolase goes viral: secretion of a sulfur metabolism enzyme in the form of virus-like particles.  

PubMed

In the course of screening for virus-host systems in extreme thermal environments, we have isolated a strain of the hyperthermophilic archaeaon Acidianus hospitalis producing unusual filamentous particles with a zipper-like appearance. The particles were shown to represent a secreted form of a genuine cellular enzyme, tetrathionate hydrolase, involved in sulfur metabolism. PMID:22636008

Krupovic, Mart; Peixeiro, Nuno; Bettstetter, Marcus; Rachel, Reinhard; Prangishvili, David

2012-08-01

277

Xenobiotic Metabolizing Enzyme and Transporter Gene Expression in Primary Cultures of Human Hepatocytes Modulated by ToxCast Chemicals  

EPA Science Inventory

ToxCast chemicals were assessed for induction or suppression of xenobiotic metabolizing enzyme and transporter gene expression using primary human hepatocytes. The mRNA levels of 14 target and 2 control genes were measured: ABCB1, ABCB11, ABCG2, SLCO1B1, CYP1A1, CYP1A2, CYP2B6, C...

278

Female mice lacking active nadph-oxidase enzymes are protected against “western diet”--induced obesity and metabolic syndrome  

Technology Transfer Automated Retrieval System (TEKTRAN)

NADPH oxidase (Nox) enzymes have been implicated in regulation of adipocyte differentiation and inflammation in a variety of tissues. We examined the effects of feeding AIN-93G or a “Western diet” (WD) (45% fat, 0.5% cholesterol) on development of obesity and “metabolic syndrome” in wild type (WT) m...

279

Comparative genomic, phylogenetic, and functional investigation of the xenobiotic metabolizing arylamine N-acetyltransferase enzyme family among fungi  

Technology Transfer Automated Retrieval System (TEKTRAN)

Arylamine N-acetyltransferases (NATs) are xenobiotic metabolizing enzymes well-characterized in several bacteria and higher eukaryotes. The role of NATs in fungal biology has only recently been investigated (Glenn and Bacon, 2009; Glenn et al., 2010). The NAT1 gene of Gibberella moniliformis was the...

280

Modulation of Xenobiotic Metabolizing Enzyme and Transporter Gene Expression in Primary Cultures of Human Hepatocytes by ToxCast Chemicals  

EPA Science Inventory

ToxCast chemicals were assessed for induction or suppression of xenobiotic metabolizing enzyme and transporter gene expression using primary human hepatocytes. The mRNA levels of 14 target and 2 control genes were measured: ABCB1, ABCB11, ABCG2, SLCO1B1, CYP1A1, CYP1A2, CYP2B6, C...

281

Metabolism by rat liver cytosol of illudin S, a toxic substance of Lampteromyces japonicus. II. Characterization of illudin S-metabolizing enzyme.  

PubMed

1. Enzyme systems responsible for formation of cyclopropane ring-cleavage metabolites (M1 and M2) of illudin S in rat liver were characterized. 2. The enzymes were localized in the cytosol fraction and utilized NADPH alone as electron donor; they were not affected by oxygen and had low pH optima. 3. Formation of metabolites M1 and M2 was inhibited completely by dicumarol (10(-4) M), an inhibitor of DT-diaphorase. 4. Menadione (10(-4) M) and quercetin (10(-4) M) both inhibited formation of M1 and M2 by 35% and 15%, respectively, but quinacrine, barbital, pyrazole and p-chloromercuribenzoic acid had no significant effect. 5. Results show that the enzyme systems may differ from DT-diaphorase, aldehyde oxidase, xanthine oxidase, ketone reductase, aldose reductase, aldehyde reductase and alcohol dehydrogenase, known cytosolic enzymes responsible for xenobiotic metabolism. PMID:1377439

Tanaka, K; Inoue, T; Kadota, S; Kikuchi, T

1992-01-01

282

TM6SF2 and MAC30, new enzyme homologs in sterol metabolism and common metabolic disease  

PubMed Central

Carriers of the Glu167Lys coding variant in the TM6SF2 gene have recently been identified as being more susceptible to non-alcoholic fatty liver disease (NAFLD), yet exhibit lower levels of circulating lipids and hence are protected against cardiovascular disease. Despite the physiological importance of these observations, the molecular function of TM6SF2 remains unknown, and no sequence similarity with functionally characterized proteins has been identified. In order to trace its evolutionary history and to identify functional domains, we embarked on a computational protein sequence analysis of TM6SF2. We identified a new domain, the EXPERA domain, which is conserved among TM6SF, MAC30/TMEM97 and EBP (D8, D7 sterol isomerase) protein families. EBP mutations are the cause of chondrodysplasia punctata 2 X-linked dominant (CDPX2), also known as Conradi-Hünermann-Happle syndrome, a defective cholesterol biosynthesis disorder. Our analysis of evolutionary conservation among EXPERA domain-containing families and the previously suggested catalytic mechanism for the EBP enzyme, indicate that TM6SF and MAC30/TMEM97 families are both highly likely to possess, as for the EBP family, catalytic activity as sterol isomerases. This unexpected prediction of enzymatic functions for TM6SF and MAC30/TMEM97 is important because it now permits detailed experiments to investigate the function of these key proteins in various human pathologies, from cardiovascular disease to cancer. PMID:25566323

Sanchez-Pulido, Luis; Ponting, Chris P.

2014-01-01

283

Effects of smoking and alcohol consumption on 5-fluorouracil-related metabolic enzymes in oral squamous cell carcinoma  

PubMed Central

Lifestyle, particularly smoking and alcohol consumption, may induce and/or inhibit drug metabolism. In order to reveal the effects of smoking and alcohol consumption on the 5-fluorouracil (5-FU)-related metabolic enzymes, namely thymidylate synthase, dihydropyrimidine dehydrogenase (DPD; a sole catabolic enzyme of 5-FU), orotate phosphoribosyl transferase (OPRT) and thymidine phosphorylase, in oral squamous cell carcinomas, the mRNA expression of these enzymes was investigated in 29 surgical specimens and compared by the Brinkman index and drinking years. The surgical specimens were divided into normal and tumor regions and were independently analyzed using quantitative reverse transcription-polymerase chain reaction. There was a significantly positive correlation between DPD mRNA expression in these tissues and Brinkman index/drinking years, with OPRT mRNA expression being significantly correlated to the Brinkman index in tumor tissues. These results revealed that lifestyle habits, including smoking and alcohol consumption, may vary the activity of the 5-FU-related metabolic enzymes. DPD is the initial and rate-limiting enzyme in the catabolic pathway of 5-FU. Therefore, smoking and alcohol consumption may reduce the anticancer activity of 5-FU, possibly through the induction of DPD activity. PMID:24772313

YAMASHITA, TOMOMI; KATO, KEIZO; LONG, NGUYEN KHANH; MAKITA, HIROKI; YONEMOTO, KAZUHIRO; IIDA, KAZUKI; TAMAOKI, NARITAKA; HATAKEYAMA, DAIJIRO; SHIBATA, TOSHIYUKI

2014-01-01

284

Identification and expression of isoflavone synthase, the key enzyme for biosynthesis of isoflavones in legumes  

Microsoft Academic Search

Isoflavones have drawn much attention because of their benefits to human health. These compounds, which are produced almost exclusively in legumes, have natural roles in plant defense and root nodulation. Isoflavone synthase catalyzes the first committed step of isoflavone biosynthesis, a branch of the phenylpropanoid pathway. To identify the gene encoding this enzyme, we used a yeast expression assay to

Woosuk Jung; Oliver Yu; Sze-Mei Cindy Lau; Daniel P. O'Keefe; Joan Odell; Gary Fader; Brian McGonigle

2000-01-01

285

Distinct patterns of dysregulated expression of enzymes involved in androgen synthesis and metabolism in metastatic prostate cancer tumors  

PubMed Central

Androgen receptor (AR) signaling persists in castration-resistant prostate carcinomas (CRPCs), due to several mechanisms that include increased AR expression and intratumoral androgen metabolism. We investigated the mechanisms underlying aberrant expression of transcripts involved in androgen metabolism in CRPC. We compared gene expression profiles and DNA copy number alteration (CNA) data from 29 normal prostate tissue samples, 127 primary prostate carcinomas (PCas) and 19 metastatic PCas. Steroidogenic enzyme transcripts were evaluated by qRT-PCR in PCa cell lines and circulating tumor cells (CTCs) from CRPC patients. Metastatic PCas expressed higher transcript levels for AR and several steroidogenic enzymes, including SRD5A1, SRD5A3, and AKR1C3, while expression of SRD5A2, CYP3A4, CYP3A5 and CYP3A7 was decreased. This aberrant expression was rarely associated with CNAs. Instead, our data suggest distinct patterns of coordinated aberrant enzyme expression. Inhibition of AR activity by itself stimulated AKR1C3 expression. The aberrant expression of the steroidogenic enzyme transcripts were detected in CTCs from CRPC patients. In conclusion, our findings identify substantial interpatient heterogeneity and distinct patterns of dysregulated expression of enzymes involved in intratumoral androgen metabolism in PCa. These steroidogenic enzymes represent targets for complete suppression of systemic and intratumoral androgen levels, an objective that is supported by the clinical efficacy of the CYP17 inhibitor abiraterone. A comprehensive AR axis targeting approach via simultaneous, frontline enzymatic blockade and/or transcriptional repression of several steroidogenic enzymes, in combination with GnRH analogs and potent anti-androgens, would represent a powerful future strategy for PCa management. PMID:22971343

Mitsiades, Nicholas; Sung, Clifford C.; Schultz, Nikolaus; Danila, Daniel C.; He, Bin; Eedunuri, Vijay Kumar; Fleisher, Martin; Sander, Chris; Sawyers, Charles L.; Scher, Howard I.

2012-01-01

286

Nicotinamide Adenine Dinucleotide-specific “Malic” Enzyme in Kalanchoë daigremontiana and Other Plants Exhibiting Crassulacean Acid Metabolism 1  

PubMed Central

NAD-specific “malic” enzyme (EC 1.1.1.39) has been isolated and purified 1200-fold from leaves of Kalanchoë daigremontiana. Kinetic studies of this enzyme, which is activated 14-fold by CoA, acetyl-CoA, and SO42?, suggest allosteric properties. Cofactor requirements show an absolute specificity for NAD and for Mn2+, which cannot be replaced by NADP or Mg2+. For maintaining enzyme activity in crude leaf extracts a thiol reagent, Mn2+, and PVP-40 were required. The latter could be omitted from purified preparations. By sucrose density gradient centrifugation NAD-malic enzyme could be localized in mitochondria. A survey of plants with crassulacean acid metabolism revealed the presence of NAD-malic enzyme in all 31 plants tested. Substantial levels of this enzyme (121-186 ?mole/hr·mg of Chl) were detected in all members tested of the family Crassulaceae. It is proposed that NAD-malic enzyme in general supplements activity of NADP-malic enzyme present in these plants and may be specifically employed to increase internal concentrations of CO2 for recycling during cessation of gas exchange in periods of severe drought. PMID:16659473

Dittrich, Peter

1976-01-01

287

Temperature during embryonic development has persistent effects on metabolic enzymes in the muscle of zebrafish.  

PubMed

Global warming is intensifying interest in the physiological consequences of temperature change in ectotherms, but we still have a relatively poor understanding of the effects of temperature on early life stages. This study determined how embryonic temperature (TE) affects development and the activity of metabolic enzymes in the swimming muscle of zebrafish. Embryos developed successfully to hatching (survival ? 88%) from 22 to 32°C, but suffered sharp increases in mortality outside of this range. Embryos that were incubated until hatching at a control TE (27°C) or near the extremes for successful development (22 or 32°C) were next raised to adulthood under control conditions at 27°C. Growth trajectories after hatching were altered in the 22°C and 32°C TE groups compared with 27°C TE controls, but growth slowed after 3 months of age in all groups. Maximal enzyme activities of cytochrome c oxidase (COX), citrate synthase (CS), hydroxyacyl-coA dehydrogenase (HOAD), pyruvate kinase (PK) and lactate dehydrogenase (LDH) were measured across a range of assay temperatures (22, 27, 32 and 36°C) in adults from each TE group that were acclimated to 27 or 32°C. Substrate affinities (Km) were also determined for COX and LDH. In adult fish acclimated to 27°C, COX and PK activities were higher in 22°C and 32°C TE groups than in 27°C TE controls, and the temperature optimum for COX activity was higher in the 32°C TE group than in the 22°C TE group. Warm acclimation reduced COX, CS and/or PK activities in the 22 and 32°C TE groups, possibly to compensate for thermal effects on molecular activity. This response did not occur in the 27°C TE controls, which instead increased LDH and HOAD activities. Warm acclimation also increased thermal sensitivity (Q10) of HOAD to cool temperatures across all TE groups. We conclude that the temperature experienced during early development can have a persistent impact on energy metabolism pathways and acclimation capacity in later life. PMID:24363419

Schnurr, Meghan E; Yin, Yi; Scott, Graham R

2014-04-15

288

Coordinated changes in xenobiotic metabolizing enzyme gene expression in aging male rats.  

PubMed

In order to gain insight into the effects of aging on susceptibility to environmental toxins, we characterized the expression of xenobiotic metabolizing enzymes (XMEs) from the livers of male F344 and Brown Norway (BN) rats across the adult lifespan. Using full-genome Affymetrix arrays, principal component analysis showed a clear age-dependent separation between young and old animals in both rat strains. Out of 1135 or 1435 genes altered between the old and young groups in the F344 or BN rats, 7 or 3% were XMEs and included members of the phase I, II, and III classes of genes. There was a 20 or 32% overlap in the gene expression profile between the two strains for F344 or BN, respectively. Lipid, ergosterol, alcohol, and fatty acid metabolism genes were also altered with age in both strains. Some of the genes altered by age exhibited a gender-dependent expression pattern in young adult rats, suggesting an increasingly feminized pattern of gene expression with age in male rats. To examine transcriptional responses across lifespan after challenge with a xenobiotic compound, BN rats were exposed to toluene by oral gavage. Toluene exposure decreased the expression of glutathione synthetase, and dramatically increased the number of phase III genes being downregulated. The expression of CYP2B2 and glutathione-S-transferase decreased with age but increased in all age groups after toluene exposure. Decreased ability to detoxify and transport chemicals out of the body with age could result in increased susceptibility to some classes of chemicals in the aging population. PMID:18653662

Lee, Janice S; Ward, William O; Wolf, Douglas C; Allen, James W; Mills, Camilla; DeVito, Michael J; Corton, J Christopher

2008-11-01

289

Hepatic Xenobiotic Metabolizing Enzyme and Transporter Gene Expression through the Life Stages of the Mouse  

PubMed Central

Background Differences in responses to environmental chemicals and drugs between life stages are likely due in part to differences in the expression of xenobiotic metabolizing enzymes and transporters (XMETs). No comprehensive analysis of the mRNA expression of XMETs has been carried out through life stages in any species. Results Using full-genome arrays, the mRNA expression of all XMETs and their regulatory proteins was examined during fetal (gestation day (GD) 19), neonatal (postnatal day (PND) 7), prepubescent (PND32), middle age (12 months), and old age (18 and 24 months) in the C57BL/6J (C57) mouse liver and compared to adults. Fetal and neonatal life stages exhibited dramatic differences in XMET mRNA expression compared to the relatively minor effects of old age. The total number of XMET probe sets that differed from adults was 636, 500, 84, 5, 43, and 102 for GD19, PND7, PND32, 12 months, 18 months and 24 months, respectively. At all life stages except PND32, under-expressed genes outnumbered over-expressed genes. The altered XMETs included those in all of the major metabolic and transport phases including introduction of reactive or polar groups (Phase I), conjugation (Phase II) and excretion (Phase III). In the fetus and neonate, parallel increases in expression were noted in the dioxin receptor, Nrf2 components and their regulated genes while nuclear receptors and regulated genes were generally down-regulated. Suppression of male-specific XMETs was observed at early (GD19, PND7) and to a lesser extent, later life stages (18 and 24 months). A number of female-specific XMETs exhibited a spike in expression centered at PND7. Conclusions The analysis revealed dramatic differences in the expression of the XMETs, especially in the fetus and neonate that are partially dependent on gender-dependent factors. XMET expression can be used to predict life stage-specific responses to environmental chemicals and drugs. PMID:21931700

Lee, Janice S.; Ward, William O.; Liu, Jie; Ren, Hongzu; Vallanat, Beena; Delker, Don; Corton, J. Christopher

2011-01-01

290

Lipid metabolism enzyme ACSVL3 supports glioblastoma stem cell maintenance and tumorigenicity  

PubMed Central

Background Targeting cell metabolism offers promising opportunities for the development of drugs to treat cancer. We previously found that the fatty acyl-CoA synthetase VL3 (ACSVL3) is elevated in malignant brain tumor tissues and involved in tumorigenesis. This study investigates the role of ACSVL3 in the maintenance of glioblastoma multiforme (GBM) stem cell self-renewal and the capacity of GBM stem cells to initiate tumor xenograft formation. Methods We examined ACSVL3 expression during differentiation of several GBM stem cell enriched neurosphere cultures. To study the function of ACSVL3, we performed loss-of-function by using small interfering RNAs to target ACSVL3 and examined stem cell marker expression, neurosphere formation and tumor initiation properties. Results ACSVL3 expression levels were substantially increased in GBM stem cell enriched neurosphere cultures and decreased after differentiation of the neurospheres. Down-regulating ACSVL3 with small inhibiting RNAs decreased the expression of markers and regulators associated with stem cell self-renewal, including CD133, ALDH, Musashi-1 and Sox-2. ACSVL3 knockdown in neurosphere cells led to increased expression of differentiation markers GFAP and Tuj1. Furthermore, ACSVL3 knockdown reduced anchorage-independent neurosphere cell growth, neurosphere-forming capacity as well as self-renewal of these GBM stem cell enriched neurosphere cultures. In vivo studies revealed that ACSVL3 loss-of-function substantially inhibited the ability of neurosphere cells to propagate orthotopic tumor xenografts. A link between ACSVL3 and cancer stem cell phenotype was further established by the findings that ACSVL3 expression was regulated by receptor tyrosine kinase pathways that support GBM stem cell self-renewal and tumor initiation, including EGFR and HGF/c-Met pathways. Conclusions Our findings indicate that the lipid metabolism enzyme ACSVL3 is involved in GBM stem cell maintenance and the tumor-initiating capacity of GBM stem cell enriched-neurospheres in animals. PMID:24893952

2014-01-01

291

Carboxylation of phenylphosphate by phenol carboxylase, an enzyme system of anaerobic phenol metabolism.  

PubMed Central

Several lines of evidence indicate that the first step in the anaerobic metabolism of phenol is phenol carboxylation to 4-hydroxybenzoate; this reaction is considered a biological Kolbe-Schmitt carboxylation. A phenol carboxylase system was characterized by using a denitrifying Pseudomonas strain, K 172, which catalyzes an isotope exchange between 14CO2 and the carboxyl group of 4-hydroxybenzoate. The enzymatic isotope exchange activity (100 nmol min-1 mg-1 of protein) requires Mn2+ and K+. We show that this system also catalyzes the carboxylation of phenylphosphate (the phosphoric acid monophenyl ester) to 4-hydroxybenzoate and phosphate. The specific activity of phenylphosphate carboxylation at the optimal pH of 6.5 is 12 nmol of CO2 fixed min-1 mg-1 of protein. Phenylphosphate cannot be replaced by Mg(2+)-ATP and phenol. The carboxylase activity requires Mn2+ but, in contrast to the isotope exchange activity, does not require K+. The apparent Km values are 1.5 mM dissolved CO2 and 0.2 mM phenylphosphate. Several convenient assays for phenylophosphate carboxylation are described. The isotope exchange reaction and the net carboxylation reaction are catalyzed by the same oxygen-sensitive enzyme, which has a half-life in an air-saturated solution of less than 1 min. Both activities cochromatographed with a protein with a Mr of 280,000, and both activities were induced only after anaerobic growth on phenol. The carboxylation of phenylphosphate suggests that phenylphosphate itself is the physiological CO2 acceptor molecular of this novel CO2 fixation reaction. Alternatively, phenylphosphate could simulate the unknown natural precursor. It is suggested that the formation of an enzyme-bound phenolate anion from the activated phenolic compound is the rate-determining step in the carboxylation reaction. PMID:1592817

Lack, A; Fuchs, G

1992-01-01

292

Polymorphisms in carcinogen metabolism enzymes, fish intake, and risk of prostate cancer  

PubMed Central

Cooking fish at high temperature can produce potent carcinogens such as heterocyclic amines and polycyclic aromatic hydrocarbons. The effects of these carcinogens may undergo modification by the enzymes responsible for their detoxification and/or activation. In this study, we investigated genetic polymorphisms in nine carcinogen metabolism enzymes and their modifying effects on the association between white or dark fish consumption and prostate cancer (PCA) risk. We genotyped 497 localized and 936 advanced PCA cases and 760 controls from the California Collaborative Case–Control Study of Prostate Cancer. Three polymorphisms, EPHX1 Tyr113His, CYP1B1 Leu432Val and GSTT1 null/present, were associated with localized PCA risk. The PTGS2 765 G/C polymorphism modified the association between white fish consumption and advanced PCA risk (interaction P 5 0.002), with high white fish consumption being positively associated with risk only among carriers of the C allele. This effect modification by PTGS2 genotype was stronger when restricted to consumption of well-done white fish (interaction P 5 0.021). These findings support the hypotheses that changes in white fish brought upon by high-temperature cooking methods, such as carcinogen accumulation and/or fatty acid composition changes, may contribute to prostate carcinogenesis. However, the gene–diet interactions should be interpreted with caution given the limited sample size. Thus, our findings require further validation with additional studies. Abbreviations: AA African American; BMI body mass index; CI confidence interval; CNV copy number variant; EPIC European Prospective Investigation into Cancer and Nutrition; HCA heterocyclic amine; HCFA Health Care Financing Administration; LAC Los Angeles county; MAF minor allele frequency; NHW non-Hispanic White; OR odds ratio; PAH polycyclic aromatic hydrocarbon; PCA prostate cancer; PTGS2 prostaglandin- endoperoxide synthase 2; PUFA polyunsaturated fatty acids; RDD random-digit dialing; SEER Surveillance, Epidemiology, and End Result; SES socio-economic status; SFBA San Francisco Bay Area; SNP single-nucleotide polymorphism PMID:22610071

Stern, Mariana C.

2012-01-01

293

Arachidonic Acid-metabolizing Cytochrome P450 Enzymes Are Targets of ?-3 Fatty Acids*  

PubMed Central

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) protect against cardiovascular disease by largely unknown mechanisms. We tested the hypothesis that EPA and DHA may compete with arachidonic acid (AA) for the conversion by cytochrome P450 (CYP) enzymes, resulting in the formation of alternative, physiologically active, metabolites. Renal and hepatic microsomes, as well as various CYP isoforms, displayed equal or elevated activities when metabolizing EPA or DHA instead of AA. CYP2C/2J isoforms converting AA to epoxyeicosatrienoic acids (EETs) preferentially epoxidized the ?-3 double bond and thereby produced 17,18-epoxyeicosatetraenoic (17,18-EEQ) and 19,20-epoxydocosapentaenoic acid (19,20-EDP) from EPA and DHA. We found that these ?-3 epoxides are highly active as antiarrhythmic agents, suppressing the Ca2+-induced increased rate of spontaneous beating of neonatal rat cardiomyocytes, at low nanomolar concentrations. CYP4A/4F isoforms ?-hydroxylating AA were less regioselective toward EPA and DHA, catalyzing predominantly ?- and ? minus 1 hydroxylation. Rats given dietary EPA/DHA supplementation exhibited substantial replacement of AA by EPA and DHA in membrane phospholipids in plasma, heart, kidney, liver, lung, and pancreas, with less pronounced changes in the brain. The changes in fatty acids were accompanied by concomitant changes in endogenous CYP metabolite profiles (e.g. altering the EET/EEQ/EDP ratio from 87:0:13 to 27:18:55 in the heart). These results demonstrate that CYP enzymes efficiently convert EPA and DHA to novel epoxy and hydroxy metabolites that could mediate some of the beneficial cardiovascular effects of dietary ?-3 fatty acids. PMID:20732876

Arnold, Cosima; Markovic, Marija; Blossey, Katrin; Wallukat, Gerd; Fischer, Robert; Dechend, Ralf; Konkel, Anne; von Schacky, Clemens; Luft, Friedrich C.; Muller, Dominik N.; Rothe, Michael; Schunck, Wolf-Hagen

2010-01-01

294

(CANCER RESEARCH 48, 1390-1397, March 15, 1988] Epoxide-metabolizing Enzymes in Mammary Gland and Liver from BALB/c Mice  

E-print Network

(CANCER RESEARCH 48, 1390-1397, March 15, 1988] Epoxide-metabolizing Enzymes in Mammary Gland and Liver from BALB/c Mice and Effects of Inducers on Enzyme Activity1 Marilyn H. Suva, Roger N. Wixtrom.3.23) (EH) are hydrolytic enzymes which may play an important role in the activation and detoxification

Hammock, Bruce D.

295

[Comparative efficacy and safety of contemporary Angiotensin converting enzyme inhibitors moexipril and spirapril in women with postmenopausal metabolic syndrome].  

PubMed

Moexipril (7.4-15 mg/day) was given to 34, spirapril (3-6 mg/day) -- to 18 postmenopausal women with hypertension and metabolic syndrome for 16 weeks. Hydrochlorthiazide was added when therapy was not sufficiently effective. Both angiotensin converting enzyme inhibitors had similar hypotensive activity: blood pressure normalized in 71 and 61% of moexipril and spirapril treated women, respectively. Both drugs promoted normalization of metabolism of lipid (lowering of levels of cholesterol, atherogenic lipoproteins and triglycerides) and carbohydrates (lowering of hyperinsulinemia). Patients with postmenopausal metabolic syndrome had elevation of leptin level up to 27.5+/-5.5 pg/ml. Moexipril and spirapril caused lowering of elevated levels of leptin. These drugs did not affect levels of sex hormones. They exerted vasoprotective (normalization of endothelium dependent and independent vasodilatation) and nephroprotective (attenuation and normalization of microalbuminuria) effects. Thus spirapril and moexipril are effective in treatment of hypertension in patients with postmenopausal metabolic syndrome. PMID:16474309

Leonova, M V; Demidova, M A; Tarasov, A V; Belousov, Iu B

2006-01-01

296

Mycobacterium tuberculosis Glucosyl-3-Phosphoglycerate Synthase: Structure of a Key Enzyme in Methylglucose Lipopolysaccharide Biosynthesis  

PubMed Central

Tuberculosis constitutes today a serious threat to human health worldwide, aggravated by the increasing number of identified multi-resistant strains of Mycobacterium tuberculosis, its causative agent, as well as by the lack of development of novel mycobactericidal compounds for the last few decades. The increased resilience of this pathogen is due, to a great extent, to its complex, polysaccharide-rich, and unusually impermeable cell wall. The synthesis of this essential structure is still poorly understood despite the fact that enzymes involved in glycosidic bond synthesis represent more than 1% of all M. tuberculosis ORFs identified to date. One of them is GpgS, a retaining glycosyltransferase (GT) with low sequence homology to any other GTs of known structure, which has been identified in two species of mycobacteria and shown to be essential for the survival of M. tuberculosis. To further understand the biochemical properties of M. tuberculosis GpgS, we determined the three-dimensional structure of the apo enzyme, as well as of its ternary complex with UDP and 3-phosphoglycerate, by X-ray crystallography, to a resolution of 2.5 and 2.7 Ĺ, respectively. GpgS, the first enzyme from the newly established GT-81 family to be structurally characterized, displays a dimeric architecture with an overall fold similar to that of other GT-A-type glycosyltransferases. These three-dimensional structures provide a molecular explanation for the enzyme's preference for UDP-containing donor substrates, as well as for its glucose versus mannose discrimination, and uncover the structural determinants for acceptor substrate selectivity. Glycosyltransferases constitute a growing family of enzymes for which structural and mechanistic data urges. The three-dimensional structures of M. tuberculosis GpgS now determined provide such data for a novel enzyme family, clearly establishing the molecular determinants for substrate recognition and catalysis, while providing an experimental scaffold for the structure-based rational design of specific inhibitors, which lay the foundation for the development of novel anti-tuberculosis therapies. PMID:19015727

Pereira, Pedro José Barbosa; Empadinhas, Nuno; Albuquerque, Luciana; Sá-Moura, Bebiana; da Costa, Milton S.; Macedo-Ribeiro, Sandra

2008-01-01

297

[Effects of exogenous NO3- on cherry root function and enzyme activities related to nitrogen metabolism under hypoxia stress].  

PubMed

A water culture experiment with controlled dissolved oxygen concentration was conducted to explore the effects of exogenous NO3- on the root function and enzyme activities related to nitrogen metabolism of cherry (Prunun cerasus x P. canescens) seedlings under hypoxia stress. Comparing with the control (7.5 mmol NO3- x L(-1)), treatments 15 and 22.5 mmol NO3- x L(-1) made the materials for plant metabolism abundant, ensured the synthesis of enzyme proteins, increased root activity, maintained root respiration, improved the activities of enzymes related to nitrogen metabolism, such as nitrate reductase (NR), glutamine synthethase (GS), and glutamate dehydrogenase (NADH-GDH) in roots, and thereby, supplied enough energy for root respiration and NAD+ to glycolytic pathway, ensured electron transfer, and avoid ammonium toxicity under hypoxia stress. As a result, the injury of hypoxia stress to cherry plant was alleviated. Applying NO3- at the concentration of 22.5 mmol x L(-1) was more advisable. However, NO3- deficiency (0 mmol x L(-1)) showed opposite results. The above results suggested that applying exogenous NO3- to growth medium could regulate cherry root function and nitrogen metabolism, and antagonize the damage of hypoxia stress on cherry roots. PMID:21443020

Feng, Li-guo; Sheng, Li-xi; Shu, Huai-rui

2010-12-01

298

Genome-wide identification of gibberellins metabolic enzyme genes and expression profiling analysis during seed germination in maize.  

PubMed

Gibberellin (GA) is an essential phytohormone that controls many aspects of plant development. To enhance our understanding of GA metabolism in maize, we intensively screened and identified 27 candidate genes encoding the seven GA metabolic enzymes including ent-copalyl diphosphate synthase (CPS), ent-kaurene synthase (KS), ent-kaurene oxidase (KO), ent-kaurenoic acid oxidase (KAO), GA 20-oxidase (GA20ox), GA 3-oxidase (GA3ox), and GA 2-oxidase (GA2ox), using all available public maize databases. The results indicate that maize genome contains three CPS, four KS, two KO and one KAO genes, and most of them are arranged separately on the maize genome, which differs from that in rice. In addition, the enzymes catalyzing the later steps (ZmGA20ox, ZmGA3ox and ZmGA2ox) are also encoded by gene families in maize, but GA3ox enzyme is likely to be encoded by single gene. Expression profiling analysis exhibited that transcripts of 15 GA metabolic genes could be detected during maize seed germination, which provides further evidence for the notion that increased synthesis of active GA in the embryo is required for triggering germination events. Moreover, a variety of temporal genes expression patterns of GA metabolic genes were detected, which revealed the complexity of underlying mechanism for GA regulated seed germination. PMID:21640170

Song, Jian; Guo, Baojian; Song, Fangwei; Peng, Huiru; Yao, Yingyin; Zhang, Yirong; Sun, Qixin; Ni, Zhongfu

2011-08-15

299

Pyruvate decarboxylase and alcohol dehydrogenase overexpression in Escherichia coli resulted in high ethanol production and rewired metabolic enzyme networks.  

PubMed

Pyruvate decarboxylase and alcohol dehydrogenase are efficient enzymes for ethanol production in Zymomonas mobilis. These two enzymes were over-expressed in Escherichia coli, a promising candidate for industrial ethanol production, resulting in high ethanol production in the engineered E. coli. To investigate the intracellular changes to the enzyme overexpression for homoethanol production, 2-DE and LC-MS/MS were performed. More than 1,000 protein spots were reproducibly detected in the gel by image analysis. Compared to the wild-type, 99 protein spots showed significant changes in abundance in the recombinant E. coli, in which 46 were down-regulated and 53 were up-regulated. Most proteins related to tricarboxylic acid cycle, glycerol metabolism and other energy metabolism were up-regulated, whereas proteins involved in glycolysis and glyoxylate pathway were down-regulated, indicating the rewired metabolism in the engineered E. coli. As glycolysis is the main pathway for ethanol production, and it was inhibited significantly in engineered E. coli, further efforts should be directed at minimizing the repression of glycolysis to optimize metabolism network for higher yields of ethanol production. PMID:25217026

Yang, Mingfeng; Li, Xuefeng; Bu, Chunya; Wang, Hui; Shi, Guanglu; Yang, Xiushan; Hu, Yong; Wang, Xiaoqin

2014-11-01

300

Enzymic synthesis of indole-3-acetyl-1-O-beta-d-glucose. II. Metabolic characteristics of the enzyme  

NASA Technical Reports Server (NTRS)

The synthesis of indole-3-acetyl-1-O-beta-D-glucose from indole-3-acetic acid (IAA) and uridine diphosphoglucose (UDPG) has been shown to be a reversible reaction with the equilibrium away from ester formation and toward formation of IAA. The enzyme occurs primarily in the liquid endosperm of the corn kernel but some activity occurs in the embryo. It is relatively specific showing no glucose ester formation with oxindole-3-acetic acid or 7-hydroxy-oxindole-3-acetic acid, and low activity with phenylpropene acids, such as rho-coumaric acid. The enzyme is also specific for the nucleotide sugar showing no activity with UDPGalactose or UDPXylose. The enzyme is inhibited by inorganic pyrophosphate, by phosphate esters and by phospholipids, particularly phosphatidyl ethanolamine. The enzyme is inhibited by zeatin, by 2,4-dichlorophenoxy-acetic acid, by IAA-myo-inositol and IAA-glucan, but not by zeatin riboside, and only weakly by gibberellic acid, abscisic acid and kinetin. The reaction is slightly stimulated by both calcium and calmodulin and, in some cases, by thiol compounds. The role of this enzyme in the homeostatic control of indole-3-acetic acid levels in Zea mays is discussed.

Leznicki, A. J.; Bandurski, R. S.

1988-01-01

301

Sequence and organization of genes encoding enzymes involved in pyruvate metabolism in Mycoplasma capricolum.  

PubMed Central

The region of the genome of Mycoplasma capricolum upstream of the portion encompassing the genes for Enzymes I and IIAglc of the phosphoenolpyruvate:sugar phosphotransferase system (PTS) was cloned and sequenced. Examination of the sequence revealed open reading frames corresponding to numerous genes involved with the oxidation of pyruvate. The deduced gene organization is naox (encoding NADH oxidase)-lplA (encoding lipoate-protein ligase)-odpA (encoding pyruvate dehydrogenase EI alpha)-odpB (encoding pyruvate dehydrogenase EI beta)-odp2(encoding pyruvate dehydrogenase EII)-dldH (encoding dihydrolipoamide dehydrogenase)-pta (encoding phosphotransacetylase)-ack (encoding acetate kinase)-orfA (an unknown open reading frame)-kdtB-ptsI-crr. Analysis of the DNA sequence suggests that the naox and lplA genes are part of a single operon, odpA and odpB constitute an additional operon, odp2 and dldH a third operon, and pta and ack an additional transcription unit. Phylogenetic analyses of the protein products of the odpA and odpB genes indicate that they are most similar to the corresponding proteins from Mycoplasma genitalium, Acholeplasma laidlawii, and Gram-positive organisms. The product of the odp2 gene contains a single lipoyl domain, as is the case with the corresponding proteins from M. genitalium and numerous other organisms. An evolutionary tree places the M. capricolum odp2 gene product in close relationship to the corresponding proteins from A. laidlawii and M.genitalium. The dldH gene encodes an unusual form of dihydrolipoamide dehydrogenase that contains an aminoterminal extension corresponding to a lipoyl domain, a property shared by the corresponding proteins from Alcaligenes eutrophus and Clostridium magnum. Aside from that feature, the protein is related phylogenetically to the corresponding proteins from A. laidlawii and M. genitalium. The phosphotransacetylase from M. capricolum is related most closely to the corresponding protein from M. genitalium and is distinguished easily from the enzymes from Escherichia coli and Haemophilus influenzae by the absence of the characteristic amino-terminal extension. The acetate kinase from M. capricolum is related evolutionarily to the homologous enzyme from M. genitalium. Map position comparisons of genes encoding proteins involved with pyruvate metabolism show that, whereas all the genes are clustered in M. capricolum, they are scattered in M. genitalium. PMID:8844861

Zhu, P. P.; Peterkofsky, A.

1996-01-01

302

Effect of increasing body condition on key regulators of fat metabolism in subcutaneous adipose tissue depot and circulation of nonlactating dairy cows.  

PubMed

In response to negative energy balance, overconditioned cows mobilize more body fat than thin cows and subsequently are prone to develop metabolic disorders. Changes in adipose tissue (AT) metabolism are barely investigated in overconditioned cows. Therefore, the objective was to investigate the effect of increasing body condition on key regulator proteins of fat metabolism in subcutaneous AT and circulation of dairy cows. Nonlactating, nonpregnant dairy cows (n=8) investigated in the current study served as a model to elucidate the changes in the course of overcondition independent from physiological changes related to gestation, parturition, and lactation. Cows were fed diets with increasing portions of concentrate during the first 6wk of the experiment until 60% were reached, which was maintained for 9wk. Biopsy samples from AT of the subcutaneous tailhead region were collected every 8wk, whereas blood was sampled monthly. Within the experimental period cows had an average BW gain of 243±33.3 kg. Leptin and insulin concentrations were increased until wk 12. Based on serum concentrations of glucose, insulin, and nonesterified fatty acids, the surrogate indices for insulin sensitivity were calculated. High-concentrate feeding led to decreased quantitative insulin sensitivity check index and homeostasis model assessment due to high insulin and glucose concentrations indicating decreased insulin sensitivity. Adiponectin, an adipokine-promoting insulin sensitivity, decreased in subcutaneous AT, but remained unchanged in the circulation. The high-concentrate diet affected key enzymes reflecting AT metabolism such as AMP-activated protein kinase and hormone-sensitive lipase, both represented as the proportion of the phosphorylated protein to total protein, as well as fatty acid synthase. The extent of phosphorylation of AMP-activated protein kinase and the protein expression of fatty acid synthase were inversely regulated throughout the experimental period, whereas the extent of phosphorylation of hormone-sensitive lipase was consistently decreasing by the high-concentrate diet. Overcondition in nonpregnant, nonlactating dairy cows changed the expression of key regulator proteins of AT metabolism and circulation accompanied by impaired insulin sensitivity, which might increase the risk for metabolic disorders. PMID:25497790

Locher, L; Häussler, S; Laubenthal, L; Singh, S P; Winkler, J; Kinoshita, A; Kenéz, Á; Rehage, J; Huber, K; Sauerwein, H; Dänicke, S

2015-02-01

303

Expression pattern of glycoside hydrolase genes in Lutzomyia longipalpis reveals key enzymes involved in larval digestion  

PubMed Central

The sand fly Lutzomyia longipalpis is the most important vector of American Visceral Leishmaniasis. Adults are phytophagous (males and females) or blood feeders (females only), and larvae feed on solid detritus. Digestion in sand fly larvae has scarcely been studied, but some glycosidase activities putatively involved in microorganism digestion were already described. Nevertheless, the molecular nature of these enzymes, as the corresponding genes and transcripts, were not explored yet. Catabolism of microbial carbohydrates in insects generally involves ?-1,3-glucanases, chitinases, and digestive lysozymes. In this work, the transcripts of digestive ?-1,3-glucanase and chitinases were identified in the L. longipalpis larvae throughout analysis of sequences and expression patterns of glycoside hydrolases families 16, 18, and 22. The activity of one i-type lysozyme was also registered. Interestingly, this lysozyme seems to play a role in immunity, rather than digestion. This is the first attempt to identify the molecular nature of sand fly larval digestive enzymes. PMID:25140153

Moraes, Caroline da Silva; Diaz-Albiter, Hector M.; Faria, Maiara do Valle; Sant'Anna, Maurício R. V.; Dillon, Rod J.; Genta, Fernando A.

2014-01-01

304

Age-related alteration of PKC, a key enzyme in memory processes  

Microsoft Academic Search

Brain aging is characterized by a progressive decline of the cognitive and memory functions. It is becoming increasingly clear\\u000a that protein phosphorylation and, in particular, the activity of the calcium-phospholipid-dependent protein kinase C (PKC)\\u000a may be one of the fundamental cellular changes associated with memory function. PKC is a multigene family of enzymes highly\\u000a expressed in brain tissues. The activation

A. Pascale; S. Govoni; F. Battaini

1998-01-01

305

Quantitative studies of enzyme-substrate compartmentation, functional coupling and metabolic channelling in muscle cells  

Microsoft Academic Search

Some historical aspects of development of the concepts of functional coupling, metabolic channelling, compartmentation and energy transfer networks are reviewed. Different quantitative approaches, including kinetic and mathematical modeling of energy metabolism, intracellular energy transfer and metabolic regulation of energy production and fluxes in the cells in vivo are analyzed. As an example of the system with metabolic channelling, thermodynamic aspects

Valdur Saks; Pierre Dos Santos; Frank N. Gellerich; Philippe Diolez

1998-01-01

306

The evolution of pyrroline-5-carboxylate synthase in plants: a key enzyme in proline synthesis.  

PubMed

Many plants synthesize and accumulate proline in response to osmotic stress conditions. A central enzyme in the proline biosynthesis is the bifunctional enzyme Delta(1)-pyrroline-5-carboxylate synthase (P5CS) that includes two functional catalytic domains: the gamma-glutamyl kinase and the glutamic-gamma-semialdehyde dehydrogenase. This enzyme catalyzes the first two steps of the proline biosynthetic pathway and plays a central role in the regulation of this process in plants. To determine the evolutionary events that occurred in P5CS genes, partial sequences from four Neotropical trees were cloned and compared to those of other plant taxa. Molecular phylogenetic analysis indicated that P5CS duplication events have occurred several times following the emergence of flowering plants and at different frequencies throughout the evolution of monocots and dicots. Despite the high number of conserved residues in plant P5CS sequences, positive selection was observed at different regions of P5CS paralogous genes and also when dicots and monocots were contrasted. PMID:19002717

Turchetto-Zolet, Andreia Carina; Margis-Pinheiro, Marcia; Margis, Rogerio

2009-01-01

307

A Metabolic Enzyme as a Primary Virulence Factor of Mycoplasma mycoides subsp. mycoides Small Colony  

PubMed Central

During evolution, pathogenic bacteria have developed complex interactions with their hosts. This has frequently involved the acquisition of virulence factors on pathogenicity islands, plasmids, transposons, or prophages, allowing them to colonize, survive, and replicate within the host. In contrast, Mycoplasma species, the smallest self-replicating organisms, have regressively evolved from gram-positive bacteria by reduction of the genome to a minimal size, with the consequence that they have economized their genetic resources. Hence, pathogenic Mycoplasma species lack typical primary virulence factors such as toxins, cytolysins, and invasins. Consequently, little is known how pathogenic Mycoplasma species cause host cell damage, inflammation, and disease. Here we identify a novel primary virulence determinant in Mycoplasma mycoides subsp. mycoides Small Colony (SC), which causes host cell injury. This virulence factor, released in significant amounts in the presence of glycerol in the growth medium, consists of toxic by-products such as H2O2 formed by l-?-glycerophosphate oxidase (GlpO), a membrane-located enzyme that is involved in the metabolism of glycerol. When embryonic calf nasal epithelial cells are infected with M. mycoides subsp. mycoides SC in the presence of physiological amounts of glycerol, H2O2 is released inside the cells prior to cell death. This process can be inhibited with monospecific anti-GlpO antibodies. PMID:16166545

Pilo, Paola; Vilei, Edy M.; Peterhans, Ernst; Bonvin-Klotz, Laetitia; Stoffel, Michael H.; Dobbelaere, Dirk; Frey, Joachim

2005-01-01

308

A metabolic enzyme as a primary virulence factor of Mycoplasma mycoides subsp. mycoides small colony.  

PubMed

During evolution, pathogenic bacteria have developed complex interactions with their hosts. This has frequently involved the acquisition of virulence factors on pathogenicity islands, plasmids, transposons, or prophages, allowing them to colonize, survive, and replicate within the host. In contrast, Mycoplasma species, the smallest self-replicating organisms, have regressively evolved from gram-positive bacteria by reduction of the genome to a minimal size, with the consequence that they have economized their genetic resources. Hence, pathogenic Mycoplasma species lack typical primary virulence factors such as toxins, cytolysins, and invasins. Consequently, little is known how pathogenic Mycoplasma species cause host cell damage, inflammation, and disease. Here we identify a novel primary virulence determinant in Mycoplasma mycoides subsp. mycoides Small Colony (SC), which causes host cell injury. This virulence factor, released in significant amounts in the presence of glycerol in the growth medium, consists of toxic by-products such as H2O2 formed by l-alpha-glycerophosphate oxidase (GlpO), a membrane-located enzyme that is involved in the metabolism of glycerol. When embryonic calf nasal epithelial cells are infected with M. mycoides subsp. mycoides SC in the presence of physiological amounts of glycerol, H2O2 is released inside the cells prior to cell death. This process can be inhibited with monospecific anti-GlpO antibodies. PMID:16166545

Pilo, Paola; Vilei, Edy M; Peterhans, Ernst; Bonvin-Klotz, Laetitia; Stoffel, Michael H; Dobbelaere, Dirk; Frey, Joachim

2005-10-01

309

Hydroxylated PAHs in bile of deep-sea fish. Relationship with xenobiotic metabolizing enzymes  

SciTech Connect

Polycyclic aromatic hydrocarbon (PAH) pollution in deep-sea environments has been assessed by measuring bile PAH metabolites in deep-sea fish. Five species from the NW Mediterranean were selected for the study: Coryphaenoides guentheri, Lepidion lepidion, Mora moro, Bathypterois mediterraneus, and Alepocephalus rostratus. Bile crude samples were directly analyzed by HPLC-fluorescence at the excitation/emission wavelengths of benzo[a]pyrene. Differences among sampling sites were recorded, which suggests that coastal discharges of contaminants may reach these remote areas. Subsequently, a number of bile samples were hydrolyzed and analyzed by gas chromatography--mass spectrometry (GC-MS) for the determination of individual PAHs. 1-Pyrenol and 2-phenylphenol were among the most abundant compounds detected. The results obtained confirm the long-range transport of PAHs to deep-sea environments, subsequent exposure of fish inhabiting those remote areas, and its ability to metabolize and excrete them through the bile. The data also describe hepatic enzymes (cytochrome P450 and glutathione S-transferases) that appear to be as catalytically efficient as those in shallow water species.

Escartin, E.; Porte, C. [IIQAB-CSIC, Barcelona (Spain). Environmental Chemistry Dept.] [IIQAB-CSIC, Barcelona (Spain). Environmental Chemistry Dept.

1999-08-15

310

Structural Insights into Maize Viviparous14, a Key Enzyme in the Biosynthesis of the Phytohormone Abscisic Acid  

SciTech Connect

The key regulatory step in the biosynthesis of abscisic acid (ABA), a hormone central to the regulation of several important processes in plants, is the oxidative cleavage of the 11,12 double bond of a 9-cis-epoxycarotenoid. The enzyme viviparous14 (VP14) performs this cleavage in maize (Zea mays), making it a target for the rational design of novel chemical agents and genetic modifications that improve plant behavior through the modulation of ABA levels. The structure of VP14, determined to 3.2-{angstrom} resolution, provides both insight into the determinants of regio- and stereospecificity of this enzyme and suggests a possible mechanism for oxidative cleavage. Furthermore, mutagenesis of the distantly related CCD1 of maize shows how the VP14 structure represents a template for all plant carotenoid cleavage dioxygenases (CCDs). In addition, the structure suggests how VP14 associates with the membrane as a way of gaining access to its membrane soluble substrate.

Messing, Simon A.J.; Gabelli, Sandra B.; Echeverria, Ignacia; Vogel, Jonathan T.; Guan, Jiahn Chou; Tan, Bao Cai; Klee, Harry J.; McCarty, Donald R.; Amzel, L. Mario (JHU); (Florida)

2011-09-06

311

Structural Insights into Maize Viviparous14, a Key Enzyme in the Biosynthesis of the Phytohormone Abscisic Acid W  

SciTech Connect

The key regulatory step in the biosynthesis of abscisic acid (ABA), a hormone central to the regulation of several important processes in plants, is the oxidative cleavage of the 11,12 double bond of a 9-cis-epoxycarotenoid. The enzyme viviparous14 (VP14) performs this cleavage in maize (Zea mays), making it a target for the rational design of novel chemical agents and genetic modifications that improve plant behavior through the modulation of ABA levels. The structure of VP14, determined to 3.2-{angstrom} resolution, provides both insight into the determinants of regio- and stereospecificity of this enzyme and suggests a possible mechanism for oxidative cleavage. Furthermore, mutagenesis of the distantly related CCD1 of maize shows how the VP14 structure represents a template for all plant carotenoid cleavage dioxygenases (CCDs). In addition, the structure suggests how VP14 associates with the membrane as a way of gaining access to its membrane soluble substrate.

Messing, S.; Gabelli, S; Echeverria, I; Vogel, J; Guan, J; Tan, B; Klee, H; McCarty, D; Amzela, M

2010-01-01

312

Histopathological findings of the pancreas, liver, and carbohydrate metabolizing enzymes in STZ-induced diabetic rats improved by administration of myrtenal.  

PubMed

This study aims to evaluate the efficacy of myrtenal, a natural monoterpene, for its antihyperglycemic effects and ? cell protective properties in streptozotocin (STZ)-induced diabetic rats. Oral administration of myrtenal at doses of 20, 40, and 80 mg/kg body weight to diabetic rats for 28 days resulted in a significant reduction (P < 0.05) in the levels of plasma glucose, glycosylated hemoglobin (HbA1c), and an increase in the levels of insulin and hemoglobin (Hb). Protection of body weight loss of diabetic rats by myrtenal was noted. The altered activities of the key metabolic enzymes involved in carbohydrate metabolism such as hexokinase, glucose-6-phosphatase, fructose-1,6-bisphosphatase, glucose-6-phosphate dehydrogenase, and hepatic enzymes AST, ALT, and ALP levels of diabetic rats were significantly improved by the administration of myrtenal in STZ-induced diabetic rats. Moreover, myrtenal treatment improved hepatic and muscle glycogen content in diabetic rats. Histopathological studies further revealed that the reduced islet cells were restored to near-normal conditions on treatment with myrtenal in STZ-induced diabetic rats. An alteration in liver architecture was also prevented by myrtenal treatment. Our results suggest that myrtenal possess antihyperglycemic and ? cell protective effects. Hence, myrtenal could be considered as a potent phytochemical for development as a new antidiabetic agent. PMID:25292424

Rathinam, Ayyasamy; Pari, Leelavinothan; Chandramohan, Ramasamy; Sheikh, Bashir Ahmad

2014-12-01

313

(Pyruvate decarboxylase: A key enzyme for alcohol production): (Annual) performance report  

SciTech Connect

The overall objectives of this grant are to investigate the properties and regulation of pyruvate decarboxylase from Z. mobilis and to identify the unique features which allow its high level of expression. Steps in the project include enzyme purification and antibody production, characterization of physical and kinetic properties, investigations of how environmental conditions affect the level of pyruvate decarboxylase, it's biosynthesis and turnover, cloning and sequencing, investigations of the effects of increased gene dosage, control of m-RNA synthesis and turnover, and promoter structure. 11 refs.

Ingram, L.O.

1989-01-01

314

Mono-hydroxy methoxychlor alters levels of key sex steroids and steroidogenic enzymes in cultured mouse antral follicles  

SciTech Connect

Methoxychlor (MXC) is an organochlorine pesticide that reduces fertility in female rodents by decreasing antral follicle numbers and increasing follicular death. MXC is metabolized in the body to mono-hydroxy MXC (mono-OH). Little is known about the effects of mono-OH on the ovary. Thus, this work tested the hypothesis that mono-OH exposure decreases production of 17{beta}-estradiol (E{sub 2}) by cultured mouse antral follicles. Antral follicles were isolated from CD-1 mice (age 35-39 days) and exposed to dimethylsulfoxide (DMSO), or mono-OH (0.1-10 {mu}g/mL) for 96 h. Media and follicles were collected for analysis of sex steroid levels and mRNA expression, respectively. Mono-OH treatment (10 {mu}g/mL) decreased E{sub 2} (DMSO: 3009.72 {+-} 744.99 ng/mL; mono-OH 0.1 {mu}g/mL: 1679.66 {+-} 461.99 ng/mL; 1 {mu}g/mL: 1752.72 {+-} 532.41 ng/mL; 10 {mu}g/mL: 45.89 {+-} 33.83 ng/mL), testosterone (DMSO: 15.43 {+-} 2.86 ng/mL; mono-OH 0.1 {mu}g/mL: 17.17 {+-} 4.71 ng/mL; 1 {mu}g/mL: 13.64 {+-} 3.53 ng/mL; 10 {mu}g/mL: 1.29 {+-} 0.23 ng/mL), androstenedione (DMSO: 1.92 {+-} 0.34 ng/mL; mono-OH 0.1 {mu}g/mL: 1.49 {+-} 0.43 ng/mL; 1 {mu}g/mL: 0.64 {+-} 0.31 ng/mL; 10 {mu}g/mL: 0.12 {+-} 0.06 ng/mL) and progesterone (DMSO: 24.11 {+-} 4.21 ng/mL; mono-OH 0.1 {mu}g/mL: 26.77 {+-} 4.41 ng/mL; 1 {mu}g/mL: 20.90 {+-} 3.75 ng/mL; 10 {mu}g/mL: 9.44 {+-} 2.97 ng/mL) levels. Mono-OH did not alter expression of Star, Hsd3b1, Hsd17b1 and Cyp1b1, but it did reduce levels of Cyp11a1, Cyp17a1 and Cyp19a1 mRNA. Collectively, these data suggest that mono-OH significantly decreases levels of key sex steroid hormones and the expression of enzymes required for steroidogenesis.

Craig, Zelieann R., E-mail: zelieann@gmail.co [Department of Comparative Biosciences, University of Illinois, Urbana, IL (United States); Leslie, Traci C., E-mail: traci.leslie@gmail.co [Department of Comparative Biosciences, University of Illinois, Urbana, IL (United States); Hatfield, Kimberly P., E-mail: kpm9786@yahoo.co [Program in Toxicology and Department of Epidemiology and Preventive Medicine, University of Maryland School of Medicine, Baltimore, MD 21201 (United States); Gupta, Rupesh K., E-mail: drrupesh@illinois.ed [Department of Comparative Biosciences, University of Illinois, Urbana, IL (United States); Flaws, Jodi A., E-mail: jflaws@illinois.ed [Department of Comparative Biosciences, University of Illinois, Urbana, IL (United States)

2010-12-01

315

A pangenomic analysis of the Nannochloropsis organellar genomes reveals novel genetic variations in key metabolic genes  

PubMed Central

Background Microalgae in the genus Nannochloropsis are photosynthetic marine Eustigmatophytes of significant interest to the bioenergy and aquaculture sectors due to their ability to efficiently accumulate biomass and lipids for utilization in renewable transportation fuels, aquaculture feed, and other useful bioproducts. To better understand the genetic complement that drives the metabolic processes of these organisms, we present the assembly and comparative pangenomic analysis of the chloroplast and mitochondrial genomes from Nannochloropsis salina CCMP1776. Results The chloroplast and mitochondrial genomes of N. salina are 98.4% and 97% identical to their counterparts in Nannochloropsis gaditana. Comparison of the Nannochloropsis pangenome to other algae within and outside of the same phyla revealed regions of significant genetic divergence in key genes that encode proteins needed for regulation of branched chain amino synthesis (acetohydroxyacid synthase), carbon fixation (RuBisCO activase), energy conservation (ATP synthase), protein synthesis and homeostasis (Clp protease, ribosome). Conclusions Many organellar gene modifications in Nannochloropsis are unique and deviate from conserved orthologs found across the tree of life. Implementation of secondary and tertiary structure prediction was crucial to functionally characterize many proteins and therefore should be implemented in automated annotation pipelines. The exceptional similarity of the N. salina and N. gaditana organellar genomes suggests that N. gaditana be reclassified as a strain of N. salina. PMID:24646409

2014-01-01

316

Proline and its metabolism enzymes in cucumber cell cultures during acclimation to salinity.  

PubMed

Proline is an important osmolyte appearing as the result of salt stress response of plants. In the present study, we measured the proline concentration, activities of pyrroline-5-carboxylate synthetase (P5CS), pyrroline-5-carboxylate reductase (P5CR), and proline dehydrogenase (PDH) key regulatory enzymes in the biosynthesis and degradation of proline in the acclimated (AC20) and the non-acclimated (NAC) cucumber cell suspension cultures subjected to moderate (150 mM NaCl; AC20-150, NAC-150, respectively) and severe (200 mM NaCl; AC20-200, NAC-200, respectively) salt stress. The data showed that salt stress brought about a linear increase in proline content in both types of cultures. However, in the acclimated culture proline accumulation was observed earlier, in third hour after stress. Only in the acclimated culture moderate and severe stresses up-regulated P5CS activity throughout the experiment, whereas the activity of P5CR grew in response to both NaCl concentrations only in 24th and 48th hour. The severe salt stress resulted in decrease in P5CR in NAC-200 cultures. In response to salt stress, both types of cell suspension cultures reacted with decline in PDH activity below the spectrophotometrically detected level. Cell cultures vigor correlated with salt concentration and time of exposure to the stress factor. Both NaCl concentrations caused linear decline in vigor of the non-acclimated culture up to 80-90 % at the end of the experiment, whereas in the acclimated culture significant decrease by about 30-40 % was reached in 24th hour after stress. The presented data suggest that acclimation to salt stress up-regulated proline synthesis enzyme activity and caused intensive accumulations of proline by inhibiting its oxidation. PMID:23990108

Naliwajski, Marcin R; Sk?odowska, Maria

2014-01-01

317

The histone demethylase enzyme KDM3A is a key estrogen receptor regulator in breast cancer  

PubMed Central

Endocrine therapy has successfully been used to treat estrogen receptor (ER)-positive breast cancer, but this invariably fails with cancers becoming refractory to treatment. Emerging evidence has suggested that fluctuations in ER co-regulatory protein expression may facilitate resistance to therapy and be involved in breast cancer progression. To date, a small number of enzymes that control methylation status of histones have been identified as co-regulators of ER signalling. We have identified the histone H3 lysine 9 mono- and di-methyl demethylase enzyme KDM3A as a positive regulator of ER activity. Here, we demonstrate that depletion of KDM3A by RNAi abrogates the recruitment of the ER to cis-regulatory elements within target gene promoters, thereby inhibiting estrogen-induced gene expression changes. Global gene expression analysis of KDM3A-depleted cells identified gene clusters associated with cell growth. Consistent with this, we show that knockdown of KDM3A reduces ER-positive cell proliferation and demonstrate that KDM3A is required for growth in a model of endocrine therapy-resistant disease. Crucially, we show that KDM3A catalytic activity is required for both ER-target gene expression and cell growth, demonstrating that developing compounds which target demethylase enzymatic activity may be efficacious in treating both ER-positive and endocrine therapy-resistant disease. PMID:25488809

Wade, Mark A.; Jones, Dominic; Wilson, Laura; Stockley, Jacqueline; Coffey, Kelly; Robson, Craig N.; Gaughan, Luke

2015-01-01

318

Cellulase a key enzyme in fermentation: Annual report, 1985--1986  

SciTech Connect

Many microbial cellulase systems are comprised of multiple components namely, endoglucanase (EG), cellobiohydrolase (CBH), and betaglucosidabe. These three types of enzyme of the cellulase complex act synergistically but their mechanism of interaction in the hydrolysis of crystalline cellulose is poorly understood. As monoclonal antibodies (McAb) can be used as both snesitive and specific protein probes, we proposed the preparation of McAb specific for cellulases components, which should allow direct analysis of these enzymes. We have been generating murine McAb that react specifically with cellobiohydrolases and endoglucanases. To date a McAb specific for a fungal (Trichoderma reesei) cellobiohydrolase I (CBH I) that does not show cross reactivity toward endoglucanases, has been obtained (Riskeet et al; 1986a). This McAb preparation has been used for purification of CBH I via affinity chromatography. Thus the purified McAb to CBH I was immobilized on CnBr-Sepharose-4b to yield an immunomatrix which was analogous studies are also being developed with the cellulase form Microbispora bispora. 5 refs.

Eveleigh, D.E.; Macmillan, J.D.

1986-05-04

319

Potato steroidal glycoalkaloid levels and the expression of key isoprenoid metabolic genes.  

PubMed

The potato steroidal glycoalkaloids (SGA) are toxic secondary metabolites, and their total content in tubers should not exceed 20 mg/100 g fresh weight. The two major SGA in cultivated potato (Solanum tuberosum) are alpha-chaconine and alpha-solanine. SGA biosynthetic genes and the genetic factors that control their expression have not yet been determined. In the present study, potato genotypes exhibiting different levels of SGA content showed an association between high SGA levels in their leaves and tubers and high expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase 1 (hmg1) and squalene synthase 1 (pss1), genes of the mevalonic/isoprenoid pathway. Transcripts of other key enzymes of branches of the isoprenoid pathway, vetispiradiene/sesquiterpene synthase (pvs1) and sterol C24-methyltransferase type1 (smt1), were undetectable or exhibited stable expression regardless of SGA content, respectively, suggesting facilitated precursor flow to the SGA biosynthetic branch. The transcript ratio of solanidine glucosyltransferase (sgt2) to solanidine galactosyltransferase (sgt1) was correlated to the documented chaconine-to-solanine ratio in the tested genotypes. Significantly higher expression of hmg1, pss1, smt1, sgt1 and sgt2 was monitored in the tuber phelloderm than in the parenchyma of the tuber's flesh, targeting the former as the main SGA-producing tissue in the tuber, in agreement with the known high SGA content in the layers directly under the tuber skin. PMID:17701426

Krits, Pinchas; Fogelman, Edna; Ginzberg, Idit

2007-12-01

320

Hypoxic training increases metabolic enzyme activity and composition of ?-myosin heavy chain isoform in rat ventricular myocardium  

Microsoft Academic Search

Cardiac muscle adaptation is essential for maintaining physical capacity after ascending to high altitude. This study examines\\u000a the effects of high altitude training on myocardial metabolic enzyme activity and composition of ?-myosin heavy chain (MHC).\\u000a Rats were randomly divided into normobaric sedentary (NS) and training (NT) groups, and hypobaric sedentary (HS) and training\\u000a (HT) groups. HS and HT rats were

Ming-Chun Cai; Qing-Yuan Huang; Wei-Gong Liao; Zhou Wu; Fu-Yu Liu; Yu-Qi Gao

2010-01-01

321

Enzymes of glycerol and glyceraldehyde metabolism in mouse liver: effects of caloric restriction and age on activities  

Microsoft Academic Search

Synopsis The influence of caloric restriction on hepatic glyceraldehyde- and glycerol-metabolizing enzyme activities of young and old mice were studied. Glycerol kinase and cytoplasmic glycerol-3-phosphate dehydrogenase activities were increa- sed in both young and old CR (calorie-restricted) mice when compared with controls, whereas triokinase increased only in old CR mice. Aldehyde dehydrogenase and aldehyde reductase activities in both young and

Kevork Hagopian; Jon J. RAMSEY; Richard Weindruch

2008-01-01

322

Associations between genetic polymorphisms of Phase I and II metabolizing enzymes, p53 and susceptibility to esophageal adenocarcinoma  

Microsoft Academic Search

The objectives of this exploratory case–control study were to evaluate whether genetic polymorphisms of selected Phase I and II metabolizing enzymes are associated with the risk of developing primary esophageal adenocarcinoma, and to investigate potential associations between genotypes and p53 tumor suppressor gene alterations. Cases comprised 45 patients with surgically resected esophageal adenocarcinomas, defined according to strict clinico-pathologic criteria. PCR-based

A. G Casson; Z Zheng; D Chiasson; K MacDonald; D. C Riddell; J. R Guernsey; D. L Guernsey; J McLaughlin

2003-01-01

323

Metabolic mechanisms of methanol\\/formaldehyde in isolated rat hepatocytes: Carbonyl-metabolizing enzymes versus oxidative stress  

Microsoft Academic Search

Methanol (CH3OH), a common industrial solvent, is metabolized to toxic compounds by several enzymatic as well as free radical pathways. Identifying which process best enhances or prevents CH3OH-induced cytotoxicity could provide insight into the molecular basis for acute CH3OH-induced hepatoxicity. Metabolic pathways studied include those found in 1) an isolated hepatocyte system and 2) cell-free systems. Accelerated Cytotoxicity Mechanism Screening

Stephanie L. MacAllister; Joanna Choi; Liana Dedina; Peter J. O’Brien

2011-01-01

324

Remarkable Reproducibility of Enzyme Activity Profiles in Tomato Fruits Grown under Contrasting Environments Provides a Roadmap for Studies of Fruit Metabolism1[W][OPEN  

PubMed Central

To assess the influence of the environment on fruit metabolism, tomato (Solanum lycopersicum ‘Moneymaker’) plants were grown under contrasting conditions (optimal for commercial, water limited, or shaded production) and locations. Samples were harvested at nine stages of development, and 36 enzyme activities of central metabolism were measured as well as protein, starch, and major metabolites, such as hexoses, sucrose, organic acids, and amino acids. The most remarkable result was the high reproducibility of enzyme activities throughout development, irrespective of conditions or location. Hierarchical clustering of enzyme activities also revealed tight relationships between metabolic pathways and phases of development. Thus, cell division was characterized by high activities of fructokinase, glucokinase, pyruvate kinase, and tricarboxylic acid cycle enzymes, indicating ATP production as a priority, whereas cell expansion was characterized by enzymes involved in the lower part of glycolysis, suggesting a metabolic reprogramming to anaplerosis. As expected, enzymes involved in the accumulation of sugars, citrate, and glutamate were strongly increased during ripening. However, a group of enzymes involved in ATP production, which is probably fueled by starch degradation, was also increased. Metabolites levels seemed more sensitive than enzymes to the environment, although such differences tended to decrease at ripening. The integration of enzyme and metabolite data obtained under contrasting growth conditions using principal component analysis suggests that, with the exceptions of alanine amino transferase and glutamate and malate dehydrogenase and malate, there are no links between single enzyme activities and metabolite time courses or levels. PMID:24474652

Biais, Benoît; Bénard, Camille; Beauvoit, Bertrand; Colombié, Sophie; Prodhomme, Duyęn; Ménard, Guillaume; Bernillon, Stéphane; Gehl, Bernadette; Gautier, Hélčne; Ballias, Patricia; Mazat, Jean-Pierre; Sweetlove, Lee; Génard, Michel; Gibon, Yves

2014-01-01

325

Enhanced enzymatic reactivity for electrochemically driven drug metabolism by confining cytochrome P450 enzyme in TiO? nanotube arrays.  

PubMed

Understanding the enzymatic reaction kinetics that occur within a confined space or interface is a significant challenge. Herein, a nanotube array enzymatic reactor (CYP2C9/Au/TNA) was constructed by electrostatically adsorbing enzyme on the inner wall of TiO2 nanotube arrays (TNAs). TNAs with different dimensions could be fabricated by the anodization of titanium foil through varying the anodization potential or time. The electrical conductivity of TNAs was improved by electrodepositing Au nanoparticles on the inner wall of TNAs. The cytochrome P450 2C9 enzyme (CYP2C9) was confined inside TNAs as a model. The enzymatic activity of CYP2C9 and tolbutamide metabolic yield could be effectively regulated by changing the nanotube diameter and length of TNAs. The enzymatic rate constant k(cat) and apparent Michaelis constant K(m)(app) were determined to be 9.89 s(-1) and 4.8 ?M at the tube inner diameter of about 64 nm and length of 1.08 ?m. The highest metabolic yield of tolbutamide reached 14.6%. Furthermore, the designed nanotube array enzymatic reactor could be also used in situ to monitor the tolbutamide concentration with sensitivity of 28.8 ?A mM(-1) and detection limit of 0.52 ?M. Therefore, the proposed nanotube array enzymatic reactor was a good vessel for studying enzyme biocatalysis and drug metabolism, and has potential applications including efficient biosensors and bioreactors for chemical synthesis. PMID:25014006

Lu, Jusheng; Li, Henan; Cui, Dongmei; Zhang, Yuanjian; Liu, Songqin

2014-08-01

326

Toxicology in the Fast Lane: Application of High-Throughput Bioassays to Detect Modulation of Key Enzymes and Receptors  

PubMed Central

Background Legislation at state, federal, and international levels is requiring rapid evaluation of the toxicity of numerous chemicals. Whole-animal toxicologic studies cannot yield the necessary throughput in a cost-effective fashion, leading to a critical need for a faster and more cost-effective toxicologic evaluation of xenobiotics. Objectives We tested whether mechanistically based screening assays can rapidly provide information on the potential for compounds to affect key enzymes and receptor targets, thus identifying those compounds requiring further in-depth analysis. Methods A library of 176 synthetic chemicals was prepared and examined in a high-throughput screening (HTS) manner using nine enzyme-based and five receptor-based bioassays. Results All the assays have high Z? values, indicating good discrimination among compounds in a reliable fashion, and thus are suitable for HTS assays. On average, three positive hits were obtained per assay. Although we identified compounds that were previously shown to inhibit a particular enzyme class or receptor, we surprisingly discovered that triclosan, a microbiocide present in personal care products, inhibits carboxylesterases and that dichlone, a fungicide, strongly inhibits the ryanodine receptors. Conclusions Considering the need to rapidly screen tens of thousands of anthropogenic compounds, our study shows the feasibility of using combined HTS assays as a novel approach toward obtaining toxicologic data on numerous biological end points. The HTS assay approach is very useful to quickly identify potentially hazardous compounds and to prioritize them for further in-depth studies. PMID:20049205

Morisseau, Christophe; Merzlikin, Oleg; Lin, Amy; He, Guochun; Feng, Wei; Padilla, Isela; Denison, Michael S.; Pessah, Isaac N.; Hammock, Bruce D.

2009-01-01

327

Yucasin is a potent inhibitor of YUCCA, a key enzyme in auxin biosynthesis.  

PubMed

Indole-3-acetic acid (IAA), an auxin plant hormone, is biosynthesized from tryptophan. The indole-3-pyruvic acid (IPyA) pathway, involving the tryptophan aminotransferase TAA1 and YUCCA (YUC) enzymes, was recently found to be a major IAA biosynthetic pathway in Arabidopsis. TAA1 catalyzes the conversion of tryptophan to IPyA, and YUC produces IAA from IPyA. Using a chemical biology approach with maize coleoptiles, we identified 5-(4-chlorophenyl)-4H-1,2,4-triazole-3-thiol (yucasin) as a potent inhibitor of IAA biosynthesis in YUC-expressing coleoptile tips. Enzymatic analysis of recombinant AtYUC1-His suggested that yucasin strongly inhibited YUC1-His activity against the substrate IPyA in a competitive manner. Phenotypic analysis of Arabidopsis YUC1 over-expression lines (35S::YUC1) demonstrated that yucasin acts in IAA biosynthesis catalyzed by YUC. In addition, 35S::YUC1 seedlings showed resistance to yucasin in terms of root growth. A loss-of-function mutant of TAA1, sav3-2, was hypersensitive to yucasin in terms of root growth and hypocotyl elongation of etiolated seedlings. Yucasin combined with the TAA1 inhibitor l-kynurenine acted additively in Arabidopsis seedlings, producing a phenotype similar to yucasin-treated sav3-2 seedlings, indicating the importance of IAA biosynthesis via the IPyA pathway in root growth and leaf vascular development. The present study showed that yucasin is a potent inhibitor of YUC enzymes that offers an effective tool for analyzing the contribution of IAA biosynthesis via the IPyA pathway to plant development and physiological processes. PMID:24299123

Nishimura, Takeshi; Hayashi, Ken-Ichiro; Suzuki, Hiromi; Gyohda, Atsuko; Takaoka, Chihiro; Sakaguchi, Yusuke; Matsumoto, Sachiko; Kasahara, Hiroyuki; Sakai, Tatsuya; Kato, Jun-Ichi; Kamiya, Yuji; Koshiba, Tomokazu

2014-02-01

328

Effects of Sublethal Exposure to a Glyphosate-Based Herbicide Formulation on Metabolic Activities of Different Xenobiotic-Metabolizing Enzymes in Rats.  

PubMed

The activities of different xenobiotic-metabolizing enzymes in liver subcellular fractions from Wistar rats exposed to a glyphosate (GLP)-based herbicide (Roundup full II) were evaluated in this work. Exposure to the herbicide triggered protective mechanisms against oxidative stress (increased glutathione peroxidase activity and total glutathione levels). Liver microsomes from both male and female rats exposed to the herbicide had lower (45%-54%, P < 0.01) hepatic cytochrome P450 (CYP) levels compared to their respective control animals. In female rats, the hepatic 7-ethoxycoumarin O-deethylase (a general CYP-dependent enzyme activity) was 57% higher (P < 0.05) in herbicide-exposed compared to control animals. Conversely, this enzyme activity was 58% lower (P < 0.05) in male rats receiving the herbicide. Lower (P < 0.05) 7-ethoxyresorufin O-deethlyase (EROD, CYP1A1/2 dependent) and oleandomycin triacetate (TAO) N-demethylase (CYP3A dependent) enzyme activities were observed in liver microsomes from exposed male rats. Conversely, in females receiving the herbicide, EROD increased (123%-168%, P < 0.05), whereas TAO N-demethylase did not change. A higher (158%-179%, P < 0.01) benzyloxyresorufin O-debenzylase (a CYP2B-dependent enzyme activity) activity was only observed in herbicide-exposed female rats. In herbicide-exposed rats, the hepatic S-oxidation of methimazole (flavin monooxygenase dependent) was 49% to 62% lower (P < 0.001), whereas the carbonyl reduction of menadione (a cytosolic carbonyl reductase-dependent activity) was higher (P < 0.05). Exposure to the herbicide had no effects on enzymatic activities dependent on carboxylesterases, glutathione transferases, and uridinediphospho-glucuronosyltransferases. This research demonstrated certain biochemical modifications after exposure to a GLP-based herbicide. Such modifications may affect the metabolic fate of different endobiotic and xenobiotic substances. The pharmacotoxicological significance of these findings remains to be clarified. PMID:24985121

Larsen, Karen; Najle, Roberto; Lifschitz, Adrián; Maté, María L; Lanusse, Carlos; Virkel, Guillermo L

2014-07-01

329

In vitro studies of Gynura divaricata (L.) DC extracts as inhibitors of key enzymes relevant for type 2 diabetes and hypertension  

Microsoft Academic Search

Aim of the studyTo evaluate traditionally used herb, Gynura divaricata (L.) DC (Bai Bei San Qi) as in vitro inhibitors of key enzymes involved in the pathogenesis of hyperglycemia and hypertension. We also determined the distribution of enzyme inhibitory activities in different aqueous and non-aqueous extracts.

Tingting Wu; Xueting Zhou; Yafei Deng; Qing Jing; Min Li; Lujaing Yuan

2011-01-01

330

Transcription factor Nrf1 negatively regulates the cystine/glutamate transporter and lipid-metabolizing enzymes.  

PubMed

Liver-specific Nrf1 (NF-E2-p45-related factor 1) knockout mice develop nonalcoholic steatohepatitis. To identify postnatal mechanisms responsible for this phenotype, we generated an inducible liver-specific Nrf1 knockout mouse line using animals harboring an Nrf1(flox) allele and a rat CYP1A1-Cre transgene (Nrf1(flox/flox)::CYP1A1-Cre mice). Administration of 3-methylcholanthrene (3-MC) to these mice (Nrf1(flox/flox)::CYP1A1-Cre+3MC mice) resulted in loss of hepatic Nrf1 expression. The livers of mice lacking Nrf1 accumulated lipid, and the hepatic fatty acid (FA) composition in such animals differed significantly from that in the Nrf1(flox/flox)::CYP1A1-Cre control. This change was provoked by upregulation of several FA metabolism genes. Unexpectedly, we also found that the level of glutathione was increased dramatically in livers of Nrf1(flox/flox)::CYP1A1-Cre+3MC mice. While expression of glutathione biosynthetic enzymes was unchanged, xCT, a component of the cystine/glutamate antiporter system x(c)(-), was significantly upregulated in livers of Nrf1(flox/flox)::CYP1A1-Cre+3MC mice, suggesting that Nrf1 normally suppresses xCT. Thus, stress-inducible expression of xCT is a two-step process: under homeostatic conditions, Nrf1 effectively suppresses nonspecific transactivation of xCT, but when cells encounter severe oxidative/electrophilic stress, Nrf1 is displaced from an antioxidant response element (ARE) in the gene promoter while Nrf2 is recruited to the ARE. Thus, Nrf1 controls both the FA and the cystine/cysteine content of hepatocytes by participating in an elaborate regulatory network. PMID:25092871

Tsujita, Tadayuki; Peirce, Vivian; Baird, Liam; Matsuyama, Yuka; Takaku, Misaki; Walsh, Shawn V; Griffin, Julian L; Uruno, Akira; Yamamoto, Masayuki; Hayes, John D

2014-10-01

331

Transcription Factor Nrf1 Negatively Regulates the Cystine/Glutamate Transporter and Lipid-Metabolizing Enzymes  

PubMed Central

Liver-specific Nrf1 (NF-E2-p45-related factor 1) knockout mice develop nonalcoholic steatohepatitis. To identify postnatal mechanisms responsible for this phenotype, we generated an inducible liver-specific Nrf1 knockout mouse line using animals harboring an Nrf1flox allele and a rat CYP1A1-Cre transgene (Nrf1flox/flox::CYP1A1-Cre mice). Administration of 3-methylcholanthrene (3-MC) to these mice (Nrf1flox/flox::CYP1A1-Cre+3MC mice) resulted in loss of hepatic Nrf1 expression. The livers of mice lacking Nrf1 accumulated lipid, and the hepatic fatty acid (FA) composition in such animals differed significantly from that in the Nrf1flox/flox::CYP1A1-Cre control. This change was provoked by upregulation of several FA metabolism genes. Unexpectedly, we also found that the level of glutathione was increased dramatically in livers of Nrf1flox/flox::CYP1A1-Cre+3MC mice. While expression of glutathione biosynthetic enzymes was unchanged, xCT, a component of the cystine/glutamate antiporter system xc?, was significantly upregulated in livers of Nrf1flox/flox::CYP1A1-Cre+3MC mice, suggesting that Nrf1 normally suppresses xCT. Thus, stress-inducible expression of xCT is a two-step process: under homeostatic conditions, Nrf1 effectively suppresses nonspecific transactivation of xCT, but when cells encounter severe oxidative/electrophilic stress, Nrf1 is displaced from an antioxidant response element (ARE) in the gene promoter while Nrf2 is recruited to the ARE. Thus, Nrf1 controls both the FA and the cystine/cysteine content of hepatocytes by participating in an elaborate regulatory network. PMID:25092871

Tsujita, Tadayuki; Peirce, Vivian; Baird, Liam; Matsuyama, Yuka; Takaku, Misaki; Walsh, Shawn V.; Griffin, Julian L.; Uruno, Akira

2014-01-01

332

Effects of tin-protoporphyrin administration on hepatic xenobiotic metabolizing enzymes in the juvenile rat  

SciTech Connect

The heme analogue tin-protoporphyrin IX (SnP) is a potent inhibitor of microsomal heme oxygenase. Administration of SnP to neonatal rats can prevent hyperbilirubinemia by blocking the postnatal increase of heme oxygenase activity. Apparently innocuous at therapeutic doses, it is of potential clinical value for chemoprevention of neonatal jaundice. We found that when 50-g male Sprague-Dawley rats were treated daily with 50 mumol of SnP/kg sc for 6 days, hepatic microsomal cytochromes b5 and P-450 were significantly diminished. Cytochrome P-450 reductase, two P-450-dependent monooxygenases, aminopyrine demethylase and benzo(a)pyrene hydroxylase, and catalase, a peroxisomal hemoprotein, were also significantly diminished. These results suggested that SnP might significantly affect the metabolism of other xenobiotics. This possibility was confirmed by the finding that hexobarbital-induced sleep lasted 4 times longer in SnP-treated rats than in controls. Inhibition of protein synthesis by SnP was ruled out as the cause of hemoprotein loss when administration of (/sup 3/H)leucine to SnP-treated and control rats demonstrated that proteins of the microsomal, cytosolic, and plasma membrane fractions of the livers from both groups incorporated similar levels of leucine. When /sup 55/FeCl/sub 3/ and (2-/sup 14/C)glycine were administered to measure heme synthesis, heme extract from the livers of SnP-treated rats contained 4 times more label from iron and glycine than did heme from control livers. Despite the apparent increased rate of heme synthesis in SnP-treated rats, each of the three cell fractions demonstrated a significant loss of heme but contained sizable amounts of SnP. These findings suggest that SnP causes a decrease of functional hemoprotein and partial loss of enzymic activity by displacing intracellular heme.

Stout, D.L.; Becker, F.F.

1988-01-01

333

Anaerobic Metabolism of Catechol by the Denitrifying Bacterium Thauera aromatica—a Result of Promiscuous Enzymes and Regulators??  

PubMed Central

The anaerobic metabolism of catechol (1,2-dihydroxybenzene) was studied in the betaproteobacterium Thauera aromatica that was grown with CO2 as a cosubstrate and nitrate as an electron acceptor. Based on different lines of evidence and on our knowledge of enzymes and genes involved in the anaerobic metabolism of other aromatic substrates, the following pathway is proposed. Catechol is converted to catechylphosphate by phenylphosphate synthase, which is followed by carboxylation by phenylphosphate carboxylase at the para position to the phosphorylated phenolic hydroxyl group. The product, protocatechuate (3,4-dihydroxybenzoate), is converted to its coenzyme A (CoA) thioester by 3-hydroxybenzoate-CoA ligase. Protocatechuyl-CoA is reductively dehydroxylated to 3-hydroxybenzoyl-CoA, possibly by 4-hydroxybenzoyl-CoA reductase. 3-Hydroxybenzoyl-CoA is further metabolized by reduction of the aromatic ring catalyzed by an ATP-driven benzoyl-CoA reductase. Hence, the promiscuity of several enzymes and regulatory proteins may be sufficient to create the catechol pathway that is made up of elements of phenol, 3-hydroxybenzoate, 4-hydroxybenzoate, and benzoate metabolism. PMID:18156265

Ding, Bin; Schmeling, Sirko; Fuchs, Georg

2008-01-01

334

In vitro metabolism of ?7 neuronal nicotinic receptor agonist AZD0328 and enzyme identification for its N-oxide metabolite.  

PubMed

1. AZD0328 was pharmacologically characterized as a ?7 neuronal nicotinic receptor agonist intended for treatment of Alzheimer's disease. In vitro AZD0328 cross species metabolite profile and enzyme identification for its N-oxide metabolite were evaluated in this study. 2. AZD0328 was very stable in the human hepatocyte incubation, whereas extensively metabolized in rat, dog and guinea pig hepatocyte incubations. The N-oxidation metabolite (M6) was the only metabolite detected in human hepatocyte incubations, and it also appeared to be the major in vitro metabolic pathway in a number of preclinical species. In addition, N-glucuronide metabolite of AZD0328 was observed in human liver microsomes. 3. Other metabolic pathways in the preclinical species include hydroxylation in azabicyclo octane or furopyridine part of the molecule. Pyridine N-methylation of AZD0328 (M2) was identified as a dog specific metabolite, not observed in human or other preclinical species. 4. Multiple enzymes including CYP2D6, CYP3A4/5, FMO1 and FMO3 catalyzed AZD0328 metabolism. The potential for AZD0328 to be inhibited clinically by co-administered drugs or genetic polymorphism is relative low. PMID:21226652

Zhou, Diansong; Zhang, Minli; Ye, Xiaomei; Gu, Chungang; Piser, Timothy M; Lanoue, Bernard A; Schock, Sara A; Cheng, Yi-Fang; Grimm, Scott W

2011-03-01

335

Mitochondrial dysfunction in obesity: potential benefit and mechanism of Co-enzyme Q10 supplementation in metabolic syndrome  

PubMed Central

Co-enzyme Q10 (Co-Q10) is an essential component of the mitochondrial electron transport chain. Most cells are sensitive to co-enzyme Q10 (Co-Q10) deficiency. This deficiency has been implicated in several clinical disorders such as heart failure, hypertension, Parkinson’s disease and obesity. The lipid lowering drug statin inhibits conversion of HMG-CoA to mevalonate and lowers plasma Co-Q10 concentrations. However, supplementation with Co-Q10 improves the pathophysiological condition of statin therapy. Recent evidence suggests that Co-Q10 supplementation may be useful for the treatment of obesity, oxidative stress and the inflammatory process in metabolic syndrome. The anti-inflammatory response and lipid metabolizing effect of Co-Q10 is probably mediated by transcriptional regulation of inflammation and lipid metabolism. This paper reviews the evidence showing beneficial role of Co-Q10 supplementation and its potential mechanism of action on contributing factors of metabolic and cardiovascular complications. PMID:24932457

2014-01-01

336

Enzymological analysis of the tumor suppressor A-C1 reveals a novel group of phospholipid-metabolizing enzymes  

PubMed Central

A-C1 protein is the product of a tumor suppressor gene negatively regulating the oncogene Ras and belongs to the HRASLS (HRAS-like suppressor) subfamily. We recently found that four members of this subfamily expressed in human tissues function as phospholipid-metabolizing enzymes. Here we examined a possible enzyme activity of A-C1. The homogenates of COS-7 cells overexpressing recombinant A-C1s from human, mouse, and rat showed a phospholipase A1/2 (PLA1/2) activity toward phosphatidylcholine (PC). This finding was confirmed with the purified A-C1. The activity was Ca2+ independent, and dithiothreitol and Nonidet P-40 were indispensable for full activity. Phosphatidylethanolamine (PE) was also a substrate and the phospholipase A1 (PLA1) activity was dominant over the PLA2 activity. Furthermore, the protein exhibited acyltransferase activities transferring an acyl group of PCs to the amino group of PEs and the hydroxyl group of lyso PCs. As for tissue distribution in human, mouse, and rat, A-C1 mRNA was abundantly expressed in testis, skeletal muscle, brain, and heart. These results demonstrate that A-C1 is a novel phospholipid-metabolizing enzyme. Moreover, the fact that all five members of the HRASLS subfamily, including A-C1, show similar catalytic properties strongly suggests that these proteins constitute a new class of enzymes showing PLA1/2 and acyltransferase activities. PMID:21880860

Shinohara, Naoki; Uyama, Toru; Jin, Xing-Hua; Tsuboi, Kazuhito; Tonai, Takeharu; Houchi, Hitoshi; Ueda, Natsuo

2011-01-01

337

Key elements of plant-based diets associated with reduced risk of metabolic syndrome.  

PubMed

Approximately 20 %-25 % of adults worldwide have metabolic syndrome. Vegetarian and vegan diets have demonstrated effectiveness in improving body weight, glycemic control, and cardiovascular risk factors, as compared with conventional therapeutic approaches, and are potentially useful in the prevention of metabolic syndrome. This article consists of two steps: (1) a review of the literature on studies examining vegetarian and vegan diets and metabolic syndrome and (2) a review of foods and nutrients that are protective against or associated with metabolic syndromes that may help to explain the beneficial effects of plant-based dietary approaches for metabolic syndrome. The present review found eight observational research studies, and no intervention studies, examining the association of plant-based dietary approaches with metabolic syndrome. These studies, conducted mostly in Asian populations, yielded varying results. The majority, however, found better metabolic risk factors and lowered risk of metabolic syndrome among individuals following plant-based diets, as compared with omnivores. Some dietary components that are lower in the diets of vegetarians, such as energy intake, saturated fat, heme iron, and red and processed meat, may influence metabolic syndrome risk. In addition, plant-based diets are higher in fruits, vegetables, and fiber, which are protective against the development of metabolic syndrome. PMID:25084991

Turner-McGrievy, Gabrielle; Harris, Metria

2014-01-01

338

Homology modeling of mosquito cytochrome P450 enzymes involved in pyrethroid metabolism: insights into differences in substrate selectivity  

PubMed Central

Background Cytochrome P450 enzymes (P450s) have been implicated in insecticide resistance. Anopheles minumus mosquito P450 isoforms CYP6AA3 and CYP6P7 are capable of metabolizing pyrethroid insecticides, however CYP6P8 lacks activity against this class of compounds. Findings Homology models of the three An. minimus P450 enzymes were constructed using the multiple template alignment method. The predicted enzyme model structures were compared and used for molecular docking with insecticides and compared with results of in vitro enzymatic assays. The three model structures comprise common P450 folds but differences in geometry of their active-site cavities and substrate access channels are prominent. The CYP6AA3 model has a large active site allowing it to accommodate multiple conformations of pyrethroids. The predicted CYP6P7 active site is more constrained and less accessible to binding of pyrethroids. Moreover the predicted hydrophobic interface in the active-site cavities of CYP6AA3 and CYP6P7 may contribute to their substrate selectivity. The absence of CYP6P8 activity toward pyrethroids appears to be due to its small substrate access channel and the presence of R114 and R216 that may prevent access of pyrethroids to the enzyme heme center. Conclusions Differences in active site topologies among CYPAA3, CYP6P7, and CYP6P8 enzymes may impact substrate binding and selectivity. Information obtained using homology models has the potential to enhance the understanding of pyrethroid metabolism and detoxification mediated by P450 enzymes. PMID:21892968

2011-01-01

339

Zinc Biochemistry: From a Single Zinc Enzyme to a Key Element of Life12  

PubMed Central

The nutritional essentiality of zinc for the growth of living organisms had been recognized long before zinc biochemistry began with the discovery of zinc in carbonic anhydrase in 1939. Painstaking analytical work then demonstrated the presence of zinc as a catalytic and structural cofactor in a few hundred enzymes. In the 1980s, the field again gained momentum with the new principle of “zinc finger” proteins, in which zinc has structural functions in domains that interact with other biomolecules. Advances in structural biology and a rapid increase in the availability of gene/protein databases now made it possible to predict zinc-binding sites from metal-binding motifs detected in sequences. This procedure resulted in the definition of zinc proteomes and the remarkable estimate that the human genome encodes ?3000 zinc proteins. More recent developments focus on the regulatory functions of zinc(II) ions in intra- and intercellular information transfer and have tantalizing implications for yet additional functions of zinc in signal transduction and cellular control. At least three dozen proteins homeostatically control the vesicular storage and subcellular distribution of zinc and the concentrations of zinc(II) ions. Novel principles emerge from quantitative investigations on how strongly zinc interacts with proteins and how it is buffered to control the remarkably low cellular and subcellular concentrations of free zinc(II) ions. It is fair to conclude that the impact of zinc for health and disease will be at least as far-reaching as that of iron. PMID:23319127

Maret, Wolfgang

2013-01-01

340

Identification of Neutral Cholesterol Ester Hydrolase, a Key Enzyme Removing Cholesterol from Macrophages*S?  

PubMed Central

Unstable lipid-rich plaques in atherosclerosis are characterized by the accumulation of macrophage foam cells loaded with cholesterol ester (CE). Although hormone-sensitive lipase and cholesteryl ester hydrolase (CEH) have been proposed to mediate the hydrolysis of CE in macrophages, circumstantial evidence suggests the presence of other enzymes with neutral cholesterol ester hydrolase (nCEH) activity. Here we show that the murine orthologue of KIAA1363, designated as neutral cholesterol ester hydrolase (NCEH), is a microsomal nCEH with high expression in murine and human macrophages. The effect of various concentrations of NaCl on its nCEH activity resembles that on endogenous nCEH activity of macrophages. RNA silencing of NCEH decreases nCEH activity at least by 50%; conversely, its overexpression inhibits the CE formation in macrophages. Immunohistochemistry reveals that NCEH is expressed in macrophage foam cells in atherosclerotic lesions. These data indicate that NCEH is responsible for a major part of nCEH activity in macrophages and may be a potential therapeutic target for the prevention of atherosclerosis. PMID:18782767

Okazaki, Hiroaki; Igarashi, Masaki; Nishi, Makiko; Sekiya, Motohiro; Tajima, Makiko; Takase, Satoru; Takanashi, Mikio; Ohta, Keisuke; Tamura, Yoshiaki; Okazaki, Sachiko; Yahagi, Naoya; Ohashi, Ken; Amemiya-Kudo, Michiyo; Nakagawa, Yoshimi; Nagai, Ryozo; Kadowaki, Takashi; Osuga, Jun-ichi; Ishibashi, Shun

2008-01-01

341

[Antioxidative enzymes play key roles in cadmium tolerance of Phytolacca americana].  

PubMed

Phytolacca americana L. has the capacity to take up and accumulate to very high levels heavy metals such as Mn and Cd, and is used for phytoextraction of heavy metal contaminated soils. The role of antioxidative enzyme of Phytolacca americana in response to Cd stress is unknown. The 6-week-old seedlings of Phytolacca americana were exposed to half strength Hoagland solution with 200 micromol/L CdCl2 or 400 micromol/L CdCl2 for 4 days. The content of H2O2 and MDA, and electrolyte leakage increased, while the photosynthetic rate decreased, indicated that the oxidative damage induced by Cd stress in Phytolacca americana was one of the metal toxicity mechanism. The activities of SOD and POD increased rapidly with elevated Cd concentration and exposure time, CAT activity was stable in response to 200 micromol/L CdCl2 stress, and increased only at 3 d later upon 400 micromol/L CdCl2, treatment. Suggested that the enzymatic antioxidation capacity played important role in Cd tolerance of hyperaccumulator plant. PMID:21634194

Zhang, Yu-Xiu; Zhang, Hong-Mei; Huang, Zhi-Bo; Li, Lin-Feng; Liu, Jin-Guang; Li, Xia

2011-03-01

342

Correlations among antral follicular echotexture, apoptosis and expression of key steroidogenic enzymes in sheep  

PubMed Central

Nineteen cycling ewes underwent transrectal ultrasonography of ovaries followed by ovariectomies during the growth phase of the first follicular wave of the interovulatory interval or the proestrus/estrus phase of the cycle. Quantitative ultrasonographic characteristics of the antrum and follicular wall in a total of forty-three ovine antral follicles were examined for correlations with the protein expression of three steroidogenic enzymes (cytochrome P450 17?-hydroxylase, CYP17; cytochrome P450 aromatase, CYP19; and 3?-hydroxysteroid dehydrogenase, 3?-HSD) determined by densitometric analysis of immunohistochemical slides, follicular dimensions, granulosa layer thickness and the percentage of apoptotic granulosa cells. Significant correlations were found between echotextural attributes of ovine antral follicles and the percentage of apoptotic granulosa cells, CYP17 expression (theca), CYP19 expression (granulosa) and 3?-HSD expression (theca cells). Computer-aided analyses of ultrasonographic images can be beneficial to the development of assisted reproductive technologies and diagnosis of hormonal imbalances without the need for ovarian biopsies or hormone assays. PMID:25109269

VANDUZER, Taylor; DUGGAVATHI, Raj; MURAWSKI, Maciej; ZIEBA, Dorota A.; SROKA, Patrycja; BARTLEWSKI, Pawel M.

2014-01-01

343

6-Gingerol Protects against Nutritional Steatohepatitis by Regulating Key Genes Related to Inflammation and Lipid Metabolism  

PubMed Central

Non-alcoholic fatty liver disease, including non-alcoholic steatohepatitis (NASH), appears to be increasingly common worldwide. The aim of the study was to investigate the effects of 6-gingerol ((S)-5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-3-decanone), a bioactive ingredient of plants belonging to the Zingiberaceae family, on experimental models of NASH. In HepG2 cells, 6-gingerol (100 ?mol/L) treatment inhibited free fatty acids mixture (0.33 mmol/L palmitate and 0.66 mmol/L oleate)-induced triglyceride and inflammatory marker accumulations. Male C57BL/6 mice were fed with a methionine and choline-deficient (MCD) diet to induce steatohepatitis. After four weeks of MCD diet feeding, the mice were dosed orally with 6-gingerol (25, 50 or 100 mg/kg/day) once daily for another four weeks. 6-Gingerol (100 mg/kg/day) attenuated liver steatosis and necro-inflammation in MCD diet-fed mice. The expressions of inflammatory cytokine genes, including those for monocyte chemoattractant protein-1, tumor necrosis factor-?, and interleukin-6, and nuclear transcription factor (NF-?B), which were increased in the livers of MCD diet-fed mice, were attenuated by 6-gingerol. 6-Gingerol possesses a repressive property on hepatic steatosis, which is associated with induction of peroxisome proliferator-activated receptor ?. Our study demonstrated the protective role of 6-gingerol in ameliorating nutritional steatohepatitis. The effect was mediated through regulating key genes related to lipid metabolism and inflammation. PMID:25658238

Tzeng, Thing-Fong; Liou, Shorong-Shii; Chang, Chia Ju; Liu, I-Min

2015-01-01

344

6-gingerol protects against nutritional steatohepatitis by regulating key genes related to inflammation and lipid metabolism.  

PubMed

Non-alcoholic fatty liver disease, including non-alcoholic steatohepatitis (NASH), appears to be increasingly common worldwide. The aim of the study was to investigate the effects of 6-gingerol ((S)-5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-3-decanone), a bioactive ingredient of plants belonging to the Zingiberaceae family, on experimental models of NASH. In HepG2 cells, 6-gingerol (100 ?mol/L) treatment inhibited free fatty acids mixture (0.33 mmol/L palmitate and 0.66 mmol/L oleate)-induced triglyceride and inflammatory marker accumulations. Male C57BL/6 mice were fed with a methionine and choline-deficient (MCD) diet to induce steatohepatitis. After four weeks of MCD diet feeding, the mice were dosed orally with 6-gingerol (25, 50 or 100 mg/kg/day) once daily for another four weeks. 6-Gingerol (100 mg/kg/day) attenuated liver steatosis and necro-inflammation in MCD diet-fed mice. The expressions of inflammatory cytokine genes, including those for monocyte chemoattractant protein-1, tumor necrosis factor-?, and interleukin-6, and nuclear transcription factor (NF-?B), which were increased in the livers of MCD diet-fed mice, were attenuated by 6-gingerol. 6-Gingerol possesses a repressive property on hepatic steatosis, which is associated with induction of peroxisome proliferator-activated receptor ?. Our study demonstrated the protective role of 6-gingerol in ameliorating nutritional steatohepatitis. The effect was mediated through regulating key genes related to lipid metabolism and inflammation. PMID:25658238

Tzeng, Thing-Fong; Liou, Shorong-Shii; Chang, Chia Ju; Liu, I-Min

2015-01-01

345

Variation in sulfur and selenium accumulation is controlled by naturally occurring isoforms of the key sulfur assimilation enzyme ADENOSINE 5'-PHOSPHOSULFATE REDUCTASE2 across the Arabidopsis species range.  

PubMed

Natural variation allows the investigation of both the fundamental functions of genes and their role in local adaptation. As one of the essential macronutrients, sulfur is vital for plant growth and development and also for crop yield and quality. Selenium and sulfur are assimilated by the same process, and although plants do not require selenium, plant-based selenium is an important source of this essential element for animals. Here, we report the use of linkage mapping in synthetic F2 populations and complementation to investigate the genetic architecture of variation in total leaf sulfur and selenium concentrations in a diverse set of Arabidopsis (Arabidopsis thaliana) accessions. We identify in accessions collected from Sweden and the Czech Republic two variants of the enzyme ADENOSINE 5'-PHOSPHOSULFATE REDUCTASE2 (APR2) with strongly diminished catalytic capacity. APR2 is a key enzyme in both sulfate and selenate reduction, and its reduced activity in the loss-of-function allele apr2-1 and the two Arabidopsis accessions Hodonín and Shahdara leads to a lowering of sulfur flux from sulfate into the reduced sulfur compounds, cysteine and glutathione, and into proteins, concomitant with an increase in the accumulation of sulfate in leaves. We conclude from our observation, and the previously identified weak allele of APR2 from the Shahdara accession collected in Tadjikistan, that the catalytic capacity of APR2 varies by 4 orders of magnitude across the Arabidopsis species range, driving significant differences in sulfur and selenium metabolism. The selective benefit, if any, of this large variation remains to be explored. PMID:25245030

Chao, Dai-Yin; Baraniecka, Patrycja; Danku, John; Koprivova, Anna; Lahner, Brett; Luo, Hongbing; Yakubova, Elena; Dilkes, Brian; Kopriva, Stanislav; Salt, David E

2014-11-01

346

Metabolism of diosbulbin B in vitro and in vivo in rats: formation of reactive metabolites and human enzymes involved.  

PubMed

Diosbulbin B (DB), a major constituent of the furano-norditerpenes in Dioscorea bulbifera Linn, exhibits potential antineoplasmic activity and hepatotoxicity. The metabolism and reactive metabolites of DB in vitro (with human and animal liver microsomes) and in vivo in rats were investigated. The human enzymes involved in DB metabolism were identified. DB was first catalyzed into reactive metabolites of 2-butene-1,4-dial derivatives dependent on NADPH and then trapped by Tris base or oxidized to hemiacetal lactones (M12 and M13) in microsomal incubations. Tris base was used as buffer constituent and as a trapping agent for aldehyde. Methoxylamine and glutathione (GSH) were also used as trapping agents. DB metabolism in vivo in rats after oral administration was consistent with that in vitro. The structures of M12 and M13, as well as mono-GSH conjugates of DB (M31), were confirmed by nuclear magnetic resonance spectroscopy of the chemically synthesized products. The bioactivation enzymes of DB were identified as CYP3A4/5, 2C9, and 2C19. CYP3A4 was found to be the primary enzyme using human recombinant cytochrome P450 enzymes, specific inhibitory studies, and a relative activity factor approach for pooled human liver microsomes. Michaelis-Menten constants K(m) and V(max) were determined by the formation of M31. The reactive metabolites may be related to the hepatotoxicity of DB. The gender difference in CYP3A expression in mice and rats contributed to the gender-related liver injury and pharmacokinetics in mice and rats, respectively. PMID:25053620

Yang, Baohua; Liu, Wei; Chen, Kaixian; Wang, Zhengtao; Wang, Changhong

2014-10-01

347

Persistent Overexpression of Phosphoglycerate Mutase, a Glycolytic Enzyme, Modifies Energy Metabolism and Reduces Stress Resistance of Heart in Mice  

PubMed Central

Background Heart failure is associated with changes in cardiac energy metabolism. Glucose metabolism in particular is thought to be important in the pathogenesis of heart failure. We examined the effects of persistent overexpression of phosphoglycerate mutase 2 (Pgam2), a glycolytic enzyme, on cardiac energy metabolism and function. Methods and Results Transgenic mice constitutively overexpressing Pgam2 in a heart-specific manner were generated, and cardiac energy metabolism and function were analyzed. Cardiac function at rest was normal. The uptake of analogs of glucose or fatty acids and the phosphocreatine/?ATP ratio at rest were normal. A comprehensive metabolomic analysis revealed an increase in the levels of a few metabolites immediately upstream and downstream of Pgam2 in the glycolytic pathway, whereas the levels of metabolites in the initial few steps of glycolysis and lactate remained unchanged. The levels of metabolites in the tricarboxylic acid (TCA) cycle were altered. The capacity for respiration by isolated mitochondria in vitro was decreased, and that for the generation of reactive oxygen species (ROS) in vitro was increased. Impaired cardiac function was observed in response to dobutamine. Mice developed systolic dysfunction upon pressure overload. Conclusions Constitutive overexpression of Pgam2 modified energy metabolism and reduced stress resistance of heart in mice. PMID:23951293

Shioi, Tetsuo; Kato, Takao; Inuzuka, Yasutaka; Kawashima, Tsuneaki; Tamaki, Yodo; Kawamoto, Akira; Tanada, Yohei; Iwanaga, Yoshitaka; Narazaki, Michiko; Matsuda, Tetsuya; Adachi, Souichi; Soga, Tomoyoshi; Takemura, Genzou; Kondoh, Hiroshi; Kita, Toru; Kimura, Takeshi

2013-01-01

348

Chromium picolinate does not improve key features of metabolic syndrome in obese nondiabetic adults  

Technology Transfer Automated Retrieval System (TEKTRAN)

The use of chromium-containing dietary supplements is widespread among patients with type 2 diabetes as a means of improving glucose metabolism. However, chromium’s role as a potential therapy for patients at high risk for developing type 2 diabetes, specifically those with metabolic syndrome, is n...

349

Caffeine as a metabolic probe: Exploration of the enzyme-inducing effect of cigarette smoking  

Microsoft Academic Search

It has been realized recently that the primary metabolism of caffeine in humans is catalyzed by P-450IA2 and that the rate of caffeine metabolism can be estimated from a metabolic ratio in a single urine sample. A population of 178 students including 19 smokers were subjected to this caffeine test to establish their P-450IA2 index. Both stated numbers of cigarettes

Werner Kalow; Bing-Kou Tang

1991-01-01

350

The Regulation of Alfalfa Saponin Extract on Key Genes Involved in Hepatic Cholesterol Metabolism in Hyperlipidemic Rats  

PubMed Central

To investigate the cholesterol-lowering effects of alfalfa saponin extract (ASE) and its regulation mechanism on some key genes involved in cholesterol metabolism, 40 healthy 7 weeks old male Sprague Dawley (SD) rats were randomly divided into four groups with 10 rats in each group: control group, hyperlipidemic group, ASE treatment group, ASE prevention group. The body weight gain, relative liver weight and serum lipid 1evels of rats were determined. Total cholesterol (TC) and total bile acids (TBA) levels in liver and feces were also measured. Furthermore, the activity and mRNA expressions of Hmgcr, Acat2, Cyp7a1 and Ldlr were investigated. The results showed the following: (1) The abnormal serum lipid levels in hyperlipidemic rats were ameliorated by ASE administration (both ASE prevention group and treatment group) (P<0.05). (2) Both ASE administration to hyperlipidemic rats significantly reduced liver TC and increased liver TBA level (P<0.05). TC and TBA levels in feces of hyperlipidemic rats were remarkably elevated by both ASE administration (P<0.05). (3) mRNA expressions of Hmgcr and Acat2 in the liver of hyperlipidemic rats were remarkably down-regulated (P<0.05), as well as mRNA expressions of Cyp7a1 and Ldlr were dramatically up-regulated by both ASE administration (P<0.05). The activities of these enzymes also paralleled the observed changes in mRNA levels. (4) There was no significant difference between ASE treatment and ASE prevention group for most parameters evaluated. Our present study indicated that ASE had cholesterol-lowering effects. The possible mechanism could be attributed to (1) the down-regulation of Hmgcr and Acat2, as well as up-regulation of Cyp7a1 and Ldlr in the liver of hyperlipidemic rats, which was involved in cholesterol biosynthesis, uptake, and efflux pathway; (2) the increase in excretion of cholesterol. The findings in our study suggested ASE had great potential usefulness as a natural agent for treating hyperlipidemia. PMID:24505463

Shi, Yinghua; Guo, Rui; Wang, Xianke; Yuan, Dedi; Zhang, Senhao; Wang, Jie; Yan, Xuebing; Wang, Chengzhang

2014-01-01

351

The regulation of alfalfa saponin extract on key genes involved in hepatic cholesterol metabolism in hyperlipidemic rats.  

PubMed

To investigate the cholesterol-lowering effects of alfalfa saponin extract (ASE) and its regulation mechanism on some key genes involved in cholesterol metabolism, 40 healthy 7 weeks old male Sprague Dawley (SD) rats were randomly divided into four groups with 10 rats in each group: control group, hyperlipidemic group, ASE treatment group, ASE prevention group. The body weight gain, relative liver weight and serum lipid 1evels of rats were determined. Total cholesterol (TC) and total bile acids (TBA) levels in liver and feces were also measured. Furthermore, the activity and mRNA expressions of Hmgcr, Acat2, Cyp7a1 and Ldlr were investigated. The results showed the following: (1) The abnormal serum lipid levels in hyperlipidemic rats were ameliorated by ASE administration (both ASE prevention group and treatment group) (P<0.05). (2) Both ASE administration to hyperlipidemic rats significantly reduced liver TC and increased liver TBA level (P<0.05). TC and TBA levels in feces of hyperlipidemic rats were remarkably elevated by both ASE administration (P<0.05). (3) mRNA expressions of Hmgcr and Acat2 in the liver of hyperlipidemic rats were remarkably down-regulated (P<0.05), as well as mRNA expressions of Cyp7a1 and Ldlr were dramatically up-regulated by both ASE administration (P<0.05). The activities of these enzymes also paralleled the observed changes in mRNA levels. (4) There was no significant difference between ASE treatment and ASE prevention group for most parameters evaluated. Our present study indicated that ASE had cholesterol-lowering effects. The possible mechanism could be attributed to (1) the down-regulation of Hmgcr and Acat2, as well as up-regulation of Cyp7a1 and Ldlr in the liver of hyperlipidemic rats, which was involved in cholesterol biosynthesis, uptake, and efflux pathway; (2) the increase in excretion of cholesterol. The findings in our study suggested ASE had great potential usefulness as a natural agent for treating hyperlipidemia. PMID:24505463

Shi, Yinghua; Guo, Rui; Wang, Xianke; Yuan, Dedi; Zhang, Senhao; Wang, Jie; Yan, Xuebing; Wang, Chengzhang

2014-01-01

352

Fine quantitative trait loci mapping of carbon and nitrogen metabolism enzyme activities and seedling biomass in the intermated maize IBM mapping population  

Technology Transfer Automated Retrieval System (TEKTRAN)

Understanding the genetic basis of nitrogen and carbon metabolism will accelerate development of plant varieties with high yield and improved nitrogen use efficiency. In this study, we measured the activities of ten enzymes from carbon and nitrogen metabolism and seedling/juvenile biomass in the mai...

353

Identification of mitogen-activated protein kinase docking sites in enzymes that metabolize phosphatidylinositols and inositol phosphates  

PubMed Central

Background Reversible interactions between the components of cellular signaling pathways allow for the formation and dissociation of multimolecular complexes with spatial and temporal resolution and, thus, are an important means of integrating multiple signals into a coordinated cellular response. Several mechanisms that underlie these interactions have been identified, including the recognition of specific docking sites, termed a D-domain and FXFP motif, on proteins that bind mitogen-activated protein kinases (MAPKs). We recently found that phosphatidylinositol-specific phospholipase C-?1 (PLC-?1) directly binds to extracellular signal-regulated kinase 2 (ERK2), a MAPK, via a D-domain-dependent mechanism. In addition, we identified D-domain sequences in several other PLC isozymes. In the present studies we sought to determine whether MAPK docking sequences could be recognized in other enzymes that metabolize phosphatidylinositols (PIs), as well as in enzymes that metabolize inositol phosphates (IPs). Results We found that several, but not all, of these enzymes contain identifiable D-domain sequences. Further, we found a high degree of conservation of these sequences and their location in human and mouse proteins; notable exceptions were PI 3-kinase C2-?, PI 4-kinase type II?, and inositol polyphosphate 1-phosphatase. Conclusion The results indicate that there may be extensive crosstalk between MAPK signaling and signaling pathways that are regulated by cellular levels of PIs or IPs. PMID:16445858

Caldwell, Kevin K; Sosa, Marcos; Buckley, Colin T

2006-01-01

354

Molecular identification of microsomal acyl-CoA:glycerol-3-phosphate acyltransferase, a key enzyme in de novo triacylglycerol synthesis  

PubMed Central

Acyl-CoA:glycerol-3-phosphate acyltransferase (GPAT) catalyzes the first step during de novo synthesis of triacylglycerol. It has been well recognized that mammals possess multiple enzymatically distinct proteins with GPAT activity. Although the mitochondrial-associated GPAT has been cloned and extensively characterized, the molecular identity of the endoplasmic reticulum (ER)-associated GPAT, which accounts for the majority of total GPAT activity in most tissues, has remained elusive. Here we report the identification of genes encoding human and mouse ER-associated GPAT (termed GPAT3). GPAT3 is a member of the acyltransferase family predominantly expressed in tissues characterized by active lipid metabolism, such as adipose tissue, small intestine, kidney, and heart. Ectopic expression of GPAT3 leads to a significant increase in N-ethylmaleimide-sensitive GPAT activity, whereas acyltransferase activity toward a variety of other lysophospholipids, as well as neutral lipid substrates, is not altered. Overexpression of GPAT3 in mammalian cells results in increased triacylglycerol, but not phospholipid, formation. GPAT3 is localized to the ER when overexpressed in COS-7 cells. GPAT3 mRNA is dramatically up-regulated during adipocyte differentiation, is reciprocally regulated in adipose tissue and liver of ob/ob mice, and is up-regulated in mice treated with a peroxisome proliferator-activated receptor ? (PPAR?) agonist. A substantial loss of GPAT activity in 3T3-L1 adipocytes was achieved by reducing GPAT3 mRNA levels through GPAT3-specific siRNA knockdown. These findings identify GPAT3 as a previously uncharacterized triacylglycerol biosynthetic enzyme. Similar to other lipogenic enzymes, GPAT3 may be useful as a target for the treatment of obesity. PMID:17170135

Cao, Jingsong; Li, Jian-Liang; Li, Dongmei; Tobin, James F.; Gimeno, Ruth E.

2006-01-01

355

COMPARATIVE ENZYME INDUCTION AND LINDANE METABOLISM IN RATS PRE-TREATED WITH VARIOUS ORGANOCHLORINE PESTICIDES  

EPA Science Inventory

The comparative effect of 7 days pre-treatment with one of seven organochlorine pesticides on the metabolism of lindane in vivo and on the metabolism of EPN, p-nitroanisole and methyl orange in vitro was investigated. Mirex was the most potent inducer of the oxidative hydrolysis ...

356

GENE EXPRESSION PROFILING OF XENOBIOTIC METABOLIZING ENZYMES (XMES) IN THE AGING MALE FISHER RAT  

EPA Science Inventory

Detoxification and elimination of xenobiotics is a major function of the liver and is important in maintaining the metabolic homeostasis of the organism. The degree to which aging affects hepatic metabolism is not known. The expression of XMEs, in part, determines the fate of the...

357

The Action of Antidiabetic Plants of the Canadian James Bay Cree Traditional Pharmacopeia on Key Enzymes of Hepatic Glucose Homeostasis  

PubMed Central

We determined the capacity of putative antidiabetic plants used by the Eastern James Bay Cree (Canada) to modulate key enzymes of gluconeogenesis and glycogen synthesis and key regulating kinases. Glucose-6-phosphatase (G6Pase) and glycogen synthase (GS) activities were assessed in cultured hepatocytes treated with crude extracts of seventeen plant species. Phosphorylation of AMP-dependent protein kinase (AMPK), Akt, and Glycogen synthase kinase-3 (GSK-3) were probed by Western blot. Seven of the seventeen plant extracts significantly decreased G6Pase activity, Abies balsamea and Picea glauca, exerting an effect similar to insulin. This action involved both Akt and AMPK phosphorylation. On the other hand, several plant extracts activated GS, Larix laricina and A. balsamea, far exceeding the action of insulin. We also found a significant correlation between GS stimulation and GSK-3 phosphorylation induced by plant extract treatments. In summary, three Cree plants stand out for marked effects on hepatic glucose homeostasis. P. glauca affects glucose production whereas L. laricina rather acts on glucose storage. However, A. balsamea has the most promising profile, simultaneously and powerfully reducing G6Pase and stimulating GS. Our studies thus confirm that the reduction of hepatic glucose production likely contributes to the therapeutic potential of several antidiabetic Cree traditional medicines. PMID:23864882

Nachar, Abir; Vallerand, Diane; Musallam, Lina; Lavoie, Louis; Arnason, John; Haddad, Pierre S.

2013-01-01

358

The action of antidiabetic plants of the canadian james bay cree traditional pharmacopeia on key enzymes of hepatic glucose homeostasis.  

PubMed

We determined the capacity of putative antidiabetic plants used by the Eastern James Bay Cree (Canada) to modulate key enzymes of gluconeogenesis and glycogen synthesis and key regulating kinases. Glucose-6-phosphatase (G6Pase) and glycogen synthase (GS) activities were assessed in cultured hepatocytes treated with crude extracts of seventeen plant species. Phosphorylation of AMP-dependent protein kinase (AMPK), Akt, and Glycogen synthase kinase-3 (GSK-3) were probed by Western blot. Seven of the seventeen plant extracts significantly decreased G6Pase activity, Abies balsamea and Picea glauca, exerting an effect similar to insulin. This action involved both Akt and AMPK phosphorylation. On the other hand, several plant extracts activated GS, Larix laricina and A. balsamea, far exceeding the action of insulin. We also found a significant correlation between GS stimulation and GSK-3 phosphorylation induced by plant extract treatments. In summary, three Cree plants stand out for marked effects on hepatic glucose homeostasis. P. glauca affects glucose production whereas L. laricina rather acts on glucose storage. However, A. balsamea has the most promising profile, simultaneously and powerfully reducing G6Pase and stimulating GS. Our studies thus confirm that the reduction of hepatic glucose production likely contributes to the therapeutic potential of several antidiabetic Cree traditional medicines. PMID:23864882

Nachar, Abir; Vallerand, Diane; Musallam, Lina; Lavoie, Louis; Badawi, Alaa; Arnason, John; Haddad, Pierre S

2013-01-01

359

Absolute quantification of Medicago truncatula sucrose synthase isoforms and N-metabolism enzymes in symbiotic root nodules and the detection of novel nodule phosphoproteins by mass spectrometry.  

PubMed

Mass spectrometry (MS) has become increasingly important for tissue specific protein quantification at the isoform level, as well as for the analysis of protein post-translational regulation mechanisms and turnover rates. Thanks to the development of high accuracy mass spectrometers, peptide sequencing without prior knowledge of the amino acid sequence--de novo sequencing--can be performed. In this work, absolute quantification of a set of key enzymes involved in carbon and nitrogen metabolism in Medicago truncatula 'Jemalong A17' root nodules is presented. Among them, sucrose synthase (SuSy; EC 2.4.1.13), one of the central enzymes in sucrose cleavage in root nodules, has been further characterized and the relative phosphorylation state of the three most abundant isoforms has been quantified. De novo sequencing provided sequence information of a so far unidentified peptide, most probably belonging to SuSy2, the second most abundant isoform in M. truncatula root nodules. TiO(2)-phosphopeptide enrichment led to the identification of not only a phosphorylation site at Ser11 in SuSy1, but also of several novel phosphorylation sites present in other root nodule proteins such as alkaline invertase (AI; EC 3.2.1.26) and an RNA-binding protein. PMID:18772307

Wienkoop, Stefanie; Larrainzar, Estíbaliz; Glinski, Mirko; González, Esther M; Arrese-Igor, Cesar; Weckwerth, Wolfram

2008-01-01

360

Diminution in phase 1 and phase 2 drug metabolizing enzymes of rat lung by asbestos: An in vitro study  

SciTech Connect

In the present paper studies are presented concerning the effect of three varieties of asbestos namely, chrysotile, crocidolite and amosite, and of titanium dioxide, an inert non-asbestos dust, on the enzymes of phase 1 and phase 2 drug metabolism in isolated rat lung microsomes and post-microsomal fraction. Since 3-methylcholanthrene (3-MC) is known to induce cytochromes P-450 in the IA family and their associated activity, benzo(a)pyrene hydroxylase, therefore, the effect of these mineral fibers on cytochrome P-450 and benzo(a)pyrene hydroxylase in lung microsomes isolated from 3-MC-treated rats was studied.

Khan, S.G.; Ali, S.; Rahman, Q. (Industrial Toxicology Research Centre, Lucknow (India))

1991-11-01

361

Enzymes of galactose metabolism in livers of suckling and adult tammar wallabies (Macropus eugenii) and other marsupials.  

PubMed

The activities of galactokinase, hexose-1-phosphate uridylyl transferase and UDPglucose 4-epimerase in homogenates of livers of two adult and 20 suckling tammar wallabies aged from 6 to 50 weeks were investigated. The activities of all three enzymes were high until 24-30 weeks post partum, after which they declined to low levels. The activities of the three liver enzymes were high in pouch young of six other species of marsupial. Comparison of the activities of the three liver enzymes in suckling tammar wallabies with those in suckling rats showed no difference between the two species in regard to galactokinase and uridylyl transferase, but the UDPglucose 4-epimerase activity in tammar wallabies was approximately double than found in rats. This may be related to the high galactose content of tammar wallaby milk compared with rat milk. In suckling tammar wallabies, the liver had higher enzyme activities than other tissues studied. It is concluded that, contrary to the suggestion of Stephens et al. (1975), pouch young marsupials are not deficient in their ability to metabolize galactose. PMID:6272674

Vernon, J K; Messer, M; Green, B

1981-01-01

362

High sensitivity of Nrf2 knockout mice to acetaminophen hepatotoxicity associated with decreased expression of ARE-regulated drug metabolizing enzymes and antioxidant genes.  

PubMed

Nrf2, which belongs to the basic leucine zipper (bZip) transcription factor family, has been implicated as a key molecule involved in antioxidant-responsive element (ARE)-mediated gene expression. In order to examine the role of Nrf2 in protection against xenobiotic toxicity, the sensitivity of nrf2 knockout mice to acetaminophen (N-acetyl-4-aminophenol (APAP)) was analyzed. The saturation of detoxification pathways after high levels of exposure to APAP is known to induce hepatotoxicity. Two factors important in its detoxification are UDP-glucuronosyltransferase (UDP-GT), an ARE-regulated phase-II drug-metabolizing enzyme, and glutathione (GSH), an antioxidant molecule whose synthesis depends on ARE-regulated gamma-glutamylcysteine synthetase (gammaGCS). Two- to 4-month-old male mice were orally administered a single dose of APAP at 0, 150, 300, or 600 mg/kg. Doses of 300 mg/kg APAP or greater caused death in the homozygous knockout mice only, and those that survived showed a greater severity in hepatic damage than the wild-type mice, as demonstrated by increased plasma alanine aminotransferase activity, decreased hepatic non-protein sulfhydryl (NPSH) content, and centrilobular hepatocellular necrosis. The high sensitivity of Nrf2-deficient mice was confirmed from observations made at 0, 2, 8, and 24 h after dosing with 300 mg/kg APAP; increased anti-APAP immunoreactivity was also noted in their livers at 2 h. Untreated homozygous knockout mice showed both a lower UDP-GT activity and NPSH content, which corresponded to decreased mRNA levels of UDP-GT (Ugt1a6) and the heavy chain of gammaGCS, respectively. These results show that Nrf2 plays a protective role against APAP hepatotoxicity by regulating both drug metabolizing enzymes and antioxidant genes through the ARE. PMID:11134556

Enomoto, A; Itoh, K; Nagayoshi, E; Haruta, J; Kimura, T; O'Connor, T; Harada, T; Yamamoto, M

2001-01-01

363

[Effects of krestin and p-aminobenzoic-N-xyloside sodium salt on activities of drug-metabolizing enzymes and glutathione-related enzymes in rat liver].  

PubMed

Effects of krestin (PSK) and p-aminobenzoic-N-xyloside sodium salt (K-247) (both products of Kreha Chemical Co., Japan) on activities of drug-metabolizing enzymes and glutathione (GSH)-related enzymes were investigated in rat liver. When PSK was administered at a dose of 100 mg/kg body weight, ip every day for 7 or 14 days, the action of UDP-glucuronyltransferase (UDP-GT) on o-aminophenol (o-GT) and those of GSH S-transferase (GST) on both 1, 2-dichloro-4-nitrobenzene ( DCNB ) and 1-chloro-2, 4-dinitrobenzene (CDNB) slightly increased together with increased activities of GSH-peroxidase on both H2O2 and cumene hydroperoxide, were as GSH levels were decreased. When PSK or K-247 was administered at 1% in diet for 4 or 8 weeks, o-GT activity and GST activities with both substrates increased on K-247 feeding, while GST activity for CDNB decreased on PSK and K-247 feedings. These changes were statistically significant but very small. The content of P-450 and the activity of gamma-glutamyl transpeptidase changed little in any administration schedules mentioned above. PMID:6144291

Ebina, T; Kitahara, A; Soma, Y; Ishikawa, T; Tsuchida, S; Sato, K

1984-05-01

364

Study of 'Redhaven' peach and its white-fleshed mutant suggests a key role of CCD4 carotenoid dioxygenase in carotenoid and norisoprenoid volatile metabolism  

PubMed Central

Background Carotenoids are plant metabolites which are not only essential in photosynthesis but also important quality factors in determining the pigmentation and aroma of flowers and fruits. To investigate the regulation of carotenoid metabolism, as related to norisoprenoids and other volatile compounds in peach (Prunus persica L. Batsch.), and the role of carotenoid dioxygenases in determining differences in flesh color phenotype and volatile composition, the expression patterns of relevant carotenoid genes and metabolites were studied during fruit development along with volatile compound content. Two contrasted cultivars, the yellow-fleshed 'Redhaven' (RH) and its white-fleshed mutant 'Redhaven Bianca' (RHB) were examined. Results The two genotypes displayed marked differences in the accumulation of carotenoid pigments in mesocarp tissues. Lower carotenoid levels and higher levels of norisoprenoid volatiles were observed in RHB, which might be explained by differential activity of carotenoid cleavage dioxygenase (CCD) enzymes. In fact, the ccd4 transcript levels were dramatically higher at late ripening stages in RHB with respect to RH. The two genotypes also showed differences in the expression patterns of several carotenoid and isoprenoid transcripts, compatible with a feed-back regulation of these transcripts. Abamine SG - an inhibitor of CCD enzymes - decreased the levels of both isoprenoid and non-isoprenoid volatiles in RHB fruits, indicating a complex regulation of volatile production. Conclusions Differential expression of ccd4 is likely to be the major determinant in the accumulation of carotenoids and carotenoid-derived volatiles in peach fruit flesh. More in general, dioxygenases appear to be key factors controlling volatile composition in peach fruit, since abamine SG-treated 'Redhaven Bianca' fruits had strongly reduced levels of norisoprenoids and other volatile classes. Comparative functional studies of peach carotenoid cleavage enzymes are required to fully elucidate their role in peach fruit pigmentation and aroma. PMID:21269483

2011-01-01

365

Metabolic mechanisms of methanol/formaldehyde in isolated rat hepatocytes: carbonyl-metabolizing enzymes versus oxidative stress.  

PubMed

Methanol (CH(3)OH), a common industrial solvent, is metabolized to toxic compounds by several enzymatic as well as free radical pathways. Identifying which process best enhances or prevents CH(3)OH-induced cytotoxicity could provide insight into the molecular basis for acute CH(3)OH-induced hepatoxicity. Metabolic pathways studied include those found in 1) an isolated hepatocyte system and 2) cell-free systems. Accelerated Cytotoxicity Mechanism Screening (ACMS) techniques demonstrated that CH(3)OH had little toxicity towards rat hepatocytes in 95% O(2), even at 2M concentration, whereas 50 mM was the estimated LC(50) (2h) in 1% O(2), estimated to be the physiological concentration in the centrilobular region of the liver and also the target region for ethanol toxicity. Cytotoxicity was attributed to increased NADH levels caused by CH(3)OH metabolism, catalyzed by ADH1, resulting in reductive stress, which reduced and released ferrous iron from Ferritin causing oxygen activation. A similar cytotoxic mechanism at 1% O(2) was previous found for ethanol. With 95% O(2), the addition of Fe(II)/H(2)O(2), at non-toxic concentrations were the most effective agents for increasing hepatocyte toxicity induced by 1M CH(3)OH, with a 3-fold increase in cytotoxicity and ROS formation. Iron chelators, desferoxamine, and NADH oxidizers and ATP generators, e.g. fructose, also protected hepatocytes and decreased ROS formation and cytotoxicity. Hepatocyte protein carbonylation induced by formaldehyde (HCHO) formation was also increased about 4-fold, when CH(3)OH was oxidized by the Fenton-like system, Fe(II)/H(2)O(2), and correlated with increased cytotoxicity. In a cell-free bovine serum albumin system, Fe(II)/H(2)O(2) also increased CH(3)OH oxidation as well as HCHO protein carbonylation. Nontoxic ferrous iron and a H(2)O(2) generating system increased HCHO-induced cytotoxicity and hepatocyte protein carbonylation. In addition, HCHO cytotoxicity was markedly increased by ADH1 and ALDH2 inhibitors or GSH-depleted hepatocytes. Increased HCHO concentration levels correlated with increased HCHO-induced protein carbonylation in hepatocytes. These results suggest that CH(3)OH at 1% O(2) involves activation of the Fenton system to form HCHO. However, at higher O(2) levels, radicals generated through Fe(II)/H(2)O(2) can oxidize CH(3)OH/HCHO to form pro-oxidant radicals and lead to increased oxidative stress through protein carbonylation and ROS formation which ultimately causes cell death. PMID:21276436

MacAllister, Stephanie L; Choi, Joanna; Dedina, Liana; O'Brien, Peter J

2011-05-30

366

The evolution of new enzyme function: lessons from xenobiotic metabolizing bacteria versus insecticide-resistant insects  

PubMed Central

Here, we compare the evolutionary routes by which bacteria and insects have evolved enzymatic processes for the degradation of four classes of synthetic chemical insecticide. For insects, the selective advantage of such degradative activities is survival on exposure to the insecticide, whereas for the bacteria the advantage is simply a matter of access to additional sources of nutrients. Nevertheless, bacteria have evolved highly efficient enzymes from a wide variety of enzyme families, whereas insects have relied upon generalist esterase-, cytochrome P450- and glutathione-S-transferase-dependent detoxification systems. Moreover, the mutant insect enzymes are less efficient kinetically and less diverged in sequence from their putative ancestors than their bacterial counterparts. This presumably reflects several advantages that bacteria have over insects in the acquisition of new enzymatic functions, such as a broad biochemical repertoire from which new functions can be evolved, large population sizes, high effective mutation rates, very short generation times and access to genetic diversity through horizontal gene transfer. Both the insect and bacterial systems support recent theory proposing that new biochemical functions often evolve from ‘promiscuous’ activities in existing enzymes, with subsequent mutations then enhancing those activities. Study of the insect enzymes will help in resistance management, while the bacterial enzymes are potential bioremediants of insecticide residues in a range of contaminated environments. PMID:25567970

Russell, Robyn J; Scott, Colin; Jackson, Colin J; Pandey, Rinku; Pandey, Gunjan; Taylor, Matthew C; Coppin, Christopher W; Liu, Jian-Wei; Oakeshott, John G

2011-01-01

367

Phase I to II cross-induction of xenobiotic metabolizing enzymes: A feedforward control mechanism for potential hormetic responses  

SciTech Connect

Hormetic responses to xenobiotic exposure likely occur as a result of overcompensation by the homeostatic control systems operating in biological organisms. However, the mechanisms underlying overcompensation that leads to hormesis are still unclear. A well-known homeostatic circuit in the cell is the gene induction network comprising phase I, II and III metabolizing enzymes, which are responsible for xenobiotic detoxification, and in many cases, bioactivation. By formulating a differential equation-based computational model, we investigated in this study whether hormesis can arise from the operation of this gene/enzyme network. The model consists of two feedback and one feedforward controls. With the phase I negative feedback control, xenobiotic X activates nuclear receptors to induce cytochrome P450 enzyme, which bioactivates X into a reactive metabolite X'. With the phase II negative feedback control, X' activates transcription factor Nrf2 to induce phase II enzymes such as glutathione S-transferase and glutamate cysteine ligase, etc., which participate in a set of reactions that lead to the metabolism of X' into a less toxic conjugate X''. The feedforward control involves phase I to II cross-induction, in which the parent chemical X can also induce phase II enzymes directly through the nuclear receptor and indirectly through transcriptionally upregulating Nrf2. As a result of the active feedforward control, a steady-state hormetic relationship readily arises between the concentrations of the reactive metabolite X' and the extracellular parent chemical X to which the cell is exposed. The shape of dose-response evolves over time from initially monotonically increasing to J-shaped at the final steady state-a temporal sequence consistent with adaptation-mediated hormesis. The magnitude of the hormetic response is enhanced by increases in the feedforward gain, but attenuated by increases in the bioactivation or phase II feedback loop gains. Our study suggests a possibly common mechanism for the hormetic responses observed with many mutagens/carcinogens whose activities require bioactivation by phase I enzymes. Feedforward control, often operating in combination with negative feedback regulation in a homeostatic system, may be a general control theme responsible for steady-state hormesis.

Zhang Qiang [Division of Computational Biology, Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States)], E-mail: qzhang@thehamner.org; Pi Jingbo [Division of Translational Biology, Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); Woods, Courtney G. [Division of Computational Biology, Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States); ExxonMobil Biomedical Sciences, Annandale, NJ 08801 (United States); Andersen, Melvin E. [Division of Computational Biology, Hamner Institutes for Health Sciences, 6 Davis Drive, Research Triangle Park, NC 27709 (United States)

2009-06-15

368

Phase I to II Cross-Induction of Xenobiotic Metabolizing Enzymes: a Feedforward Control Mechanism for Potential Hormetic Responses  

PubMed Central

Hormetic responses to xenobiotic exposure likely occur as a result of overcompensation by the homeostatic control systems operating in biological organisms. However, the mechanisms underlying overcompensation that leads to hormesis are still unclear. A well-known homeostatic circuit in the cell is the gene induction network comprising phase I, II and III metabolizing enzymes, which are responsible for xenobiotic detoxification, and in many cases, bioactivation. By formulating a differential equation-based computational model, we investigated in this study whether hormesis can arise from the operation of this gene/enzyme network. The model consists of two feedback and one feedforward controls. With the phase I negative feedback control, xenobiotic X activates nuclear receptors to induce cytochrome P450 enzyme, which bioactivates X into a reactive metabolite X?. With the phase II negative feedback control, X? activates transcription factor Nrf2 to induce phase II enzymes such as glutathione S-transferase and glutamate cysteine ligase, etc., which participate in a set of reactions that lead to the metabolism of X? into a less toxic conjugate X?. The feedforward control involves phase I to II cross-induction, in which the parent chemical X can also induce phase II enzymes directly through the nuclear receptor and indirectly through transcriptionally upregulating Nrf2. As a result of the active feedforward control, a steady-state hormetic relationship readily arises between the concentrations of the reactive metabolite X? and the extracellular parent chemical X to which the cell is exposed. The shape of dose response evolves over time from initially monotonically increasing to J-shaped at the final steady state - a temporal sequence consistent with adaptation-mediated hormesis. The magnitude of the hormetic response is enhanced by increases in the feedforward gain, but attenuated by increases in the bioactivation or phase II feedback loop gains. Our study suggests a possibly common mechanism for the hormetic responses observed with many mutagens/carcinogens whose activities require bioactivation by phase I enzymes. Feedforward control, often operating in combination with negative feedback regulation in a homeostatic system, may be a general control theme responsible for steady-state hormesis. PMID:19371757

Zhang, Qiang; Pi, Jingbo; Woods, Courtney G.; Andersen, Melvin E.

2009-01-01

369

Phosphotransferase protein EIIANtr interacts with SpoT, a key enzyme of the stringent response, in Ralstonia eutropha H16.  

PubMed

EIIA(Ntr) is a member of a truncated phosphotransferase (PTS) system that serves regulatory functions and exists in many Proteobacteria in addition to the sugar transport PTS. In Escherichia coli, EIIA(Ntr) regulates K(+) homeostasis through interaction with the K(+) transporter TrkA and sensor kinase KdpD. In the ?-Proteobacterium Ralstonia eutropha H16, EIIA(Ntr) influences formation of the industrially important bioplastic poly(3-hydroxybutyrate) (PHB). PHB accumulation is controlled by the stringent response and induced under conditions of nitrogen deprivation. Knockout of EIIA(Ntr) increases the PHB content. In contrast, absence of enzyme I or HPr, which deliver phosphoryl groups to EIIA(Ntr), has the opposite effect. To clarify the role of EIIA(Ntr) in PHB formation, we screened for interacting proteins that co-purify with Strep-tagged EIIA(Ntr) from R. eutropha cells. This approach identified the bifunctional ppGpp synthase/hydrolase SpoT1, a key enzyme of the stringent response. Two-hybrid and far-Western analyses confirmed the interaction and indicated that only non-phosphorylated EIIA(Ntr) interacts with SpoT1. Interestingly, this interaction does not occur between the corresponding proteins of E. coli. Vice versa, interaction of EIIA(Ntr) with KdpD appears to be absent in R. eutropha, although R. eutropha EIIA(Ntr) can perfectly substitute its homologue in E. coli in regulation of KdpD activity. Thus, interaction with KdpD might be an evolutionary 'ancient' task of EIIA(Ntr) that was subsequently replaced by interaction with SpoT1 in R. eutropha. In conclusion, EIIA(Ntr) might integrate information about nutritional status, as reflected by its phosphorylation state, into the stringent response, thereby controlling cellular PHB content in R. eutropha. PMID:24515609

Karstens, Katja; Zschiedrich, Christopher P; Bowien, Botho; Stülke, Jörg; Görke, Boris

2014-04-01

370

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions  

PubMed Central

Acetic acid bacteria (AAB) play an important role during cocoa fermentation, as their main product, acetate, is a major driver for the development of the desired cocoa flavors. Here, we investigated the specialized metabolism of these bacteria under cocoa pulp fermentation-simulating conditions. A carefully designed combination of parallel 13C isotope labeling experiments allowed the elucidation of intracellular fluxes in the complex environment of cocoa pulp, when lactate and ethanol were included as primary substrates among undefined ingredients. We demonstrate that AAB exhibit a functionally separated metabolism during coconsumption of two-carbon and three-carbon substrates. Acetate is almost exclusively derived from ethanol, while lactate serves for the formation of acetoin and biomass building blocks. Although this is suboptimal for cellular energetics, this allows maximized growth and conversion rates. The functional separation results from a lack of phosphoenolpyruvate carboxykinase and malic enzymes, typically present in bacteria to interconnect metabolism. In fact, gluconeogenesis is driven by pyruvate phosphate dikinase. Consequently, a balanced ratio of lactate and ethanol is important for the optimum performance of AAB. As lactate and ethanol are individually supplied by lactic acid bacteria and yeasts during the initial phase of cocoa fermentation, respectively, this underlines the importance of a well-balanced microbial consortium for a successful fermentation process. Indeed, AAB performed the best and produced the largest amounts of acetate in mixed culture experiments when lactic acid bacteria and yeasts were both present. PMID:24837393

Adler, Philipp; Frey, Lasse Jannis; Berger, Antje; Bolten, Christoph Josef; Hansen, Carl Erik

2014-01-01

371

Toward "stable-on-the-table" enzymes: improving key properties of catalase by covalent conjugation with poly(acrylic acid).  

PubMed

Several key properties of catalase such as thermal stability, resistance to protease degradation, and resistance to ascorbate inhibition were improved, while retaining its structure and activity, by conjugation to poly(acrylic acid) (PAA, Mw 8000) via carbodiimide chemistry where the amine groups on the protein are appended to the carboxyl groups of the polymer. Catalase conjugation was examined at three different pH values (pH 5.0, 6.0, and 7.0) and at three distinct mole ratios (1:100, 1:500, and 1:1000) of catalase to PAA at each reaction pH. The corresponding products are labeled as Cat-PAA(x)-y, where x is the protein to polymer mole ratio and y is the pH used for the synthesis. The coupling reaction consumed about 60-70% of the primary amines on the catalase; all samples were completely water-soluble and formed nanogels, as evidenced by gel electrophoresis and electron microscopy. The UV circular dichroism (CD) spectra indicated substantial retention of protein secondary structure for all samples, which increased to 100% with increasing pH of the synthesis and polymer mole fraction. Soret CD bands of all samples indicated loss of ?50% of band intensities, independent of the reaction pH. Catalytic activities of the conjugates increased with increasing synthesis pH, where 55-80% and 90-100% activity was retained for all samples synthesized at pH 5.0 and pH 7.0, respectively, and the Km or Vmax values of Cat-PAA(100)-7 did not differ significantly from those of the free enzyme. All conjugates synthesized at pH 7.0 were thermally stable even when heated to ?85-90 °C, while native catalase denatured between 55 and 65 °C. All conjugates retained 40-90% of their original activities even after storing for 10 weeks at 8 °C, while unmodified catalase lost all of its activity within 2 weeks, under similar storage conditions. Interestingly, PAA surrounding catalase limited access to the enzyme from large molecules like proteases and significantly increased resistance to trypsin digestion compared to unmodified catalase. Similarly, negatively charged PAA surrounding the catalase in these conjugates protected the enzyme against inhibition by negatively charged inhibitors such as ascorbate. While Cat-PAA(100)-7 did not show any inhibition by ascorbate in the presence of 270 ?M ascorbate, unmodified catalase lost ?70% of its activity under similar conditions. This simple, facile, and rational methodology produced thermostable, storable catalase that is also protected from protease digestion and ascorbate inhibition and most likely prevented the dissociation of the multimer. Using synthetic polymers to protect and improve enzyme properties could be an attractive approach for making "Stable-on-the-Table" enzymes, as a viable alternative to protein engineering. PMID:25046001

Riccardi, Caterina M; Cole, Kyle S; Benson, Kyle R; Ward, Jessamyn R; Bassett, Kayla M; Zhang, Yiren; Zore, Omkar V; Stromer, Bobbi; Kasi, Rajeswari M; Kumar, Challa V

2014-08-20

372

Intestinal bile acid sensing is linked to key endocrine and metabolic signalng pathways  

Technology Transfer Automated Retrieval System (TEKTRAN)

Bile acids have historically been considered to mainly function in cholesterol homeostasis and facilitate fat digestion in the gastrointestinal tract. Recent discoveries show that bile acids also function as signaling molecules that exert diverse endocrine and metabolic actions by activating G prote...

373

Regulation of the expression of key genes involved in HDL metabolism by unsaturated fatty acids  

Technology Transfer Automated Retrieval System (TEKTRAN)

The aim of this study was to determine the effects, and possible mechanisms of action, of unsaturated fatty acids on the expression of genes involved in HDL metabolism in HepG2 cells. The mRNA concentration of target genes was assessed by real time PCR. Protein concentrations were determined by wes...

374

Proline induces heat tolerance in chickpea (Cicer arietinum L.) plants by protecting vital enzymes of carbon and antioxidative metabolism.  

PubMed

Chickpea is a heat sensitive crop hence its potential yield is considerably reduced under high temperatures exceeding 35 °C. In the present study, we evaluated the efficacy of proline in countering the damage caused by heat stress to growth and to enzymes of carbon and antioxidative metabolism in chickpea. The chickpea seeds were raised without (control) and with proline (10 ?M) at temperatures of 30/25 °C, 35/30 °C, 40/35 °C and 45/40 °C as day/ night (12 h/12 h) in a growth chamber. The shoot and root length at 40/35 °C decreased by 46 and 37 %, respectively over control while at 45/40 °C, a decrease of 63 and 47 %, respectively over control was observed. In the plants growing in the presence of 10 ?M proline at 40/35 °C and 45/40 °C, the shoot length showed improvement of 32 and 53 %, respectively over untreated plants, while the root growth was improved by 22 and 26 %, respectively. The stress injury (as membrane damage) increased with elevation of temperatures while cellular respiration, chlorophyll content and relative leaf water content reduced as the temperature increased to 45/40 °C. The endogenous proline was elevated to 46 ?mol g(-1) dw at 40/35 °C but declined to 19 ?mol g(-1) dw in plants growing at 45/40 °C that was associated with considerable inhibition of growth at this temperature. The oxidative damage measured as malondialdehyde and hydrogen peroxide content increased manifolds in heat stressed plants coupled with inhibition in the activities of enzymatic (superoxide dismutase, catalase, ascorbate peroxidase, glutathione reductase) and levels of non-enzymatic (ascorbic acid, glutathione, proline) antioxidants. The enzymes associated with carbon fixation (RUBISCO), sucrose synthesis (sucrose phosphate synthase) and sucrose hydrolysis (invertase) were strongly inhibited at 45/40 °C. The plants growing in the presence of proline accumulated proline up to 63 ?mol g(-1) dw and showed less injury to membranes, had improved content of chlorophyll and water, especially at 45/40 °C. Additionally, the oxidative injury was significantly reduced coupled with elevated levels of enzymatic and non-enzymatic antioxidants. A significant improvement was also noticed in the activities of enzymes of carbon metabolism in proline-treated plants. We report here that proline imparts partial heat tolerance to chickpea's growth by reducing the cellular injury and protection of some vital enzymes related to carbon and oxidative metabolism and exogenous application of proline appears to have a countering effect against elevated high temperatures on chickpea. PMID:23573011

Kaushal, Neeru; Gupta, Kriti; Bhandhari, Kalpna; Kumar, Sanjeev; Thakur, Prince; Nayyar, Harsh

2011-07-01

375

Enzymes of glucose and methanol metabolism in the actinomycete Amycolatopsis methanolica.  

PubMed Central

The actinomycete Amycolatopsis methanolica was found to employ the normal bacterial set of glycolytic and pentose phosphate pathway enzymes, except for the presence of a PPi-dependent phosphofructokinase (PPi-PFK) and a 3-phosphoglycerate mutase that is stimulated by 2,3-bisphosphoglycerate. Screening of a number of actinomycetes revealed PPi-PFK activity only in members of the family Pseudonocardiaceae. The A. methanolica PPi-PFK and 3-phosphoglycerate mutase enzymes were purified to homogeneity. PPi-PFK appeared to be insensitive to the typical effectors of ATP-dependent PFK enzymes. Nevertheless, strong N-terminal amino acid sequence homology was found with ATP-PFK enzymes from other bacteria. The A. methanolica pyruvate kinase was purified over 250-fold and characterized as an allosteric enzyme, sensitive to inhibition by P(i) and ATP but stimulated by AMP. By using mutants, evidence was obtained for the presence of transketolase isoenzymes functioning in the pentose phosphate pathway and ribulose monophosphate cycle during growth on glucose and methanol, respectively. PMID:7961441

Alves, A M; Euverink, G J; Hektor, H J; Hessels, G I; van der Vlag, J; Vrijbloed, J W; Hondmann, D; Visser, J; Dijkhuizen, L

1994-01-01

376

Enzyme chemistry and the evolution of metabolic diversity: the. beta. -ketoadipate pathway  

SciTech Connect

The two converging catechol and protocatechuate branches of the ..beta..-ketoadipate pathway in Pseudomonas putida have long been considered a paradigm of evolutionary divergence of specialized enzymes from a common ancestor. The structural similarities of substrates, products and the enzymes themselves have supported this hypothesis. Employing chemical and /sup 1/H NMR techniques, they have determined the absolute stereochemical courses of the reactions catalyzed by ..beta..-carboxymuconate cycloisomerase, muconolactone isomerase, and ..gamma..-carboxymuconolactone decarboxylase. Surprisingly, ..beta..-carboxymuconate cycloisomerase proceeds via an anti addition while the corresponding muconate cycloisomerase has been shown to catalyze a syn addition. Moreover, the chiral centers generated in the products of both enzymes are of the opposite relative configuration. They believe that the shift in mechanism may reflect basic energetic differences of the two reactions. The stereochemistries of the isomerase and decarboxylase have been established by /sup 1/H NMR using a ricochet analysis. Both reactions proceed via a syn process; the relative configurations of muconolactone and ..gamma..-carboxymuconolactone necessitate that the enzymes operate on opposite faces of the common enol-lactone product. These findings suggest that either critical active site changes have occurred in these enzymes to accommodate preferred mechanistic pathways or the evolutionary relationship of the two branches is more remote than previously believed.

Kozarich, J.W.

1986-05-01

377

Interleukin-1 Receptor-Associated Kinase-3 Is a Key Inhibitor of Inflammation in Obesity and Metabolic Syndrome  

PubMed Central

Background Visceral obesity is associated with the rising incidence of type 2 diabetes and metabolic syndrome. Low-grade chronic inflammation and oxidative stress synergize in obesity and obesity-induced disorders. Objective We searched a cluster of molecules that support interactions between these stress conditions in monocytes. Methods RNA expressions in blood monocytes of two independent cohorts comprising 21 and 102 obese persons and 46 age-matched controls were determined by microarray and independently validated by quantitative RT-PCR analysis. The effect of three-month weight loss after bariatric surgery was determined. The effect of RNA silencing on inflammation and oxidative stress was studied in human monocytic THP-1 cells. Results Interleukin-1 receptor-associated kinase-3 (IRAK3), key inhibitor of IRAK/NF?B-mediated chronic inflammation, is downregulated in monocytes of obese persons. Low IRAK3 was associated with high superoxide dismutase-2 (SOD2), a marker of mitochondrial oxidative stress. A comparable expression profile was also detected in visceral adipose tissue of the same obese subjects. Low IRAK3 and high SOD2 was associated with a high prevalence of metabolic syndrome (odds ratio: 9.3; sensitivity: 91%; specificity: 77%). By comparison, the odds ratio of high-sensitivity C-reactive protein, a widely used marker of systemic inflammation, was 4.3 (sensitivity: 69%; specificity: 66%). Weight loss was associated with an increase in IRAK3 and a decrease in SOD2, in association with a lowering of systemic inflammation and a decreasing number of metabolic syndrome components. We identified the increase in reactive oxygen species in combination with obesity-associated low adiponectin and high glucose and interleukin-6 as cause of the decrease in IRAK3 in THP-1 cells in vitro. Conclusion IRAK3 is a key inhibitor of inflammation in association with obesity and metabolic syndrome. Our data warrant further evaluation of IRAK3 as a diagnostic and prognostic marker, and as a target for intervention. PMID:22272346

De Keyzer, Dieuwke; Mertens, Ann; Lannoo, Matthias; Vanaudenaerde, Bart; Hoylaerts, Marc; Benhabilčs, Nora; Tsatsanis, Christos; Mathieu, Chantal; Holvoet, Paul

2012-01-01

378

Sucrose non-fermenting related kinase enzyme is essential for cardiac metabolism  

PubMed Central

ABSTRACT In this study, we have identified a novel member of the AMPK family, namely Sucrose non-fermenting related kinase (Snrk), that is responsible for maintaining cardiac metabolism in mammals. SNRK is expressed in the heart, and brain, and in cell types such as endothelial cells, smooth muscle cells and cardiomyocytes (CMs). Snrk knockout (KO) mice display enlarged hearts, and die at postnatal day 0. Microarray analysis of embryonic day 17.5 Snrk hearts, and blood profile of neonates display defect in lipid metabolic pathways. SNRK knockdown CMs showed altered phospho-acetyl-coA carboxylase and phospho-AMPK levels similar to global and endothelial conditional KO mouse. Finally, adult cardiac conditional KO mouse displays severe cardiac functional defects and lethality. Our results suggest that Snrk is essential for maintaining cardiac metabolic homeostasis, and shows an autonomous role for SNRK during mammalian development. PMID:25505152

Cossette, Stephanie M.; Gastonguay, Adam J.; Bao, Xiaoping; Lerch-Gaggl, Alexandra; Zhong, Ling; Harmann, Leanne M.; Koceja, Christopher; Miao, Robert Q.; Vakeel, Padmanabhan; Chun, Changzoon; Li, Keguo; Foeckler, Jamie; Bordas, Michelle; Weiler, Hartmut; Strande, Jennifer; Palecek, Sean P.; Ramchandran, Ramani

2015-01-01

379

Sucrose non-fermenting related kinase enzyme is essential for cardiac metabolism.  

PubMed

In this study, we have identified a novel member of the AMPK family, namely Sucrose non-fermenting related kinase (Snrk), that is responsible for maintaining cardiac metabolism in mammals. SNRK is expressed in the heart, and brain, and in cell types such as endothelial cells, smooth muscle cells and cardiomyocytes (CMs). Snrk knockout (KO) mice display enlarged hearts, and die at postnatal day 0. Microarray analysis of embryonic day 17.5 Snrk hearts, and blood profile of neonates display defect in lipid metabolic pathways. SNRK knockdown CMs showed altered phospho-acetyl-coA carboxylase and phospho-AMPK levels sim