Kegg Pathway: Citrate cycle (TCA cycle)

Am J Physiol Regul Integr Comp Physiol. 2008 Jul 16;
Green HJ, Bombardier EB, Burnett M, Iqbal S, D'Arsigny CL, O'Donnell DE, Ouyang J, Webb KA

The objective of this study was to determine if patients with chronic obstructive lung disease (COPD) display differences in organization of the metabolic pathways and segments involved in energy supply as compared to healthy control subjects. Metabolic pathway potential, based on the measurement of the maximal activity (Vmax) of representative enzymes, was assessed in tissue extracted from the vastus lateralis in 7 patients with COPD (age 67+/-4 years; FEV1/FVC= 44+/-3 %; mean+/-SE) and 9 healthy age-matched controls (CON)(age 68+/-2 years; FEV1/FVC= 75+/-2 %). Compared to CON, the COPD patients displayed lower (P<0.05) Vmax (mol.kg protein-1.h-1) for Citrate synthase (CS, 2.20+/-0.16 vs 3.19+/-0.5), cytochrome-c oxidase (COX, 21.2+/-2.0 vs 28.7+/-2.2) and 3-hydroxyacyl-CoA dehydrogenase (HADH, 2.54+/-0.14 vs 3.74+/-0.12) but not Citrate synthase (CS, 2.20+/-0.16 vs 3.19+/-0.5). While no differences between groups were observed in Vmax for creatine phosphokinase (CPK), phosphorylase (PHOSPH), phosphofructokinase (PFK), pyruvate kinase (PK) and lactate dehydrogenase (LDH), hexokinase (HEX) was elevated in COPD (P0.05). Enzyme activity ratios were higher (P0.05) for HEX/CS, HEX/COX, PHOSPH/HADH and PFK/HADH in COPD compared to CON. It is concluded that COPD patients exhibit a reduced potential for both the electron transport system and fat oxidation and an increased potential for glucose phosphorylation while the potential for glycogenolysis and glycolysis remains normal. A comparison of enzyme ratios indicated greater potentials for glucose phosphorylation relative to the citric acid cycle and the electron transport chain, glycolysis relative to the electron transport chain, and glycogenolysis and glycolysis relative to beta-oxidation. Key words: Lung disease, skeletal muscle, enzymes, oxidative, glycolytic.

Rapid Commun Mass Spectrom. 2008 Jul 15; 22(16): 2436-2446
Law WS, Huang PY, Ong ES, Ong CN, Li SF, Pasikanti KK, Chan EC

A method using gas chromatography/mass spectrometry (GC/MS), liquid chromatography/mass spectrometry (LC/MS) and (1)H NMR with pattern recognition tools such as principle components analysis (PCA) was used to study the human urinary metabolic profiles after the intake of green tea. From the normalized peak areas obtained from GC/MS and LC/MS and peak heights from (1)H NMR, statistical analyses were used in the identification of potential biomarkers. Metabolic profiling by GC/MS provided a different set of quantitative signatures of metabolites that can be used to characterize the molecular changes in human urine samples. A comparison of normalized metabonomics data for selected metabolites in human urine samples in the presence of potential overlapping peaks after tea ingestion from LC/MS and (1)H NMR showed the reliability of the current approach and method of normalization. The close agreements of LC/MS with (1)H NMR data showed that the effects of ion suppression in LC/MS for early eluting metabolites were not significant. Concurrently, the specificity of detecting the stated metabolites by (1)H NMR and LC/MS was demonstrated. Our data showed that a number of metabolites involved in glucose metabolism, citric acid cycle and amino acid metabolism were affected immediately after the intake of green tea. The proposed approach provided a more comprehensive picture of the metabolic changes after intake of green tea in human urine. The multiple analytical approach together with pattern recognition tools is a useful platform to study metabolic profiles after ingestion of botanicals and medicinal plants. Copyright (c) 2008 John Wiley & Sons, Ltd.

Biotechnol Bioeng. 1995 May 20; 46(4): 325-32
Rane KD, Sims KA

The effect of dissolved oxygen on citric acid production and oxygen uptake by Candida lipolytica Y 1095 was evaluated in cell recycle and fed-batch fermentation systems. The maximum observed volumetric productivity, which occurred at a dilution rate of 0.06 h(-1), a dissolved oxygen concentration of 80%, and a biomass concentration of 5% w/v, in the cell recycle system, was 1.32 g citric acid/L . h. At these same conditions, the citric acid yield was 0.65 g/g and the specific citric acid productivity was 24.9 mg citric acid/g cell . h. In the cell recycle system, citric acid yields ranged from 0.45 to 0.72 g/g. Both the volumetric and specific citric acid productivities were dependent on the dilution rate and the concentration of dissolved oxygen in the fermentor. Similar productivities (1.29 g citric acid/L . h) were obtained in the fed-batch system operated at a cycle time of 36 h, a dissolved oxygen concentration of 80%, and 60 g total biomass. Citric acid yields in the fed-batch fermentor were consistently lower than those obtained in the cell recycle system and ranged from 0.40 to 0.59 g/g. Although citric acid yields in the fed-batch fermentor were lower than those obtained in the cell recycle system, higher citric:isocitric acid ratios were obtained in the fed-batch fermentor. As in the cell recycle system, both the volumetric and specific citric acid productivities in the fed-batch fermentor were dependent on the cycle time and dissolved oxygen concentration. (c) 1995 John Wiley & Sons, Inc.

Biosystems. 2008 Jun 20;
Egri-Nagy A, Nehaniv CL, Rhodes JL, Schilstra MJ

We propose a modeling and analysis method for biochemical reactions based on finite state automata. This is a completely different approach compared to traditional modeling of reactions by differential equations. Our method aims to explore the algebraic structure behind chemical reactions using automatically generated coordinate systems. In this paper we briefly summarize the underlying mathematical theory (the algebraic hierarchical decomposition theory of finite state automata) and describe how such automata can be derived from the description of chemical reaction networks. We also outline techniques for the flexible manipulation of existing models. As a real-world example we use the Krebs citric acid cycle.

J Mol Cell Cardiol. 2008 May 27;
Khairallah RJ, Khairallah M, Gélinas R, Bouchard B, Young ME, Allen BG, Lopaschuk GD, Deschepper CF, Des Rosiers C

While the balance between carbohydrates and fatty acids for energy production appears to be crucial for cardiac homeostasis, much remains to be learned about the molecular mechanisms underlying this relationship. Given the reported benefits of cGMP signaling on the myocardium, we investigated the impact of its chronic activation on cardiac energy metabolism using mice overexpressing a constitutively active cytoplasmic guanylate cyclase (GC(+/0)) in cardiomyocytes. Ex vivo working GC(+/0) heart perfusions with (13)C-labeled substrates revealed an altered pattern of exogenous substrate fuel selection compared to controls, namely a 38+/-9% lower contribution of exogenous fatty acids to acetyl-CoA formation, while that of carbohydrates remains unchanged despite a two-fold increase in glycolysis. The lower contribution of exogenous fatty acids to energy production is not associated with changes in energy demand or supply (contractile function, oxygen consumption, tissue acetyl-CoA or CoA levels, citric acid cycle flux rate) or in the regulation of beta-oxidation (acetyl-CoA carboxylase activity, tissue malonyl-CoA levels). However, GC(+/0) hearts show a two-fold increase in the incorporation of exogenous oleate into triglycerides. Furthermore, the following molecular data are consistent with a concomitant increase in triglyceride hydrolysis: (i) increased abundance of hormone sensitive lipase (HSL) protein (24+/-11%) and mRNA (22+/-4%) as well as (ii) several phosphorylation events related to HSL inhibitory (AMPK) and activation (ERK 1/2) sites, which should contribute to enhance its activity. These changes in exogenous fatty acid trafficking in GC(+/0) hearts appear to be functionally relevant, as demonstrated by their resistance to fasting-induced triglyceride accumulation. While the documented metabolic profile of GC(+/0) mouse hearts is partly reminiscent of hypertrophied hearts, the observed changes in lipid trafficking have not been previously documented, and may be part of the molecular mechanism underlying the benefits of cGMP signaling on the myocardium.

Biosci Rep. 2008 Jun 30;
Wang J, Wang X, Liu C, Zhang J, Zhu C, Guo X

A novel gene, named NgAOX1a, was isolated from Nicotiana glutinosa by reverse transcription-PCR (RT-PCR). The full-length cDNA of NgAOX1a was 1448 bp, including a 1062 bp open reading frame (ORF), a 124 bp 5' untranslated region (UTR) and a 262 bp 3' UTR. The ORF encodes a 353-amino acid protein which contains two conserved cysteine residues, four iron binding motifs, five helix regions and six conserved histidine residues. The phylogenetic tree showed that NgAOX1a belonged to AOX1-type. Alignment analysis showed that NgAOX1a shared high similarity with other known alternative oxidases. Four extrons and three introns were detected in the genomic DNA sequence, and Southern blotting analysis suggested that NgAOX1a is a single copy gene. A series of putative cis-acting elements were examined in the 5' flanking region of NgAOX1a. Northern blotting analysis showed that the transcript levels of NgAOX1a can be markedly accumulated when tobacco seedlings were treated with various abiotic stimuli, such as exogenous signaling molecules for plant defense response, salicylic acid (SA) and H2O2, and exogenous tricarboxylic acid (TCA) cycle metabolite Citrate. However, it could be suppressed by abiotic stress, such as CoCl2, an inhibitor of ethylene (ET), which indicated that the expression of NgAOX1a may be regulated by ET. In addition, NgAOX1a can also be strongly induced by three viral pathogens, Tobacco mosaic virus (TMV), Potato virus X (PVX) and Potato virus Y (PVY). Those results indicate that NgAOX1a may be involved in multi signal transduction pathways and may play an important role in defense response.

Int J Pharm. 2008 May 3;
Kanazawa T, Takashima Y, Hirayama S, Okada H

Human immunodeficiency virus (HIV) infections mainly occur through the vaginal and rectal mucosal membranes. In the present study, to develop a DNA vaginal vaccine against viral and bacterial infections, the effects of the menstrual cycle on DNA transfection through the vaginal mucosa in female mice and transfection enhancement by electroporation, a chelating agent, cell-penetrating peptides (CPP) and nuclear localizing signals (NLS) were investigated. The transfection efficiencies of a marker plasmid DNA (pDNA), pCMV-Luc, on the vaginal mucosal membrane in mice at the stages of metestrus and diestrus were significantly higher than those at the stages of proestrus and estrus. The gene expression was markedly enhanced by electroporation and by pretreatment with the chelating agent. The highest level of expression was obtained by 2h pretreatment with 5% citric acid solution combined with electroporation with 15 pulses at 250V/cm for 5milliseconds (ms). Furthermore, a synergistic promoting effect on pDNA transfection was obtained by co-administration of CPP, the Tat peptide analog, and NLS, the NF-kappaB analog. These results indicate that effective DNA vaccination administered through the vaginal tract is possible by selecting the menstrual stage and overcoming the mucosal barrier using a combination of methods that promotes uptake.

J Biol Chem. 2008 Jun 10;
Yang L, Kombu RS, Kasumov T, Zhu SH, Cendrowski AV, David F, Anderson VE, Kelleher JK, Brunengraber H

We conducted a study coupling metabolomics and mass isotopomer analysis of liver gluconeogenesis and citric acid cycle. Rat livers were perfused with lactate or pyruvate +/- aminooxyacetate or mercaptopicolinate, in the presence of 40% enriched NaH13CO3. Other livers were perfused with dimethyl-[1,4 -13C2]succinate +/- mercaptopicolinate. In this first of two companion reports, we show that a substantial fraction of gluconeogenic carbon leaves the liver as citric acid cycle intermediates, mostly a-ketoglutarate. The efflux of gluconeogenic carbon ranges from 10% to 200% of the rate of liver gluconeogenesis. This cataplerotic efflux of gluconeogenic carbon may contribute to renal gluconeogenesis in vivo. Multiple crossover analyses of concentrations of gluconeogenic intermediates, and redox measurements expand previous reports on the regulation of gluconeogenesis and the effects of inhibitors. We also demonstrate the formation of adducts from the condensation, in the liver, of (i) aminooxyacetate with pyruvate, a-ketoglutarate, and oxaloacetate, and (ii) mercaptopicolinate and pyruvate. These adducts may exert metabolic effects unrelated to their effect on gluconeogenesis.

J Biol Chem. 2008 Jun 10;
Yang L, Kasumov T, Kombu RS, Zhu SH, Cendrowski AV, David F, Anderson VE, Kelleher JK, Brunengraber H

In this second of two companion reports, we compare the mass isotopomer distribution of metabolites of liver gluconeogenesis and citric acid cycle labelled from NaH13CO3 or dimethyl-[1,4-13C2]succinate. The mass isoto-pomer distribution of intermediates reveals the reversibility of the isoCitrate dehydrogenase + aconitase reactions, even in the absence of a source of a-ketoglutarate. In addition, in many cases, a number of labeling incompatibilities were found such as (i) glucose vs triose phosphates and phosphoenolpyruvate, (ii) differences in the labeling ratios C4/C3 of glucose vs [glyceraldehyde-3-phosphate]/[dihydroxyacetone phosphate]and (iii) labeling of citric acid cycle intermediates in tissue vs effluent perfusate. Overall, our data show that gluconeogenic and citric acid cycle intermediates cannot be considered as sets of homogeneously labeled pools. This probably results from the zonation of hepatic metabolism and, in some cases, from differences in the labeling pattern of mitochondrial vs extra-mitochondrial metabolites. Our data have implications for the use of labeling patterns for the calculation of metabolic rates or fractional syntheses in liver, as well as for modeling liver intermediary metabolism.

Med Arh. 2008; 62(1): 45-8
Becić F, Kapić E, Rakanović-Todić M

Alpha lipoic acid is important intramolecular redox system. It is coenzyme of piruvate dehydrogenase and ketoglutarate dehydrogenase. Alpha lipoic acid has enzymatic and cytoprotective effect. It has key role in citric acid cycle, as a coenzyme. Therapeutic efficacy of alpha lipoic acid in diabetic neuropathy is based on reaction with free radicals and lipophylic antioxydans properties. Clinical studies results showed efficacy and safety of alpha liponic acid application in patients with diabetic neuropathy.

Extremophiles. 2008 Jun 5;
Voordeckers JW, Do MH, Hügler M, Ko V, Sievert SM, Vetriani C

The bacterial and archaeal communities of three deep-sea hydrothermal vent systems located on the Mid-Atlantic Ridge (MAR; Rainbow, Logatchev and Broken Spur) were investigated using an integrated culture-dependent and independent approach. Comparative molecular phylogenetic analyses, using the 16S rRNA gene and the deduced amino acid sequences of the alpha and beta subunits of the ATP Citrate lyase encoding genes were carried out on natural microbial communities, on an enrichment culture obtained from the Broken Spur chimney, and on novel chemolithoautotrophic bacteria and reference strains originally isolated from several different deep-sea vents. Our data showed that the three MAR hydrothermal vent chimneys investigated in this study host very different microbial assemblages. The microbial community of the Rainbow chimney was dominated by thermophilic, autotrophic, hydrogen-oxidizing, sulfur- and nitrate-reducing Epsilonproteobacteria related to the genus Caminibacter. The detection of sequences related to sulfur-reducing bacteria and archaea (Archaeoglobus) indicated that thermophilic sulfate reduction might also be occurring at this site. The Logatchev bacterial community included several sequences related to mesophilic sulfur-oxidizing bacteria, while the archaeal component of this chimney was dominated by sequences related to the ANME-2 lineage, suggesting that anaerobic oxidation of methane may be occurring at this site. Comparative analyses of the ATP Citrate lyase encoding genes from natural microbial communities suggested that Epsilonproteobacteria were the dominant primary producers using the reverse TCA cycle (rTCA) at Rainbow, while Aquificales of the genera Desulfurobacterium and Persephonella were prevalent in the Broken Spur chimney.

Am J Physiol Endocrinol Metab. 2008 Jun 3;
Hyyti OM, Olson A, Ge M, Ning XH, Buroker NE, Chung Y, Jue T, Portman MA

Dominant negative thyroid hormone receptors (TR) show elevated expression relative to ligand binding TRs during cardiac hypertrophy. We tested the hypothesis that over-expression of a dominant negative TR alters cardiac metabolism and contractile efficiency (CE). We used mice expressing the cardioselective dominant negative TR beta1 mutation, Delta337T. Isolated working Delta337T hearts and nontransgenic controls (CON) were perfused with (13)carbon labeled free fatty acids (FFA), acetoacetate (ACAC), lactate, and glucose at physiological concentrations for 30 minutes. (13)C NMR spectroscopy and isotopomer analyses were used to determine substrate flux and fractional contributions (Fc) of acetyl-CoA to the citric acid cycle (CAC). Delta337T hearts exhibited rate depression, but higher developed pressure and CE, defined as work per oxygen consumption (MVO2). Unlabeled substrate Fc from endogenous sources was higher in Delta337T, but Fc -ACAC was lower. Fluxes through CAC, lactate, ACAC, and FFA were reduced in Delta337T. CE and Fc differences were reversed by pacing Delta337T to CON rates, accompanied with an increase in FFA Fc. Delta337T hearts lacked the ability to increase MVO2. Decreases in protein expression for glucose transporter-4 and hexokinase-2 and increases in puryvate dehydrogenase kinase-2 and -4 suggest that these hearts are unable to increase carbohydrate oxidation in response to stress. These data show that Delta337T alters the metabolic phenotype in murine heart by reducing substrate flux for multiple pathways. Some of these changes are heart rate dependent, indicating that the substrate shift may represent an accommodation to altered contractile protein kinetics, which can be disrupted by pacing stress. Key words: thyroid hormone, glucose metabolism, free fatty acids.

Proc Natl Acad Sci U S A. 2008 Jun 3; 105(22): 7851-6
Huber H, Gallenberger M, Jahn U, Eylert E, Berg IA, Kockelkorn D, Eisenreich W, Fuchs G

Ignicoccus hospitalis is an anaerobic, autotrophic, hyperthermophilic Archaeum that serves as a host for the symbiotic/parasitic Archaeum Nanoarchaeum equitans. It uses a yet unsolved autotrophic CO(2) fixation pathway that starts from acetyl-CoA (CoA), which is reductively carboxylated to pyruvate. Pyruvate is converted to phosphoenol-pyruvate (PEP), from which glucogenesis as well as oxaloacetate formation branch off. Here, we present the complete metabolic cycle by which the primary CO(2) acceptor molecule acetyl-CoA is regenerated. Oxaloacetate is reduced to succinyl-CoA by an incomplete reductive citric acid cycle lacking 2-oxoglutarate dehydrogenase or synthase. Succinyl-CoA is reduced to 4-hydroxybutyrate, which is then activated to the CoA thioester. By using the radical enzyme 4-hydroxybutyryl-CoA dehydratase, 4-hydroxybutyryl-CoA is dehydrated to crotonyl-CoA. Finally, beta-oxidation of crotonyl-CoA leads to two molecules of acetyl-CoA. Thus, the cyclic pathway forms an extra molecule of acetyl-CoA, with pyruvate synthase and PEP carboxylase as the carboxylating enzymes. The proposal is based on in vitro transformation of 4-hydroxybutyrate, detection of all enzyme activities, and in vivo-labeling experiments using [1-(14)C]4-hydroxybutyrate, [1,4-(13)C(2)], [U-(13)C(4)]succinate, or [1-(13)C]pyruvate as tracers. The pathway is termed the dicarboxylate/4-hydroxybutyrate cycle. It combines anaerobic metabolic modules to a straightforward and efficient CO(2) fixation mechanism.

Int Angiol. 2008 Jun; 27(3): 253-9
Riccioni C, Sarcinella R, Palermo G, Izzo A, Liguori M, Koverech A, Messano M, Virmani A

AIM: The effectiveness of propionyl-L-carnitine (PLC) monotherapy regimen alone or in association with pulsed muscular compression was compared to the physical therapy by itself against obliterant arteriopathy Leriche Fontaine stage II. PLC is involved in cellular metabolism and is transformed into two active substances, free L-carnitine and propionyl-coenzyme A in the mitochondria, which take part in fatty acid transfer and in the citric acid cycle, respectively. METHODS: Forty-two patients with arterial disease were selected (22 males and 20 females; mean age: 62+/-8 years; 21 type 2 diabetic [DB] and 21 non-DB [NDB]). At enrollment all patients completed a symptoms questionnaire enabling both clinical and social evaluation of the impact of the arteriopathy on the quality of life. Then, patients had: routine blood samples, echo duplex scan; evaluation of the ankle/arm (Winsor) index; impedance plethysmography (Rheoscreen) to measure the crest time (CT), index of the pathological changes due to the sclerosis on the vascular wall, and measurement of walking distance by means of treadmill test. Patients were randomized in three groups, each of them composed by 14 patients (7 DB and 7 NDB): the first group was submitted to infusional PLC therapy at a dosage of 4 fl (total: 1,200 mg PLC) in 250 cc of physiological solution for 5 days a week for 4 weeks; the second group was treated with PLC in association with pulsed muscular compression therapy by Vascupump (5 sessions a week for 4 weeks); the third group was submitted only to Vascupump. RESULTS: The efficacy of both PLC and Vascupump in the treatment of the peripheral vasculopathies was confirmed. From a subjective point of view, patients referred benefits both in clinical terms, i.e. increased walking distance (average increaseaegroup I: DB 102%, NDB 118%; group II: DB 94%, NDB 193%; group III: DB 33%, NDB 67%) and of decreased intensity of the calf pain from the quality of life questionnaire (21.5 to 10.7). The instrumental parameters showed a trend towards normality, i.e decrease in CT and an increase of the Winsor index, indicators of increased peripheral blood circulation. CONCLUSION: Combined pharmaco- and physical therapy was most efficient treatment regime and best results were seen in NDB compared to the DB patients.

J Bacteriol. 2008 Jul; 190(14): 4933-40
Mullins EA, Francois JA, Kappock TJ

Microbes tailor macromolecules and metabolism to overcome specific environmental challenges. Acetic acid bacteria perform the aerobic oxidation of ethanol to acetic acid and are generally resistant to high levels of these two membrane-permeable poisons. The citric acid cycle (CAC) is linked to acetic acid resistance in Acetobacter aceti by several observations, among them the oxidation of acetate to CO(2) by highly resistant acetic acid bacteria and the previously unexplained role of A. aceti Citrate synthase (AarA) in acetic acid resistance at a low pH. Here we assign specific biochemical roles to the other components of the A. aceti strain 1023 aarABC region. AarC is succinyl-coenzyme A (CoA):acetate CoA-transferase, which replaces succinyl-CoA synthetase in a variant CAC. This new bypass appears to reduce metabolic demand for free CoA, reliance upon nucleotide pools, and the likely effect of variable cytoplasmic pH upon CAC flux. The putative aarB gene is reassigned to SixA, a known activator of CAC flux. Carbon overflow pathways are triggered in many bacteria during metabolic limitation, which typically leads to the production and diffusive loss of acetate. Since acetate overflow is not feasible for A. aceti, a CO(2) loss strategy that allows acetic acid removal without substrate-level (de)phosphorylation may instead be employed. All three aar genes, therefore, support flux through a complete but unorthodox CAC that is needed to lower cytoplasmic acetate levels.

Mol Cell. 2008 May 23; 30(4): 393-402
Kaelin WG, Ratcliffe PJ

HIF plays a central role in the transcriptional response to changes in oxygen availability. The PHD family of oxygen-dependent prolyl hydroxylases plays a pivotal role in regulating HIF stability. The biochemical properties of these enzymes make them well suited to act as oxygen sensors. They also respond to other intracellular signals, including reactive oxygen species, nitric oxide, and certain metabolites, that can modulate the hypoxic response. HIF transcriptional activity is further tuned by FIH1-mediated asparagine hydroxylation. HIF affects signaling pathways that influence development, metabolism, inflammation, and integrative physiology. Accordingly, HIF-modulatory drugs are now being developed for diverse diseases.

Proc Natl Acad Sci U S A. 2008 May 13; 105(19): 7028-33
Khairallah M, Khairallah RJ, Young ME, Allen BG, Gillis MA, Danialou G, Deschepper CF, Petrof BJ, Des Rosiers C

We recently demonstrated early metabolic alterations in the dystrophin-deficient mdx heart that precede overt cardiomyopathy and may represent an early "subclinical" signature of a defective nitric oxide (NO)/cGMP pathway. In this study, we used genetic and pharmacological approaches to test the hypothesis that enhancing cGMP, downstream of NO formation, improves the contractile function, energy metabolism, and sarcolemmal integrity of the mdx heart. We first generated mdx mice overexpressing, in a cardiomyocyte-specific manner, guanylyl cyclase (GC) (mdx/GC(+/0)). When perfused ex vivo in the working mode, 12- and 20-week-old hearts maintained their contractile performance, as opposed to the severe deterioration observed in age-matched mdx hearts, which also displayed two to three times more lactate dehydrogenase release than mdx/GC(+/0). At the metabolic level, mdx/GC(+/0) displayed a pattern of substrate selection for energy production that was similar to that of their mdx counterparts, but levels of citric acid cycle intermediates were significantly higher (36 +/- 8%), suggesting improved mitochondrial function. Finally, the ability of dystrophin-deficient hearts to resist sarcolemmal damage induced in vivo by increasing the cardiac workload acutely with isoproterenol was enhanced by the presence of the transgene and even more so by inhibiting cGMP breakdown using the phosphodiesterase inhibitor sildenafil (44.4 +/- 1.0% reduction in cardiomyocyte damage). Overall, these findings demonstrate that enhancing cGMP signaling, specifically downstream and independent of NO formation, in the dystrophin-deficient heart improves contractile performance, myocardial metabolic status, and sarcolemmal integrity and thus constitutes a potential clinical avenue for the treatment of the dystrophin-related cardiomyopathies.

In Silico Biol. 2007; 7(6): 583-93
Kundu S, Roy D

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) of pathogenic protozoa Trichomonas vaginalis (TvGAPDH) is an attractive drug target since this parasite lacks functional citric acid cycle and is dependent solely on glycolysis for its energy requirements. The three dimensional structure of TvGAPDH dimer has been generated by homology modelling based on the crystal structure of human liver GAPDH. Comparison of the NAD;{+} binding pocket of the modeled TvGAPDH with human GAPDH (hGAPDH) reveals the presence of a hydrophobic pocket near the N-6 position of adenine ring as well as a hydrophobic cleft near O-2' of the adenosine ribose that are absent in the human enzyme. In order to exploit these structural differences adenosine and several adenosine analogs with substitution on N-6 position of adenine ring or 2' position of ribose sugar or both have been studied by docking experiments using the program AutoDock version 3.0.5. Our docking result suggests that bulkier hydrophobic substitution at the N-6 position of the adenine ring could form more stable complexes with TvGAPDH than with hGAPDH. An improvement of binding occurs in TvGAPDH when methoxybenzamido group has been introduced at the O-2' position of the ribose sugar. The combination of N-6 and O-2' substitutions may have produced significantly improved inhibitors. Our study may help in identifying structural elements involved in the origin of selectivity at the NAD;{+} binding pocket of TvGAPDH. This study could further be extended for future anti-trichomonal drug design strategies in order to control trichomoniasis.

J Bacteriol. 2008 Jul; 190(13): 4687-96
Sattley WM, Madigan MT, Swingley WD, Cheung PC, Clocksin KM, Conrad AL, Dejesa LC, Honchak BM, Jung DO, Karbach LE, Kurdoglu A, Lahiri S, Mastrian SD, Page LE, Taylor HL, Wang ZT, Raymond J, Chen M, Blankenship RE, Touchman JW

Despite the fact that heliobacteria are the only phototrophic representatives of the bacterial phylum Firmicutes, genomic analyses of these organisms have yet to be reported. Here we describe the complete sequence and analysis of the genome of Heliobacterium modesticaldum, a thermophilic species belonging to this unique group of phototrophs. The genome is a single 3.1-Mb circular chromosome containing 3,138 open reading frames. As suspected from physiological studies of heliobacteria that have failed to show photoautotrophic growth, genes encoding enzymes for known autotrophic pathways in other phototrophic organisms, including ribulose bisphosphate carboxylase (Calvin cycle), Citrate lyase (reverse citric acid cycle), and malyl coenzyme A lyase (3-hydroxypropionate pathway), are not present in the H. modesticaldum genome. Thus, heliobacteria appear to be the only known anaerobic anoxygenic phototrophs that are not capable of autotrophy. Although for some cellular activities, such as nitrogen fixation, there is a full complement of genes in H. modesticaldum, other processes, including carbon metabolism and endosporulation, are more genetically streamlined than they are in most other low-G+C gram-positive bacteria. Moreover, several genes encoding photosynthetic functions in phototrophic purple bacteria are not present in the heliobacteria. In contrast to the nutritional flexibility of many anoxygenic phototrophs, the complete genome sequence of H. modesticaldum reveals an organism with a notable degree of metabolic specialization and genomic reduction.

BMC Microbiol. 2008; 8: 62
van der Voort M, Kuipers OP, Buist G, de Vos WM, Abee T

BACKGROUND: The catabolite control protein CcpA is a transcriptional regulator conserved in many Gram-positives, controlling the efficiency of glucose metabolism. Here we studied the role of Bacillus cereus ATCC 14579 CcpA in regulation of metabolic pathways and expression of enterotoxin genes by comparative transcriptome analysis of the wild-type and a ccpA-deletion strain. RESULTS: Comparative analysis revealed the growth performance and glucose consumption rates to be lower in the B. cereus ATCC 14579 ccpA deletion strain than in the wild-type. In exponentially grown cells, the expression of glycolytic genes, including a non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase that mediates conversion of D-glyceraldehyde 3-phosphate to 3-phospho-D-glycerate in one single step, was down-regulated and expression of gluconeogenic genes and genes encoding the citric acid cycle was up-regulated in the B. cereus ccpA deletion strain. Furthermore, putative CRE-sites, that act as binding sites for CcpA, were identified to be present for these genes. These results indicate CcpA to be involved in the regulation of glucose metabolism, thereby optimizing the efficiency of glucose catabolism. Other genes of which the expression was affected by ccpA deletion and for which putative CRE-sites could be identified, included genes with an annotated function in the catabolism of ribose, histidine and possibly fucose/arabinose and aspartate. Notably, expression of the operons encoding non-hemolytic enterotoxin (Nhe) and hemolytic enterotoxin (Hbl) was affected by ccpA deletion, and putative CRE-sites were identified, which suggests catabolite repression of the enterotoxin operons to be CcpA-dependent. CONCLUSION: The catabolite control protein CcpA in B. cereus ATCC 14579 is involved in optimizing the catabolism of glucose with concomitant repression of gluconeogenesis and alternative metabolic pathways. Furthermore, the results point to metabolic control of enterotoxin gene expression and suggest that CcpA-mediated glucose sensing provides an additional mode of control in moderating the expression of the nhe and hbl operons in B. cereus ATCC 14579.