Glycolytic Reactions Research Articles

Structure preserving algorithms for mathematical model of auto-catalytic glycolysis chemical reaction and numerical simulations

This paper aims to develop positivity preserving splitting techniques for glycolysis reaction–diffusion chemical model. The positivity of state variables in the glycolysis model is an essential property that must be preserved for all choices of parameters. We propose two splitting methods that remain dynamically consistent with the continuous glycolysis reaction–diffusion model. The proposed methods converge to a true steady-state or fixed point under the given condition. On contrary to the classical operator splitting finite difference methods, we use nonstandard finite difference theory to propose a new class of operator splitting techniques.

13 C Metabolic Flux Analysis of Escherichia coli Engineered for Gamma-Aminobutyrate Production.

Escherichia coli is engineered for γ-aminobutyrate (GABA) production in glucose minimal medium. For this, overexpression of mutant glutamate decarboxylase (GadB) and mutant glutamate/GABA antiporter (GadC), as well as deletion of GABA transaminase (GabT), are accomplished. In addition, the carbon flux to the tricarboxylic acid cycle is engineered by the overexpression of gltA, ppc, or both. The overexpression of citrate synthase (CS), encoded by gltA, increases GABA productivity, as expected. Meanwhile, the overexpression of phosphoenolpyruvate carboxylase (PPC) causes a decrease in the rate of glucose uptake, resulting in a decrease in GABA production. The phenotypes of the strains are characterized by 13 C metabolic flux analysis (13 C MFA). The results reveal that CS overexpression increases glycolysis and anaplerotic reaction rates, as well as the citrate synthesis rate, while PPC overexpression causes little changes in metabolic fluxes, but reduces glucose uptake rate. The engineered strain produces 1.2gL-1 of GABA from glucose. Thus, by using 13 C MFA, important information is obtained for designing metabolically engineered strains for efficient GABA production.

Standard Gibbs energy of metabolic reactions: VI. Glyceraldehyde 3-phosphate dehydrogenase reaction

Glycolysis is a very central metabolic pathway for many organisms because it represents a key component in their energy production. For this reason, it has always been an extensively studied pathway. The glyceraldehyde 3-phosphate dehydrogenase (GDH) reaction is an important reaction of glycolysis yielding nicotinamide adenine dinucleotide (NADH). The aim of this work is to investigate the thermodynamics of the GDH reaction and determine the standard Gibbs energy of reaction ΔRg'0 and standard enthalpy of reaction ΔRh'0. Currently, so-called ‘standard’ data exist in the literature that depend on the conditions they were measured at. In this work, ΔRg'0 and ΔRh'0 values were determined that are independent from reaction conditions by accounting for the activity coefficients of the reacting substances. Therefore, the equation of state electrolyte Perturbed-Chain Statistical Associating Fluid Theory (ePC-SAFT) was used. The required ePC-SAFT parameters were taken from literature or fitted to new experimental osmotic coefficients. A value of ΔRg'0 = 51.5 ± 0.4 kJ mol−1 was determined at 298.15 K. This value deviates by up to 10 kJ mol−1 from existing literature values, caused by activity coefficients in the reaction medium. It can be used to determine the Gibbs energy of reaction ΔRg', which will allow statements concerning the feasibility of the GDH reaction. Further, a method is presented to predict influences of pH, initial substrate concentration and Mg2+ concentration on the reaction equilibrium. Finally, we measured the standard reaction enthalpy for the GDH reaction ΔRh'0 by titration calorimetric measurements (ΔRh'0 = 4.6 ± 0.1 kJ mol−1). This value was within van ’t Hoff evaluated ΔRh'0 (9 ± 16 kJ mol−1) using temperature-dependent equilibrium constants from equilibrium measurements corrected by ePC-SAFT predicted activity coefficients.

Kruppel-like factor 2 disturb non-small cell lung cancer energy metabolism by inhibited glutamine consumption.

Metabolic reprogramming is well accepted as a hallmark of cancer. This study aimed to explore the role of Kruppel-like factor 2 (KLF2) in aerobic glycolysis and glutamine consumption of energy metabolism in non-small cell lung cancer (NSCLC) cells. Two different NSCLC cells, A549 and NCI-H1299, were used to investigate the role of KLF2 in glycolysis and glutamine consumption by tracer technique and KLF2 transfection. The results showed that overexpression KLF could inhibit the energy metabolism and proliferation of NSCLC cells, but had no significant effect on glycolysis reaction and only affected the glutamine consumption of NSCLC cells. In NSCLC cells exposed to glutamine deprivation, the effect of overexpression of KLF2 on cell proliferation and energy metabolism disappeared. It was found that KLF2 could inhibit the expression of glutaminase (GLS) by metabolite tracing technique and so on. However, when GLS inhibitors were given to overexpressing KLF2 NSCLC cells, the intervention effect of KLF2 disappeared. Kruppel-like factor 2 could decrease the level of glutamine, participate in the consumption of glutamine by cancer cells, and then inhibit the energy metabolism of cancer cells.

Phosphofructokinase 1 Platelet Isoform Promotes β-Catenin Transactivation for Tumor Development.

Metabolism plays a critical role in direct regulation of a variety of cellular activities via metabolic enzymes and metabolites. Here, we demonstrate that phosphofructokinase 1 platelet isoform (PFKP), which catalyzes a rate-limiting reaction in glycolysis, promotes EGFR activation-induced nuclear translocation and activation of β-catenin, thereby enhancing the expression of its downstream genes CCND1 and MYC in human glioblastoma cells. Importantly, we showed that EGFR-phosphorylated PFKP Y64 has a critical role in AKT activation and AKT-mediated β-catenin S552 phosphorylation and subsequent β-catenin transactivation and promotion of tumor cell glycolysis, migration, invasion, proliferation, and brain tumor growth. These findings highlight a novel mechanism underlying a glycolytic enzyme-mediated β-catenin transactivation and underscore the integrated and reciprocal regulation of metabolism and gene expression, which are two fundamental biological processes in tumor development.

One-Year Old Dormant, "Non-culturable" Mycobacterium tuberculosis Preserves Significantly Diverse Protein Profile.

For adaptation to stressful conditions, Mycobacterium tuberculosis (Mtb) is prone to transit to a dormant, non-replicative state, which is believed to be the basis of the latent form of tuberculosis infection. Dormant bacteria persist in the host for a long period without multiplication, cannot be detected from biological samples by microbiological methods, however, their “non-culturable” state is reversible. Mechanisms supporting very long capacity of mycobacteria for resuscitation and further multiplication after prolonged survival in a dormant phase remain unclear. Using methods of 2D electrophoresis and MALDI-TOF analysis, in this study we characterized changes in the proteomic profile of Mtb stored for more than a year as dormant, non-replicating cells with a negligible metabolic activity, full resistance to antibiotics, and altered morphology (ovoid forms). Despite some protein degradation, the proteome of 1-year-old dormant mycobacteria retained numerous intact proteins. Their protein profile differed profoundly from that of metabolically active cells, but was similar to the proteome of the 4-month-old dormant bacteria. Such protein stability is likely to be due to the presence of a significant number of enzymes involved in the protection from oxidative stress (katG/Rv1908, sodA/Rv3846, sodC/Rv0432, bpoC/Rv0554), as well as chaperones (dnaJ1/Rv0352, htpG/Rv2299, groEL2/Rv0440, dnaK/Rv0350, groES/Rv3418, groEL1/Rv3417, HtpG/Rv2299c, hspX/Rv2031), and DNA-stabilizing proteins. In addition, dormant cells proteome contains enzymes involved in specific metabolic pathways (glycolytic reactions, shortened TCA cycle, degradative processes) potentially providing a low-level metabolism, or these proteins could be “frozen” for usage in the reactivation process before biosynthetic processes start. The observed stability of proteins in a dormant state could be a basis for the long-term preservation of Mtb cell vitality and hence for latent tuberculosis.

Biomimetic systems involving sequential redox reactions in glycolysis - the sulfur effect.

Magnesium hemithioacetates were used as model cysteine compounds to mimic natural hemithioacetals, and their biomimetic oxidation reactions using a model NAD+ compound were investigated. Cyclic hemithioacetate was found to be the best substrate for the reaction with the model NAD+ compound, which gave the corresponding NADH analog in excellent yield.

Вплив Дерозалу на обмін азотистих та фосфорних сполук у печінці і м’язовій тканині курей за умов хронічного експерименту

Pesticides are toxic chemicals which are widely used as an effective means against pests and plant diseases as well as for animals’ protection against ectoparasites. They are also used to control rodents which transmit infectious diseases to people and animals (encephalitis, malaria, leptospirosis, rabies, listeriosis, etc.). Biosynthesis of protein in tissues depends on many circumstances and, first of all, on the complete set of amino acids, the state of enzymes’ system and on the presence of stimulators and inhibitors of synthesis reaction. Xenobiotics including benzimidazole derivatives are a part of these compositions. In Ukraine there are more than 20 medicines based on carbendazim. Thus, the analysis of the its influence on protein metabolism is relevant. In the research the hens breed Rhode Island of the 38th line for egg and meat of 30 days of age, with the mass 1000-1200 gr. was used. They were divided into 2 groups - test (n=36) and 1 control (n=18). Нens were equally cared and fed with two food allowances: the first part up to 40 days of age and the second part from 40 days of age up to the end of the experiment (60 days). Every day throughout 60 days hens of the first and second testing groups received feed with the medicine Derosal (BAYER, Germany) in the form of 50 % aqueous suspension in dose of 90 and 900 mg. per 1 kg of body mass (0.01 and 0.1 LD50 for hens) accordingly. The control group of hens was fed without addition of the medicine. The research was carried out on the 30th and 60th day. During the experiment it was found that addition of carbendazim to fodder slowed down the use of its nutrients. This situation was characterized by the fact that on the 30th and 60th day of the research the decrease of body mass gaining up to 10 % occurred, the decrease of protein and non-protein nitrogen content in liver and muscles in all the hens’ groups occurred. The increase of concentration of DNA and RNA phosphorus, the content of acid-soluble phosphate, phosphorus of phospholipid as well as lipoid phosphorus that indicates the slight deceleration of protein synthesis were determined, DNA and RNA in liver and hens’ muscles, also indicated the stimulation of synthesis of phosphorus acid ether and the reaction of glycolysis under the conditions of pesticide adding.&nbsp

Spatial pattern formation in reaction-diffusion models: a computational approach.

Reaction-diffusion equations have been widely used to describe biological pattern formation. Nonuniform steady states of reaction-diffusion models correspond to stationary spatial patterns supported by these models. Frequently these steady states are not unique and correspond to various spatial patterns observed in biology. Traditionally, time-marching methods or steady state solvers based on Newton's method were used to compute such solutions. However, the solutions that these methods converge to highly depend on the initial conditions or guesses. In this paper, we present a systematic method to compute multiple nonuniform steady states for reaction-diffusion models and determine their dependence on model parameters. The method is based on homotopy continuation techniques and involves mesh refinement, which significantly reduces computational cost. The method generates one-parameter steady state bifurcation diagrams that may contain multiple unconnected components, as well as two-parameter solution maps that divide the parameter space into different regions according to the number of steady states. We applied the method to two classic reaction-diffusion models and compared our results with available theoretical analysis in the literature. The first is the Schnakenberg model which has been used to describe biological pattern formation due to diffusion-driven instability. The second is the Gray-Scott model which was proposed in the 1980s to describe autocatalytic glycolysis reactions. In each case, the method uncovers many, if not all, nonuniform steady states and their stabilities.

The usage of novel acrylic-modified water-reducible alkyd resin obtained from post-consumer PET bottles in water-based paint formulation

Glycolysis reactions of waste polyethylene terephthalate (PET) flakes obtained from post-consumer water bottles were carried out at 220–250 °C using diethylene glycol, dipropylene glycol and triethylene glycol, and molar ratios of PET/glycol in the glycolysis reactions were chosen to be 1/3. The obtained glycolysis products were used in the synthesis of medium-oil acrylic-modified water-reducible alkyd resins. Acrylic modification was carried out using methacrylic acid–maleic acid copolymer synthesized in our laboratory. The structure of the acrylic copolymer was investigated with FTIR analysis. Films of the modified alkyd resins were prepared and their physical and chemical surface coating properties were investigated. When surface coating test results were evaluated, it was observed that waste PET had no adverse effect on these properties of acrylic-modified water-reducible alkyd resins. Then, two water-based paints were prepared using PET-based acrylic-modified water-reducible alkyd resin and reference acrylic-modified water-reducible alkyd resin as binder. Wet paint properties and physical/chemical dry film properties of paints were determined. When the physical surface coating test results were evaluated, no difference was observed between the properties of both paints. If we evaluate the results of chemical surface coating properties, the hot water and alkali resistance of waste PET-based paint was considerably higher than the reference resin.

Chemical Recycling of Polyethlylene Terephthalate by Glycolysis Using Deep Eutectic Solvents

In this study, the glycolysis of polyethylene terephthalate was studied in presence of deep eutectic solvents as catalyst. In the glycolysis of PET, five different deep eutectic solvents were synthesized using different hydrogen bond donors and acceptors. Among the synthesized DESs, the most efficient catalyst was found to be DES formed by potassium carbonate and ethylene glycol. Glycolysis reaction was performed between PET and ethylene glycol. The effects of process parameters such as temperature, ratio of EG/PET and ratio of DES/PET were examined. The maximum yield for monomer product, bis(2-hydroxyethyl) terephthalate was observed as 88% at the reaction temperature of 180 °C, ethylene glycol/deep eutectic solvent ratio of 15 and deep eutectic solvent/polyethylene terephthalate ratio of 6.

Low-Cost Catalyst for Glycolysis of Polyethylene Terephthalate (PET)

A low-cost process for the depolymerization of polyethylene terephthalate (PET) was investigated in this work by the development of catalysts derived from food wastes for the glycolysis reaction of post-consumable waste of drinking bottles. Bis (2-hydroxyethyl) terephthalate (BHET) is obtained as a product from the glycolysis of polyethylene terephthalate (PET). Calcium oxide (CaO) catalysts derived from shells were used in this reaction. The yield of bis (2-hydroxyethyl) terephthalate (BHET) was obtained and the purity of BHET was confirmed by NMR spectroscopy.

Global Metabolic Shifts in Age and Alzheimer's Disease Mouse Brains Pivot at NAD+/NADH Redox Sites.

Age and Alzheimer’s disease (AD) share some common features such as cognitive impairments, memory loss, metabolic disturbances, bioenergetic deficits, and inflammation. Yet little is known on how systematic shifts in metabolic networks depend on age and AD. In this work, we investigated the global metabolomic alterations in non-transgenic (NTg) and triple-transgenic (3xTg-AD) mouse brain hippocampus as a function of age by using untargeted Ultrahigh Performance Liquid Chromatography-tandem Mass Spectroscopy (UPLC-MS/MS). We observed common metabolic patterns with aging in both NTg and 3xTg-AD brains involved in energy-generating pathways, fatty acids oxidation, glutamate, and sphingolipid metabolism. We found age-related downregulation of metabolites from reactions in glycolysis that consumed ATP and in the TCA cycle, especially at NAD+/NADH-dependent redox sites, where age- and AD-associated limitations in the free NADH may alter reactions. Conversely, metabolites increased in glycolytic reactions in which ATP is produced. With age, inputs to the TCA cycle were increased including fatty acid β-oxidation and glutamine. Overall age- and AD-related changes were > 2-fold when comparing the declines of upstream metabolites of NAD+/NADH-dependent reactions to the increases of downstream metabolites (p = 10-5, n = 8 redox reactions). Inflammatory metabolites such as ceramides and sphingosine-1-phosphate also increased with age. Age-related decreases in glutamate, GABA, and sphingolipid were seen which worsened with AD genetic load in 3xTg-AD brains, possibly contributing to synaptic, learning- and memory-related deficits. The data support the novel hypothesis that age- and AD-associated metabolic shifts respond to NAD(P)+/NAD(P)H redox-dependent reactions, which may contribute to decreased energetic capacity.

Hexokinase 2 dimerization and interaction with voltage-dependent anion channel promoted resistance to cell apoptosis induced by gemcitabine in pancreatic cancer.

Gemcitabine (GEM) is the standard chemotherapy drug for pancreatic cancer. Because of widespread drug resistance, the effect is limited. Therefore, it is urgent to reveal the underlying mechanism. Glycolysis is the most remarkable character of tumor aberrant metabolism, which plays vital roles on tumor drug resistance. Hexokinase 2 (HK2), as the key enzyme regulating the first‐step reaction of glycolysis, is overexpressed in many kinds of tumors. The putative role of HK2 resisting GEM therapy was investigated in this study. We found that HK2 was overexpressed in pancreatic cancer and associated with poor prognosis. HK2 knockdown decreased pancreatic cancer cell proliferation, migration viability, and promoted cell apoptosis in vitro. HK2 high expression in pancreatic cancer showed GEM resistance. HK2 knockdown increased the sensitivity of pancreatic cancer cell to GEM, the growth of xenograft tumor with HK2 knockdown was also further decreased with the GEM treatment compared with control in vivo. GEM‐resistant pancreatic cancer showed the increase of HK2 dimer rather than HK2 mRNA or protein. Our study revealed that the ROS derived from GEM promoted HK2 dimerization combining with voltage‐dependent anion channel, which resulted in the resistance to GEM. Meanwhile, our study established a new sight for GEM resistance in pancreatic cancer.

Abstract 4376: Targeting glutamine metabolism disables Warburg physiology by inhibiting proximal glycolysis and Krebs cycle rewiring

Abstract Glutamine plays a critical role in multiple metabolic reactions that support tumor anabolic processes. In spite of this, targeted inhibition of glutaminase, which converts exogenous glutamine into glutamate has met with limited clinical success. We hypothesized that broadly inhibiting glutamine metabolism would more effectively shut down tumor growth. To this end we employed a novel prodrug of the glutamine antagonist 6-diazo-5-oxo-L-norleucine (DON) in a number of syngeneic mouse models of cancer including MC38 (derived from colon cancer), 3LL (derived from lung cancer) and B16 (derived from melanoma). To better understand the anti-tumor mechanisms, we used LC-MS-based metabolomics to profile metabolic flux in these different tumor models with [U-13C]-glucose/glutamine as tracer. We observed that when compared to 3LL and B16, MC38 was highly sensitive to anti-glutamine treatment. As expected, tumor growth inhibition was correlated with inhibition of (the glutamine requiring) purine nucleotide synthesis. Surprisingly however, treatment with the glutamine antagonist markedly inhibited “proximal” glycolytic reactions as determined by inhibition of the generation of glucose-6-P and fructose 1,6-bisphosphate. This inhibition correlated with a decrease in FDG-PET. Likewise, we observed a marked decrease in one carbon metabolism as measured by serine. Strikingly, glutamine antagonism eliminated the glucose-derived succinate. Instead, a dramatic rewiring of the Krebs cycle was identified, which showed an alternative source of succinate was derived from the GABA shunt. In contrast, not only were these pathways upregulated in the relatively resistant B16 and 3LL tumors, glutamine antagonism only minimally affected these pathways. Furthermore, glutamine antagonism led to a marked decrease in kynurenine. Kynurenine is the result of tryptophan metabolism by IDO and is a potent immunosuppressive metabolite and promoter of metastasis. Decreased kynurenine levels were not the result of inhibition of IDO activity by the glutamine antagonist but rather due to a decrease in IDO expression. Interestingly, the hierarchy of susceptibility to glutamine metabolism and the inhibition of these specific pathways (proximal glycolysis, one carbon metabolism and Krebs cycle) correlated with the hierarchy of susceptibility to anti-PD-1 suggesting that tumor specific metabolism might contribute to resistance to immunotherapy. Overall these results accentuate the importance of glutamine requiring metabolic pathways independent of the role of exogenous glutamine uptake. Further, our results highlight the lack of plasticity of highly coordinated tumor metabolic programming; targeting glutamine metabolism inhibited proximal glycolysis, one carbon metabolism and Krebs cycle. In doing so, we identify novel susceptibilities to exploit for inhibiting tumor growth. Citation Format: Liang Zhao, Matthew Arwood, Min-Hee Oh, Wei Xu, Im-Hong Sun, Im-Meng Sun, Chirag Patel, Robert Leone, Jesse Alt, Rana Rais, Barbara Slusher, Jonathan D. Powell. Targeting glutamine metabolism disables Warburg physiology by inhibiting proximal glycolysis and Krebs cycle rewiring [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4376.

Metabolic support of the respiratory burst in blood and sputum neutrophils of patients with community-acquired pneumonia

The aim of the study was to investigate relationships between the respiratory burst in blood and sputum neutrophils and intracellular enzyme activity in patients with pneumonia. Methods. The study involved 82 patients with moderate to severe community-acquired pneumonia (CAP). The chemiluminescent assay was used to investigate synthesis of primary and secondary reactive oxygen species (ROS). The bioluminescent method was used to examine NAD(P)-dependent dehydrogenase activity in neutrophils. Results. Low-intensity respiratory burst in blood neutrophils and its predominant relations to the state of mitochondrial metabolism and malic enzyme activity were observed in healthy subjects. Blood neutrophils in patients with CAP were activated; the anaerobic respiration and mitochondrial metabolism increased in the cells. Additionally, intensity of the respiratory burst was related to terminal glycolysis reactions and the key reaction of the pentose phosphate cycle. Sputum neutrophils in patients with CAP were also activated. Conclusion. Abnormalities of metabolic support of the respiratory burst, such as decreased production of primary and secondary ROS, could appear under unfavorable conditions. Peroxidation increased in the cells, the intensity of terminal glycolysis reactions decreased, and substrates transferred from the citric acid cycle to amino acid exchange reactions. Under these conditions, malic enzyme remained the only substrate that stimulated the respiratory burst in sputum neutrophils, while substrate transfer from the tricarboxylic acid cycle inhibited synthesis of the secondary ROS.

Glucose Metabolism in Cardiac Hypertrophy and Heart Failure.

Glucose Metabolism in Cardiac Hypertrophy and Heart Failure.

Metabolic reprogramming and Notch activity distinguish between non-small cell lung cancer subtypes

BackgroundPrevious studies suggested that the metabolism is differently reprogrammed in the major subtypes of non-small cell lung cancer (NSCLC), squamous cell carcinomas (SCC) and adenocarcinomas (AdC). However, a comprehensive analysis of this differential metabolic reprogramming is lacking.MethodsPublicly available gene expression data from human lung cancer samples and cell lines were analysed. Stable isotope resolved metabolomics were performed on SCC and ADC tumours in human patients and in freshly resected tumour slices.ResultsAnalysis of multiple transcriptomics data from human samples identified a SCC-distinguishing enzyme gene signature. SCC tumours from patients infused with [U-13C]-glucose and SCC tissue slices incubated with stable isotope tracers demonstrated differential glucose and glutamine catabolism compared to AdCs or non-cancerous lung, confirming increased activity through pathways defined by the SCC metabolic gene signature. Furthermore, the upregulation of Notch target genes was a distinguishing feature of SCCs, which correlated with the metabolic signature. Notch and MYC-driven murine lung tumours recapitulated the SCC-distinguishing metabolic reprogramming. However, the differences between SCCs and AdCs disappear in established cell lines in 2D culture.ConclusionsOur data emphasise the importance of studying lung cancer metabolism in vivo. They also highlight potential targets for therapeutic intervention in SCC patients including differentially expressed enzymes that catalyse reactions in glycolysis, glutamine catabolism, serine, nucleotide and glutathione biosynthesis.

Colloidal stability and dynamic mechanical properties of asphalt modified with unsaturated polyester prepared from waste polyethylene terephthalate.

This research aims to prepare unsaturated polyesters (UP) from polyethylene terephthalate waste (WPET) and studying their effects on the colloidal stability and dynamic mechanical properties of the prepared modified asphalt samples. To achieve this aim WPET is chemically recycled through glycolysis reaction using different glycols namely, diethylene glycol (DEG), triethylene glycol (TEG) and propylene glycol(PG) at molar ratios of PET wastes/glycol; 1:1.2 separately. The glycolyzed products were converted into unsaturated polyester (UP) via reaction with maleic anhydride. The prepared unsaturated polyesters (UPd, UPt and UPpg) were characterized using FTIR, GPC and TGA and then added to asphalt 60/70 in percentages ranging from 2wt% to 8wt% using wet process to prepare modified asphalt binder with desirable physical and engineering properties. The prepared modified asphalt samples were evaluated for thermal stability using TGA, physical and chemical properties using conventional tests, colloidal stability index while their rheological properties were investigated using Superpave binder tester, namely, dynamic shear rheometer (DSR) to determine the rutting and stiffness resistance of the prepared samples. The results showed an improvement in asphalt properties which became more thermally, colloidally stable and resistant to rutting and plastic deformation.

Porphyrin Excretion Resulting From Mutation of a Gene Encoding a Class I Fructose 1,6-Bisphosphate Aldolase in Rhodobacter capsulatus.

This paper describes a mutant (called SB1707) of the Rhodobacter capsulatus wild type strain SB1003 in which a transposon-disrupted rcc01707 gene resulted in a ∼25-fold increase in the accumulation of coproporphyrin III in the medium of phototrophic (anaerobic) cultures grown in a yeast extract/peptone medium. There was little or no stimulation of pigment accumulation in aerobic cultures. Therefore, this effect of rcc01707 mutation appears to be specific for the anaerobic coproporphyrinogen III oxidase HemN as opposed to the aerobic enzyme HemF. The protein encoded by rcc01707 is homologous to Class I fructose 1,6-bisphosphate aldolases, which catalyze a glycolytic reaction that converts fructose 1, 6-bisphosphate to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate, precursors of pyruvate. There were significant differences in coproporphyrin III accumulation using defined media with individual organic acids and sugars as the sole carbon source: pyruvate, succinate and glutamate stimulated accumulation the most, whereas glucose suppressed coproporphyrin III accumulation to 10% of that of succinate. However, although quantitatively lesser, similar effects of carbon source on the amount of accumulated pigment in the culture medium were seen in a wild type control. Therefore, this mutation appears to exaggerate effects also seen in the wild type strain. It is possible that mutation of rcc01707 causes a metabolic bottleneck or imbalance that was not rectified during growth on the several carbon sources tested. However, we speculate that, analogous to other fructose 1,6-bisphosphate aldolases, the rcc01707 gene product has a “moonlighting” activity that in this case is needed for the maximal expression of the hemN gene. Indeed, it was found that the rcc01707 gene is needed for maximal expression of a hemN promoter-lacZ reporter. With the decrease in hemN expression due to the absence of the rcc01707 gene product, coproporphyrinogen III accumulates and is released from the cell, yielding the spontaneous oxidation product coproporphyrin III.