A receptor guanylyl cyclase expressed specifically in olfactory sensory neurons.

We have cloned an additional member (GC-D) of the membrane receptor guanylyl cyclase [GTP pyrophosphate-lyase (cyclizing), EC 4.6.1.2] family that is specifically expressed in a subpopulation of olfactory sensory neurons. The extracellular, putative ligand-binding domain of the olfactory cyclase is similar in primary structure to two guanylyl cyclases expressed in the retina but diverges considerably from other known guanylyl cyclases. The expression of GC-D RNA is restricted to a small, randomly dispersed population of neurons that is within a single topographic zone in the olfactory neuroepithelium and resembles the pattern of the more diverse seven-transmembrane-domain odorant receptors. These observations suggest that GC-D may function directly in odor recognition or in modulating the sensitivity of a subpopulation of sensory neurons to specific odors.

Basis of guanylate cyclase activation by carbon monoxide.

Kinetics of CO association with guanylate cyclase [GTP pyrophosphate-lyase (cyclizing), EC 4.6.1.2] and dissociation from carboxy guanylate cyclase have been studied at pH 7.5 by flash photolysis, yielding rate constants at 23 degrees C of 1.2 +/- 0.1 x 10(5) M-1.sec-1 and 28 +/- 2 sec-1, respectively. While the CO combination rate constant is the same as for the T state of hemoglobin, the CO dissociation rate constant is much higher than expected for a six-coordinate carboxyheme protein; yet the absorption spectrum is indicative of a six-coordinate heme. The two observations are reconciled by a reaction mechanism in which CO dissociation proceeds via a five-coordinate intermediate. This intermediate is structurally very similar to the five-coordinate nitrosyl heme derivative of guanylate cyclase and is presumably responsible for the observed 4-fold activation of guanylate cyclase by CO. Thus, we provide a model that explains enzyme activities of the nitrosyl and carboxy forms of the enzyme on the basis of a common mechanism.

Aerobic cultivation of Streptococcus zooepidemicus and the role of NADH oxidase

A metabolic flux model was developed for Streptococcus zooepidemicus to compare the metabolism of glucose and maltose during aerobic batch cultivation. Lactic acid was the main product of glucose metabolism whereas acetic acid was the main product of maltose metabolism. This difference was chiefly attributed to the two-fold higher flux through NADH oxidase in maltose-grown cells that enabled the ATP generation rate to remain high despite a slower maltose consumption rate. The two-fold higher flux was matched by a two-fold increase in NADH oxidase activity, 2.53 +/- 0.1 mumol NADH min(-1) mg(-1) protein on maltose versus 1.07 +/- 0.04 Rmol NADH min(-1) mg(-1) protein on glucose, indicating that NADH oxidase activity is regulated by the energy status of the cell. Surprisingly, the energy status of the cell had little impact on hyaluronic acid (HA) yield and molecular weight. (C) 2003 Elsevier Science B.V. All rights reserved.

Elicitors of induced disease resistance in postharvest horticultural crops: a brief review

Increasing loss of conventional fungicides due to pathogen resistance and general unacceptability in terms of public and environmental risk have favoured the introduction of integrated pest management (IPM) programmes. Induction of natural disease resistance (NDR) in harvested horticultural crops using physical, biological and/or chemical elicitors has received increasing attention over recent years, it being considered a preferred strategy for disease management. This article reviews the enhancement of constitutive and inducible antifungal compounds and suppression of postharvest diseases through using elicitors. The effect of timing of pre- and/or postharvest elicitor treatment and environment on the degree of elicitation and the potential for inducing local acquired resistance, systemic acquired resistance and/or induced systemic resistance to reduce postharvest disease is discussed. The review highlights that more applied and basic research is required to understand the role that induced NDR can play in achieving practical suppression of postharvest diseases as part of an IPM approach. (C) 2003 Elsevier B.V. All rights reserved.

Inhibition of cell-wall autolysis and pectin degradation by cations

Modification of cell wall components such as cellulose, hemicellulose and pectin plays an important role in cell expansion. Cell expansion is known to be diminished by cations but it is unknown if this results from cations reacting with pectin or other cell wall components. Autolysis of cell wall material purified from bean root (Phaseolus vulgaris L.) occurred optimally at pH 5.0 and released mainly neutral sugars but very little uronic acid. Autolytic release of neutral sugars and uronic acid was decreased when cell wall material was loaded with Ca, Cu, Sr, Zn, Al or La cations. Results were also extended to a metal-pectate model system, which behaved similarly to cell walls and these cations also inhibited the enzymatic degradation by added polygalacturonase (EC 3.2.1.15). The extent of sugar release from cation-loaded cell wall material and pectate gels was related to the degree of cation saturation of the substrate, but not to the type of cation. The binding strength of the cations was assessed by their influence on the buffer capacity of the cell wall and pectate. The strongly bound cations (Cu, Al or La) resulted in higher cation saturation of the substrate and decreased enzymatic degradability than the weakly held cations (Ca, Sr and Zn). The results indicate that the junction zones between pectin molecules can peel open with weakly held cations, allowing polygalacturonase to cleave the hairy region of pectin, while strongly bound cations or high concentrations of cations force the junction zone closed, minimising enzymatic attack on the pectin backbone. (C) 2004 Elsevier SAS. All rights reserved.

Alkali hydroxide-induced gelation of pectin

The mechanism of pectin gelation depends on the degree of methoxylation. High methoxyl pectin gels due to hydrophobic interactions and hydrogen bonding between pectin molecules. Low methoxyl pectin forms gels in the presence of di- and polyvalent cations which cross link and neutralise the negative charges of the pectin molecule. Monovalent cations normally do not lead to gel formation with high methoxyl pectin solutions free of divalent cations, especially Ca. The present study found that alkali (NaOH or KOH) added to high methoxyl pectin leads to gel formation in a concentration-depended manner. It was also found that monovalent cations (Na and K) induce gelation of low methoxyl pectin and the time required for gel formation (setting time) depends on the cation concentration. The results indicate that a combined char-e neutralisation and ionic strength effect is responsible for the monovalent cation-induced gelation of pectin. (C) 2003 Elsevier Ltd. All rights reserved.

Functional significance of dauciform roots: exudation of carboxylates and acid phosphatase under phosphorus deficiency in Caustis blakei (Cyperaceae)

Caustis blakei produces an intriguing morphological adaptation by inducing dauciform roots in response to phosphorus (P) deficiency. We tested the hypothesis that these hairy, swollen lateral roots play a similar role to cluster roots in the exudation of organic chelators and ectoenzymes known to aid the chemical mobilization of sparingly available soil nutrients, such as P. Dauciform-root development and exudate composition (carboxylates and acid phosphatase activity) were analysed in C. blakei plants grown in nutrient solution under P-starved conditions. The distribution of dauciform roots in the field was determined in relation to soil profile depth and matrix. The percentage of dauciform roots of the entire root mass was greatest at the lowest P concentration ([P]) in solution, and was suppressed with increasing solution [P], while in the field dauciform roots were predominately located in the upper soil horizons, and decreased with increasing soil depth. Citrate was the major carboxylate released in an exudative burst from mature dauciform roots, which also produced elevated levels of acid phosphatase activity. Malonate was the dominant internal carboxylate present, with the highest concentration in young dauciform roots. The high concentration of carboxylates and phosphatases released from dauciform roots, combined with their prolific distribution in the organic surface layer of nutrient-impoverished soils, provides an ecophysiological advantage for enhancing nutrient acquisition.

Stem End blockaged in Cut grevillea 'Crimson Yul-lo' inflorescences

Grevillea 'Crimson Yul-lo' inflorescences have cut flower potential, but their vase life is short. End of vase life is characterized by early wilting. The possibility of physiologically mediated stem end blockage was investigated. Hydraulic conductance of 2 cm long stem end segments declined rapidly and remained lower throughout vase life than that of 2 cm long stem segments from immediately above. Recutting daily to remove basal 2 cm stem ends increased solution uptake, delayed declines in inflorescence water potential and water content, and improved inflorescence vase life. S-carvone is a potential inhibitor of wound related suberin formation, via inhibition of phenylalanine ammonia-lyase. Vase solution treatments with S-carvone (0.318 and 0.636 mM) delayed the decline in hydraulic conductance of basal 2 cm long stem end segments and decreases in vase solution uptake and relative fresh weight of cut stems, and extended vase life. Treatments with the catechol oxidase inhibitor 4-hexylresorcinol (2.5-10 mM) also delayed stem end blockage. These findings suggest that stem end blockage in cut G. 'Crimson Yul-lo' stems is physiologically mediated. (C) 2006 Elsevier B.V. All rights reserved.

Unravelling the theories of pre-eclampsia:Are the protective pathways the new paradigm?

Pre-eclampsia is a vascular disorder of pregnancy where anti-angiogenic factors, systemic inflammation and oxidative stress predominate, but none can claim to cause pre-eclampsia. This review provides an alternative to the 'two-stage model' of pre-eclampsia in which abnormal spiral arteries modification leads to placental hypoxia, oxidative stress and aberrant maternal systemic inflammation. Very high maternal soluble fms-like tyrosine kinase-1 (sFlt-1 also known as sVEGFR) and very low placenta growth factor (PlGF) are unique to pre-eclampsia; however, abnormal spiral arteries and excessive inflammation are also prevalent in other placental disorders. Metaphorically speaking, pregnancy can be viewed as a car with an accelerator and brakes, where inflammation, oxidative stress and an imbalance in the angiogenic milieu act as the 'accelerator'. The 'braking system' includes the protective pathways of haem oxygenase 1 (also referred as Hmox1 or HO-1) and cystathionine-γ-lyase (also known as CSE or Cth), which generate carbon monoxide (CO) and hydrogen sulphide (H2S) respectively. The failure in these pathways (brakes) results in the pregnancy going out of control and the system crashing. Put simply, pre-eclampsia is an accelerator-brake defect disorder. CO and H2S hold great promise because of their unique ability to suppress the anti-angiogenic factors sFlt-1 and soluble endoglin as well as to promote PlGF and endothelial NOS activity. The key to finding a cure lies in the identification of cheap, safe and effective drugs that induce the braking system to keep the pregnancy vehicle on track past the finishing line.

Biosynthesis and analysis of plant oxylipins

The term oxylipin is applied to the generation of oxygenated products of polyunsaturated fatty acids that can arise either through non-enzymatic or enzymatic processes generating a complex array of products, including alcohols, aldehydes, ketones, acids and hydrocarbon gases. The biosynthetic origin of these products has revealed an array of enzymes involved in their formation and more recently a radical pathway. These include lipoxygenases and α-dioxygenase that insert both oxygen atoms in to the acyl chain to initiate the pathways, to specialised P450 monooxygenases that are responsible for their downstream processing. This latter group include enzymes at the branch points such as allene oxide synthase, leading to jasmonate signalling, hydroperoxide lyase, responsible for generating pathogen/pest defensive volatiles and divinyl ether synthases and peroxygenases involved in the formation of antimicrobial compounds. The complexity of the products generated raises significant challenges for their rapid identification and quantification using metabolic screening methods. Here the current developments in oxylipin metabolism are reviewed together with the emerging technologies required to expand this important field of research that underpins advances in plant-pest/pathogen interactions.

Resveratrol stimulates hydrogen sulfide (H2S) formation to relax murine corpus cavernosum

Introduction: Resveratrol (RVT) found in red wine protects against erectile dysfunction and relaxes penile tissue (corpus cavernosum) via a nitric oxide (NO) independent pathway. However, the mechanism remains to be elucidated. Hydrogen sulfide (H2S) is a potent vasodilator and neuromodulator generated in corpus cavernosum. Aims: We investigated whether RVT caused the relaxation of mice corpus cavernosum (MCC) through H2S. Methods: H2S formation is measured by methylene blue assay and vascular reactivity experiments have been performed by DMT strip myograph in CD1 MCC strips. Main Outcome Measures: Endothelial NO synthase (eNOS) inhibitor Nω-Nitro-L-arginine (L-NNA, 0.1mM) or H2S inhibitor aminooxyacetic acid (AOAA, 2mM) which inhibits both cystathionine-β-synthase (CBS) and cystathionine-gamma-lyase (CSE) enzyme or combination of AOAA with PAG (CSE inhibitor) has been used in the presence/absence of RVT (0.1mM, 30min) to elucidate the role of NO or H2S pathways on the effects of RVT in MCC. Concentration-dependent relaxations to RVT, L-cysteine, sodium hydrogen sulfide (NaHS) and acetylcholine (ACh) were studied. Results: Exposure of murine corpus cavernosum to RVT increased both basal and L-cysteine-stimulated H2S formation. Both of these effects were reversed by AOAA but not by L-NNA. RVT caused concentration-dependent relaxation of MCC and that RVT-induced relaxation was significantly inhibited by AOAA or AOAA+PAG but not by L-NNA. L-cysteine caused concentration-dependent relaxations, which are inhibited by AOAA or AOAA+PAG significantly. Incubation of MCC with RVT significantly increased L-cysteine-induced relaxation, and this effect was inhibited by AOAA+PAG. However, RVT did not alter the effect of exogenous H2S (NaHS) or ACh-induced relaxations. Conclusions: These results demonstrate that RVT-induced relaxation is at least partly dependent on H2S formation and acts independent of eNOS pathway. In phosphodiesterase 5 inhibitor (PDE-5i) nonresponder population, combination therapy with RVT may reverse erectile dysfunction via stimulating endogenous H2S formation. Yetik-Anacak G, Dereli MV, Sevin G, Ozzayim O, Erac Y, and Ahmed A. Resveratrol stimulates hydrogen sulfide (H2S) formation to relax murine corpus cavernosum.

Can hydrogen sulfide prevent preeclampsia and fetal growth restriction?

The exact aetiology of preeclampsia is unknown, but there is a good association with an imbalance in angiogenic growth factors and abnormal placentation [1]. Hydrogen sulphide (H2S), a gaseous messenger produced mainly by cystathionine γ-lyase (CSE), is pro-angiogenic vasodilator [2] and [3]. We hypothesized that a reduction in CSE activity may alter the angiogenic balance in pregnancy and induce abnormal placentation and maternal hypertension. Plasma levels of H2S were significantly decreased in preeclamptic women (p < 0.01), which was associated with reduced CSE message and protein expression in human placenta as determined by real-time PCR and immunohistochemistry. Inhibition of CSE activity by DL-propargylglycine (PAG) in first trimester (8–12 weeks gestation) human placental explants had reduced placenta growth factor (PlGF) production as assessed by ELISA and inhibited trophoblast invasion in vitro. Endothelial CSE knockdown by siRNA transfection increased the endogenous release of soluble fms-Like tyrosine kinase-1 (sFlt-1) and soluble endoglin, (sEng) from human umbilical vein endothelial cells while adenoviral-mediated CSE overexpression inhibited their release. Administration of PAG to pregnant mice induced hypertension, liver damage, and promoted abnormal labyrinth vascularisation in the placenta and decreased fetal growth. Finally, a slow releasing, H2S-generating compound, GYY4137, inhibited circulating sFlt-1 and sEng levels and restored fetal growth that was compromised by PAG-treatment demonstrating that the effect of CSE inhibitor was due to inhibition of H2S production. These results imply that endogenous H2S is required for healthy placental vasculature and a decrease in of CSE/H2S activity may contribute to the pathogenesis of preeclampsia. References [1] S. Ahmad, A. Ahmed, Elevated placental soluble vascular endothelial growth factor receptor-1 inhibits angiogenesis in preeclampsia, Circ Res., 95 (2004), pp. 884–891. [2] G. Yang, et al., H2S as a physiologic vasorelaxant: hypertension in mice with deletion of cystathionine gamma-lyase, Science, 322 (2008), pp. 587–590. [3] A. Papapetropoulos, et al., Hydrogen sulfide is an endogenous stimulator of angiogenesis, Proc Natl Acad Sci USA, 106 (2009), pp. 21972–21977.

Role of hydrogen sulfide in the female reproductive tract

It is well established that hydrogen sulfide (H 2S) has a signaling role in the body. So far it has been shown that H 2S is produced by intra-uterine tissues in the pregnant rat and the human placenta. Two main enzymes responsible for H 2S production, cystathionine- synthase and cystathionine-lyase, have been demonstrated in the pregnant and nonpregnant uterus, fetal membranes and placenta in the rat, and in human placenta. H 2S donors have been shown to inhibit contraction of the pregnant rat uterus. H 2S could play a role in maintaining uterine quiescence during pregnancy, as an oxygen sensor and vasodilator in the placenta, or as an anti-inflammatory. More research is required in this area to elucidate the roles of H 2S in the female reproductive tract and its mechanisms of action. © 2010 Expert Reviews Ltd.

Filogenia molecular das enzimas isocitrato liase e malato sintase e sua evolução em Viridiplanta

The plant metabolism consists of a complex network of physical and chemical events resulting in photosynthesis, respiration, synthesis and degradation of organic compounds. This is only possible due to the different kinds of responses to many environmental variations that a plant could be subject through evolution, leading also to conquering new surroundings. The glyoxylate cycle is a metabolic pathway found in glyoxysomes plant, which has unique role in the seedling establishment. Considered as a variation of the citric acid cycle, it uses an acetyl coenzyme A molecule, derived from lipids beta-oxidation to synthesize compounds which are used in carbohydrate synthesis. The Malate synthase (MLS) and Isocitrate lyase (ICL) enzyme of this cycle are unique and essential in regulating the biosynthesis of carbohydrates. Because of the absence of decarboxylation steps as rate-limiting steps, detailed studies of molecular phylogeny and evolution of these proteins enables the elucidation of the effects of this route presence in the evolutionary processes involved in their distribution across the genome from different plant species. Therefore, the aim of this study was to establish a relationship between the molecular evolution of the characteristics of enzymes from the glyoxylate cycle (isocitrate lyase and malate synthase) and their molecular phylogeny, among green plants (Viridiplantae). For this, amino acid and nucleotide sequences were used, from online repositories as UniProt and Genbank. Sequences were aligned and then subjected to an analysis of the best-fit substitution models. The phylogeny was rebuilt by distance methods (neighbor-joining) and discrete methods (maximum likelihood, maximum parsimony and Bayesian analysis). The identification of structural patterns in the evolution of the enzymes was made through homology modeling and structure prediction from protein sequences. Based on comparative analyzes of in silico models and from the results of phylogenetic inferences, both enzymes show significant structure conservation and their topologies in agreement with two processes of selection and specialization of the genes. Thus, confirming the relevance of new studies to elucidate the plant metabolism from an evolutionary perspective

Effets de mutations dans le gène ldhA et dans la protéine FhlA ainsi que de la limitation en glucose ou en soufre sur la production d'hydrogène chez Escherichia coli

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.