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1
Jovanović, S., and S. Weber. "Modélisation et accélération de réseaux de neurones profonds (CNN) en Python/VHDL/C++ et leur vérification et test à l’aide de l’environnement Pynq sur les FPGA Xilinx." J3eA 21 (2022): 1028. http://dx.doi.org/10.1051/j3ea/20220028.
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Nous présentons un ensemble de travaux pratiques qui seront dispensés au sein du Master EEA - Électronique Embarquée à l’université de Lorraine dans le cadre des modules Modélisation SystemC et Conception VLSI. Ces TP sont destinés à initier les étudiants à la compréhension, modélisation et conception des réseaux de neurones convolutifs dans des langages de description de matériel au niveau RTL (VHDL, le module Conception VLSI) et dans un langage de haut niveau (C++/SystemC, le module Modélisation SystemC). Ils sont organisés autour d’un ensemble d’outils de modélisation et de synthèse de Mentor Graphics (Modelsim, Catapult HLS) et spécifiques aux plateformes FPGA Xilinx et à l’environnement Pynq pour la simulation, test et vérification.
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Subramaniam, Prem S., Gang Xie, Tianhui Xia, and Roy A. Jensen. "Substrate Ambiguity of 3-Deoxy-d-manno-Octulosonate 8-Phosphate Synthase from Neisseria gonorrhoeae in the Context of Its Membership in a Protein Family Containing a Subset of 3-Deoxy-d-arabino-Heptulosonate 7-Phosphate Synthases." Journal of Bacteriology 180, no. 1 (January 1, 1998): 119–27. http://dx.doi.org/10.1128/jb.180.1.119-127.1998.
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ABSTRACT 3-Deoxy-d-manno-octulosonate 8-phosphate (KDOP) synthase and 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthase catalyze similar phosphoenolpyruvate-utilizing reactions. The genome of Neisseria gonorrhoeae contains one gene encoding KDOP synthase and one gene encoding DAHP synthase. Of the two nonhomologous DAHP synthase families known, the N. gonorrhoeae protein belongs to the family I assemblage. KDOP synthase exhibited an ability to replace arabinose-5-P with either erythrose-4-P or ribose-5-P as alternative substrates. The results of periodate oxidation studies suggested that the product formed by KDOP synthase with erythrose-4-P as the substrate was 3-deoxy-d-ribo-heptulosonate 7-P, an isomer of DAHP. As expected, this product was not utilized as a substrate by dehydroquinate synthase. The significance of the ability of KDOP synthase to substitute erythrose-4-P for arabinose-5-P is (i) recognition of the possibility that the KDOP synthase might otherwise be mistaken for a species of DAHP synthase and (ii) the possibility that the broad-specificity type of KDOP synthase might be a relatively vulnerable target for antimicrobial agents which mimic the normal substrates. An analysis of sequences in the database indicates that the family I group of DAHP synthase has a previously unrecognized membership which includes the KDOP synthases. The KDOP synthases fall into a subfamily grouping which includes a small group of DAHP synthases. Thus, family I DAHP synthases separate into two subfamilies, one of which includes the KDOP synthases. The two subfamilies appear to have diverged prior to the acquisition of allosteric-control mechanisms for DAHP synthases. These allosteric control specificities are highly diverse and correlate with the presence of N-terminal extensions which lack homology with one another.
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Kameya, Masafumi, Takeshi Ikeda, Miyuki Nakamura, Hiroyuki Arai, Masaharu Ishii, and Yasuo Igarashi. "A Novel Ferredoxin-Dependent Glutamate Synthase from the Hydrogen-Oxidizing Chemoautotrophic Bacterium Hydrogenobacter thermophilus TK-6." Journal of Bacteriology 189, no. 7 (January 19, 2007): 2805–12. http://dx.doi.org/10.1128/jb.01360-06.
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ABSTRACT Glutamate synthases are classified according to their specificities for electron donors. Ferredoxin-dependent glutamate synthases had been found only in plants and cyanobacteria, whereas many bacteria have NADPH-dependent glutamate synthases. In this study, Hydrogenobacter thermophilus, a hydrogen-oxidizing chemoautotrophic bacterium, was shown to possess a ferredoxin-dependent glutamate synthase like those of phototrophs. This is the first observation, to our knowledge, of a ferredoxin-dependent glutamate synthase in a nonphotosynthetic organism. The purified enzyme from H. thermophilus was shown to be a monomer of a 168-kDa polypeptide homologous to ferredoxin-dependent glutamate synthases from phototrophs. In contrast to known ferredoxin-dependent glutamate synthases, the H. thermophilus glutamate synthase exhibited glutaminase activity. Furthermore, this glutamate synthase did not react with a plant-type ferredoxin (Fd3 from this bacterium) containing a [2Fe-2S] cluster but did react with bacterial ferredoxins (Fd1 and Fd2 from this bacterium) containing [4Fe-4S] clusters. Interestingly, the H. thermophilus glutamate synthase was activated by some of the organic acids in the reductive tricarboxylic acid cycle, the central carbon metabolic pathway of this organism. This type of activation has not been reported for any other glutamate synthases, and this property may enable the control of nitrogen assimilation by carbon metabolism.
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Nesterov, Semen, Yury Chesnokov, Roman Kamyshinsky, Alisa Panteleeva, Konstantin Lyamzaev, Raif Vasilov, and Lev Yaguzhinsky. "Ordered Clusters of the Complete Oxidative Phosphorylation System in Cardiac Mitochondria." International Journal of Molecular Sciences 22, no. 3 (February 2, 2021): 1462. http://dx.doi.org/10.3390/ijms22031462.
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The existence of a complete oxidative phosphorylation system (OXPHOS) supercomplex including both electron transport system and ATP synthases has long been assumed based on functional evidence. However, no structural confirmation of the docking between ATP synthase and proton pumps has been obtained. In this study, cryo-electron tomography was used to reveal the supramolecular architecture of the rat heart mitochondria cristae during ATP synthesis. Respirasome and ATP synthase structure in situ were determined using subtomogram averaging. The obtained reconstructions of the inner mitochondrial membrane demonstrated that rows of respiratory chain supercomplexes can dock with rows of ATP synthases forming oligomeric ordered clusters. These ordered clusters indicate a new type of OXPHOS structural organization. It should ensure the quickness, efficiency, and damage resistance of OXPHOS, providing a direct proton transfer from pumps to ATP synthase along the lateral pH gradient without energy dissipation.
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Ma, Li-Ting, Pi-Ling Liu, Yang-Tui Cheng, Tz-Fan Shiu, and Fang-Hua Chu. "Unveiling Monoterpene Biosynthesis in Taiwania cryptomerioides via Functional Characterization." Plants 10, no. 11 (November 8, 2021): 2404. http://dx.doi.org/10.3390/plants10112404.
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Taiwania cryptomerioides is a monotypic species, and its terpenoid-rich property has been reported in recent years. To uncover monoterpene biosynthesis in T. cryptomerioides, this study used transcriptome mining to identify candidates with tentative monoterpene synthase activity. Along with the phylogenetic analysis and in vitro assay, two geraniol synthases (TcTPS13 and TcTPS14), a linalool synthase (TcTPS15), and a β-pinene synthase (TcTPS16), were functionally characterized. Via the comparison of catalytic residues, the Cys/Ser at region 1 might be crucial in determining the formation of α-pinene or β-pinene. In addition, the Cupressaceae monoterpene synthases were phylogenetically clustered together; they are unique and different from those of published conifer species. In summary, this study aimed to uncover the ambiguous monoterpenoid network in T. cryptomerioide, which would expand the landscape of monoterpene biosynthesis in Cupressaceae species.
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Agger, Sean A., Fernando Lopez-Gallego, Thomas R. Hoye, and Claudia Schmidt-Dannert. "Identification of Sesquiterpene Synthases from Nostoc punctiforme PCC 73102 and Nostoc sp. Strain PCC 7120." Journal of Bacteriology 190, no. 18 (July 25, 2008): 6084–96. http://dx.doi.org/10.1128/jb.00759-08.
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ABSTRACT Cyanobacteria are a rich source of natural products and are known to produce terpenoids. These bacteria are the major source of the musty-smelling terpenes geosmin and 2-methylisoborneol, which are found in many natural water supplies; however, no terpene synthases have been characterized from these organisms to date. Here, we describe the characterization of three sesquiterpene synthases identified in Nostoc sp. strain PCC 7120 (terpene synthase NS1) and Nostoc punctiforme PCC 73102 (terpene synthases NP1 and NP2). The second terpene synthase in N. punctiforme (NP2) is homologous to fusion-type sesquiterpene synthases from Streptomyces spp. shown to produce geosmin via an intermediate germacradienol. The enzymes were functionally expressed in Escherichia coli, and their terpene products were structurally identified as germacrene A (from NS1), the eudesmadiene 8a-epi-α-selinene (from NP1), and germacradienol (from NP2). The product of NP1, 8a-epi-α-selinene, so far has been isolated only from termites, in which it functions as a defense compound. Terpene synthases NP1 and NS1 are part of an apparent minicluster that includes a P450 and a putative hybrid two-component protein located downstream of the terpene synthases. Coexpression of P450 genes with their adjacent located terpene synthase genes in E. coli demonstrates that the P450 from Nostoc sp. can be functionally expressed in E. coli when coexpressed with a ferredoxin gene and a ferredoxin reductase gene from Nostoc and that the enzyme oxygenates the NS1 terpene product germacrene A. This represents to the best of our knowledge the first example of functional expression of a cyanobacterial P450 in E. coli.
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Eichman, Jennifer. "Intertextual Alliances: Huang Hui’s Synthesis of Confucian and Buddhist Paths to Liberation." T’oung pao 100, no. 1-3 (November 24, 2014): 120–63. http://dx.doi.org/10.1163/15685322-10013p04.
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This article argues for a reconsideration of how we categorize individual attempts at sanjiao heyi-style syntheses and characterize the broader late sixteenth-century milieu that nourished such attempts. In Zeng Zheng Kunyan bieyan 贈鄭昆嚴別言 (Parting Words for Zheng Kunyan), Huang Hui 黃輝 (1555–1612) synthesized a highly selective number of Chan Buddhist and Yangming Confucian ideas to create a path to self-cultivation rooted in the interstitial dialogue between the branch of third-generation Yangming Confucians headed by Zhou Rudeng 周汝登 (1547–1629) and the Buddhist teachings expounded by the monk Zhuhong 袾宏 (1535–1615). Unlike Confucian scholars who wrote polemical sanjiao heyi texts, Huang was an enthusiastic synthesizer intent on benefiting from both Buddhist and Confucian traditions. A close analysis of his work offers one illustration of how such syntheses were constructed while further revealing the broader philosophical discourse generated by Huang’s circle. Cet article invite à reconsidérer la façon dont sont catégorisées les tentatives individuelles de synthèse entre les “trois religions” (sanjiao heyi) et dont on décrit plus généralement le milieu qui produisait ce genre de tentative à la fin du xvie siècle. Dans ses Paroles d’adieu pour Zheng Kunyan (Zeng Zheng Kunyan bieyan 贈鄭昆嚴別言), Huang Hui 黃輝 (1555–1612) synthétise un certain nombre d’idées soigneusement choisies au sein du bouddhisme chan et du confucianisme de l’école de Wang Yangming pour inventer une voie de perfectionnement moral enracinée dans le dialogue interstitiel entre le groupe de la troisième génération des disciples de Wang Yangming mené par Zhou Rudeng 周汝登 (1547–1629) et les enseignements bouddhistes du moine Zhuhong 袾宏 (1535–1615). Contrairement aux lettrés confucéens qui produisaient des textes polémiques sur les trois religions, Huang se révèle un ardent partisan de la synthèse et cherche à tirer parti des deux traditions, bouddhiste et confucéenne. L’examen attentif de son œuvre illustre la façon dont de telles synthèses étaient édifiées, tout en mettant en évidence le discours philosophique plus général émanant du cercle auquel appartenait Huang.
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Xu, Meimei, P. Ross Wilderman, and Reuben J. Peters. "Following evolution's lead to a single residue switch for diterpene synthase product outcome." Proceedings of the National Academy of Sciences 104, no. 18 (April 24, 2007): 7397–401. http://dx.doi.org/10.1073/pnas.0611454104.
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There have been few insights into the biochemical origins of natural product biosynthesis from primary metabolism. Of particular interest are terpene synthases, which often mediate the committed step in particular biosynthetic pathways so that alteration of their product outcome is a key step in the derivation of novel natural products. These enzymes also catalyze complex reactions of significant mechanistic interest. Following an evolutionary lead from two recently diverged, functionally distinct diterpene synthase orthologs from different subspecies of rice, we have identified a single residue that can switch product outcome. Specifically, the mutation of a conserved isoleucine to threonine that acts to convert not only the originally targeted isokaurene synthase into a specific pimaradiene synthase but also has a much broader effect, which includes conversion of the ent-kaurene synthases found in all higher plants for gibberellin phytohormone biosynthesis to the production of pimaradiene. This surprisingly facile switch for diterpene synthase catalytic specificity indicates the ease with which primary (gibberellin) metabolism can be subverted to secondary biosynthesis and may underlie the widespread occurrence of pimaradiene-derived natural products. In addition, because this isoleucine is required for the mechanistically more complex cyclization to tetracyclic kaurene, whereas substitution with threonine “short-circuits” this mechanism to produce the “simpler” tricyclic pimaradiene, our results have some implications regarding the means by which terpene synthases specify product outcome.
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Antonsson, Bruno E., and Lisa S. Klig. "Candida albicans phosphatidylinositol synthase has common features with both Saccharomyces cerevisiae and mammalian phosphatidylinositol synthases." Yeast 12, no. 5 (April 1996): 449–56. http://dx.doi.org/10.1002/(sici)1097-0061(199604)12:5<449::aid-yea927>3.0.co;2-p.
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Tang, Eva H. C., and Paul M. Vanhoutte. "Gene expression changes of prostanoid synthases in endothelial cells and prostanoid receptors in vascular smooth muscle cells caused by aging and hypertension." Physiological Genomics 32, no. 3 (February 2008): 409–18. http://dx.doi.org/10.1152/physiolgenomics.00136.2007.
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The present study was designed to assess whether or not changes in genomic expression of cyclooxygenases (COX-1, COX-2), endothelial nitric oxide synthase (eNOS), and prostanoid synthases in the endothelium and of prostanoid receptors in vascular smooth muscle contribute to the occurrence of endothelium-dependent contractions during aging and hypertension. Gene expression was quantified by real-time PCR using isolated endothelial cells and smooth muscle cells (SMC) from the aorta of Wistar-Kyoto and spontaneously hypertensive rats. Genes for all known prostanoid synthases and receptors were present in endothelial cells and SMC, respectively. Aging caused overexpression of eNOS, COX-1, COX-2, thromboxane synthase, hematopoietic-type prostaglandin D synthase, membrane prostaglandin E synthase-2, and prostaglandin F synthase in endothelial cells and COX-1 and prostaglandin E2 (EP)4 receptors in SMC. Hypertension augmented the expression of COX-1, prostacyclin synthase, thromboxane synthase, and hematopoietic-type prostaglandin D synthase in endothelial cells and prostaglandin D2 (DP), EP3, and EP4 receptors in SMC. The increase in genomic expression of endothelial COX-1 explains why in aging and hypertension the endothelium has greater propensity to release cyclooxygenase-derived vasoconstrictive prostanoids. The expression of prostacyclin synthase was by far the most abundant, explaining why the majority of the COX-1-derived endoperoxides are transformed into prostacyclin, substantiating the role of prostacyclin as an endothelium-derived contracting factor. The expression of thromboxane synthase was increased in the cells of aging or hypertensive rats, explaining why the prostanoid can contribute to endothelium-dependent contractions. It is uncertain whether the gene modifications caused by aging and hypertension directly contribute to endothelium-dependent contractions or rather to vascular aging and the vascular complications of the hypertensive process.
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Shu, Shokoku, Mao Kobayashi, Kana Marunaka, Yuta Yoshino, Makiko Goto, Yuji Katsuta, and Akira Ikari. "Magnesium Supplementation Attenuates Ultraviolet-B-Induced Damage Mediated through Elevation of Polyamine Production in Human HaCaT Keratinocytes." Cells 11, no. 15 (July 22, 2022): 2268. http://dx.doi.org/10.3390/cells11152268.
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Magnesium ions (Mg2+) have favorable effects such as the improvement of barrier function and the reduction of inflammation reaction in inflammatory skin diseases. However, its mechanisms have not been fully understood. Microarray analysis has shown that the gene expressions of polyamine synthases are upregulated by MgCl2 supplementation in human HaCaT keratinocytes. Here, we investigated the mechanism and function of polyamine production. The mRNA and protein levels of polyamine synthases were dose-dependently increased by MgCl2 supplementation, which were inhibited by U0126, a MEK inhibitor; CHIR-99021, a glycogen synthase kinase-3 (GSK3) inhibitor; and Naphthol AS-E, a cyclic AMP-response-element-binding protein (CREB) inhibitor. Similarly, reporter activities of polyamine synthases were suppressed by these inhibitors, suggesting that MEK, GSK3, and CREB are involved in the transcriptional regulation of polyamine synthases. Cell viability was reduced by ultraviolet B (UVB) exposure, which was rescued by MgCl2 supplementation. The UVB-induced elevation of reactive oxygen species was attenuated by MgCl2 supplementation, which was inhibited by cysteamine, a polyamine synthase inhibitor. Our data indicate that the expression levels of polyamine synthases are upregulated by MgCl2 supplementation mediated through the activation of the MEK/GSK3/CREB pathway. MgCl2 supplementation may be useful in reducing the UVB-induced oxidative stress in the skin.
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Fan, Zeyu, Xinhao Li, Ruoyu Jiang, Jinqian Li, Fangyu Cao, Mingjuan Sun, and Lianghua Wang. "Molecular Dynamics Simulation Reveal the Structure–Activity Relationships of Kainoid Synthases." Marine Drugs 22, no. 7 (July 22, 2024): 326. http://dx.doi.org/10.3390/md22070326.
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Kainoid synthases are key enzymes in the biosynthesis of kainoids. Kainoids, as represented by DA and KA, are a class of naturally occurring non-protein amino acids with strong neurotransmitter activity in the mammalian central nervous system. Marine algae kainoid synthases include PnDabC from diatoms, which synthesizes domoic acid (DA), and DsKabC and GfKabC from red algae, which synthesize kainic acid (KA). Elucidation of the catalytic mechanism of kainoid synthases is of great significance for the rational design of better biocatalysts to promote the industrial production of kainoids for use in new drugs. Through modeling, molecular docking, and molecular dynamics simulations, we investigated the conformational dynamics of kainoid synthases. We found that the kainoid synthase complexes showed different stability in the simulation, and the binding and catalytic processes showed significant conformational transformations of kainoid synthase. The residues involved in specific interactions with the substrate contributed to the binding energy throughout the simulation process. Binding energy, the relaxed active pocket, electrostatic potential energy of the active pocket, the number and rotation of aromatic residues interacting with substrates during catalysis, and the number and frequency of hydrogen bonds between the individual functional groups revealed the structure–activity relationships and affected the degree of promiscuity of kainoid synthases. Our research enriches the understanding of the conformational dynamics of kainoid synthases and has potential guiding significance for their rational design.
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Fischer, M., W. Römisch, B. Illarionov, W. Eisenreich, and A. Bacher. "Structures and reaction mechanisms of riboflavin synthases of eubacterial and archaeal origin." Biochemical Society Transactions 33, no. 4 (August 1, 2005): 780–84. http://dx.doi.org/10.1042/bst0330780.
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The biosynthesis of one riboflavin molecule requires one molecule of GTP and two molecules of ribulose 5-phosphate as substrates. GTP is hydrolytically opened, converted into 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione by a sequence of deamination, side chain reduction and dephosphorylation. Condensation with 3,4-dihydroxy-2-butanone 4-phosphate obtained from ribulose 5-phosphate leads to 6,7-dimethyl-8-ribityllumazine. The dismutation of 6,7-dimethyl-8-ribityllumazine catalysed by riboflavin synthase produces riboflavin and 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione. A pentacyclic adduct of two 6,7-dimethyl-8-ribityllumazines has been identified earlier as a catalytically competent reaction intermediate of the Escherichia coli enzyme. Acid quenching of reaction mixtures of riboflavin synthase of Methanococcus jannaschii, devoid of similarity to riboflavin synthases of eubacteria and eukaryotes, afforded a compound whose optical absorption and NMR spectra resemble that of the pentacyclic E. coli riboflavin synthase intermediate, whereas the CD spectra of the two compounds have similar envelopes but opposite signs. Each of the compounds could serve as a catalytically competent intermediate for the enzyme by which it was produced, but not vice versa. All available data indicate that the respective pentacyclic intermediates of the M. jannaschii and E. coli enzymes are diastereomers. Whereas the riboflavin synthase of M. jannaschii is devoid of similarity with those of eubacteria and eukaryotes, it has significant sequence similarity with 6,7-dimethyl-8-ribityllumazine synthases catalysing the penultimate step of riboflavin biosynthesis. 6,7-Dimethyl-8-ribityllumazine synthase and the archaeal riboflavin synthase appear to have diverged early in the evolution of Archaea from a common ancestor.
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Ebizuka, Yutaka, Yuji Katsube, T. Tsutsumi, Tetsuo Kushiro, and M. Shibuya. "Functional genomics approach to the study of triterpene biosynthesis." Pure and Applied Chemistry 75, no. 2-3 (January 1, 2003): 369–74. http://dx.doi.org/10.1351/pac200375020369.
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The Arabidopsis thaliana genome-sequencing project has identified the presence of 13 oxidosqualene cyclase homologs in this plant. In addition to the already identified clones, namely, CAS1 cycloartenol synthase, LUP1 lupeol synthase, and YUP8H12R.43 multifunctional triterpene synthase, two new cDNAs of the putative oxidosqualene cyclase genes, F1019.4 and T30F21.16, were obtained by polymerase chain reaction (PCR) and functionally expressed in yeast. Liquid chromatography/mass spectrometry (LC/MS) analysis led to the identification of some of their reaction products. Interestingly, except for CAS1 for sterol biosynthesis of primary metabolism, so-far-obtained all triterpene synthases of this plant are multifunctional, producing more than one cyclization product. A feeding experiment of 13C-labeled acetate with LUP1 lupeol synthase transformant demonstrated the stereospecific water addition to lupenyl cation intermediate, yielding 3β,20 dihydroxylupane, which accounts for the multiproduct nature of this synthase.
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Webby, Celia J., Mark L. Patchett, and Emily J. Parker. "Characterization of a recombinant type II 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase from Helicobacter pylori." Biochemical Journal 390, no. 1 (August 9, 2005): 223–30. http://dx.doi.org/10.1042/bj20050259.
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DAH7P (3-Deoxy-D-arabino-heptulosonate 7-phosphate) synthase catalyses the condensation reaction between phosphoenolpyruvate (PEP) and D-erythrose 4-phosphate (E4P) as the first committed step in the biosynthesis of aromatic compounds in plants and micro-organisms. Previous work has identified two families of DAH7P synthases based on sequence similarity and molecular mass, with the majority of the mechanistic and structural studies being carried out on the type I paralogues from Escherichia coli. Whereas a number of organisms possess genes encoding both type I and type II DAH7P synthases, the pathogen Helicobacter pylori has only a single, type II, enzyme. Recombinant DAH7P synthase from H. pylori was partially solubilized by co-expression with chaperonins GroEL/GroES in E. coli, and purified to homogeneity. The enzyme reaction follows an ordered sequential mechanism with the following kinetic parameters: Km (PEP), 3 μM; Km (E4P), 6 μM; and kcat, 3.3 s−1. The enzyme reaction involves interaction of the si face of PEP with the re face of E4P. H. pylori DAH7P synthase is not inhibited by phenylalanine, tyrosine, tryptophan or chorismate. EDTA inactivates the enzyme, and activity is restored by a range of bivalent metal ions, including (in order of decreasing effectiveness) Co2+, Mn2+, Ca2+, Mg2+, Cu2+ and Zn2+. Analysis of type II DAH7P synthase sequences reveals several highly conserved motifs, and comparison with the type I enzymes suggests that catalysis by these two enzyme types occurs on a similar active-site scaffold and that the two DAH7P synthase families may indeed be distantly related.
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King, Ross D., and Chuan Lu. "An investigation into eukaryotic pseudouridine synthases." Journal of Bioinformatics and Computational Biology 12, no. 04 (August 2014): 1450015. http://dx.doi.org/10.1142/s0219720014500152.
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A common post-transcriptional modification of RNA is the conversion of uridine to its isomer pseudouridine. We investigated the biological significance of eukaryotic pseudouridine synthases using the yeast Saccharomyces cerevisiae. We conducted a comprehensive statistical analysis on growth data from automated perturbation (gene deletion) experiments, and used bi-logistic curve analysis to characterise the yeast phenotypes. The deletant strains displayed different alteration in growth properties, including in some cases enhanced growth and/or biphasic growth curves not seen in wild-type strains under matched conditions. These results demonstrate that disrupting pseudouridine synthases can have a significant qualitative effect on growth. We further investigated the significance of post-transcriptional pseudouridine modification through investigation of the scientific literature. We found that (1) In Toxoplasma gondii, a pseudouridine synthase gene is critical in cellular differentiation between the two asexual forms: Tachyzoites and bradyzoites; (2) Mutation of pseudouridine synthase genes has also been implicated in human diseases (mitochondrial myopathy and sideroblastic anemia (MLASA); dyskeratosis congenita). Taken together, these results are consistent with pseudouridine synthases having a Gene Ontology function of "biological regulation".
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Ma, Li, and John L. Wallace. "Endothelial nitric oxide synthase modulates gastric ulcer healing in rats." American Journal of Physiology-Gastrointestinal and Liver Physiology 279, no. 2 (August 1, 2000): G341—G346. http://dx.doi.org/10.1152/ajpgi.2000.279.2.g341.
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Nitric oxide has been shown to be beneficial for gastric ulcer healing. We determined the relative effects of endothelial and inducible nitric oxide synthases on gastric ulcer healing in rats. Ulcers were induced by serosal application of acetic acid. Ulcer severity, angiogenesis, and nitric oxide synthase expression were assessed 3–10 days later. The effects of inhibitors of nitric oxide synthase were also examined. Inducible nitric oxide synthase mRNA was only detected in ulcerated tissue (maximal at day 3), whereas the endothelial isoform mRNA was detected in normal tissue and increased during ulcer healing. Inducible nitric oxide synthase was expressed in inflammatory cells in the ulcer bed, whereas endothelial nitric oxide synthase was found in the vascular endothelium and in some mucosal cells in both normal and ulcerated tissues. Angiogenesis changed in parallel with endothelial nitric oxide synthase expression. N 6-(iminoethyl)-l-lysine did not affect angiogenesis or ulcer healing, while N G-nitro-l-arginine methyl ester significantly reduced both. In conclusion, endothelial nitric oxide synthase, but not the inducible isoform, plays a significant role in gastric ulcer healing.
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Liu, Mengyu, Xiaofeng Liu, Junhua Hu, Yang Xue, and Xiaochun Zhao. "Genetic diversity of limonene synthase genes in Rongan kumquat (Fortunella crassifolia)." Functional Plant Biology 47, no. 5 (2020): 425. http://dx.doi.org/10.1071/fp19051.
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D-limonene is the main component of citrus essential oils. In the monoterpene biosynthetic pathway, geranyl diphosphate reacts with monoterpenes to form the prenyl-carbocation intermediate to produce d-limonene. In this study, d-limonene synthase (FcLS) genes were first isolated from Rongan kumquat (Fortunella crassifolia Swingle). Sequencing analysis revealed that the open reading frames of 18 FcLS genes contain 12 single nucleotide polymorphisms, which resulted in the variation of FcLS proteins, indicating that the limonene synthase genes are a large family in F. crassifolia. This phenomenon has not been reported in Citrus. The predicted FcLS proteins showed a high amino acid sequence identity with other Citrus limonene synthases and also had the typical structures of limonene synthase protein. FcLS1 was validated to be a functional d-limonene synthase by prokaryotic expression.
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Yamada, Yuuki, Tomohisa Kuzuyama, Mamoru Komatsu, Kazuo Shin-ya, Satoshi Omura, David E. Cane, and Haruo Ikeda. "Terpene synthases are widely distributed in bacteria." Proceedings of the National Academy of Sciences 112, no. 3 (December 22, 2014): 857–62. http://dx.doi.org/10.1073/pnas.1422108112.
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Odoriferous terpene metabolites of bacterial origin have been known for many years. In genome-sequencedStreptomycetaceaemicroorganisms, the vast majority produces the degraded sesquiterpene alcohol geosmin. Two minor groups of bacteria do not produce geosmin, with one of these groups instead producing other sesquiterpene alcohols, whereas members of the remaining group do not produce any detectable terpenoid metabolites. Because bacterial terpene synthases typically show no significant overall sequence similarity to any other known fungal or plant terpene synthases and usually exhibit relatively low levels of mutual sequence similarity with other bacterial synthases, simple correlation of protein sequence data with the structure of the cyclized terpene product has been precluded. We have previously described a powerful search method based on the use of hidden Markov models (HMMs) and protein families database (Pfam) search that has allowed the discovery of monoterpene synthases of bacterial origin. Using an enhanced set of HMM parameters generated using a training set of 140 previously identified bacterial terpene synthase sequences, a Pfam search of 8,759,463 predicted bacterial proteins from public databases and in-house draft genome data has now revealed 262 presumptive terpene synthases. The biochemical function of a considerable number of these presumptive terpene synthase genes could be determined by expression in a specially engineered heterologousStreptomyceshost and spectroscopic identification of the resulting terpene products. In addition to a wide variety of terpenes that had been previously reported from fungal or plant sources, we have isolated and determined the complete structures of 13 previously unidentified cyclic sesquiterpenes and diterpenes.
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Moustafa, Amira, and Yoshiaki Habara. "Hydrogen sulfide: a novel gaseous signaling molecule and intracellular Ca2+ regulator in rat parotid acinar cells." American Journal of Physiology-Cell Physiology 309, no. 7 (October 1, 2015): C480—C490. http://dx.doi.org/10.1152/ajpcell.00147.2015.
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In addition to nitric oxide (NO), hydrogen sulfide (H2S) is recognized as a crucial gaseous messenger that exerts many biological actions in various tissues. An attempt was made to assess the roles and underlying mechanisms of both gases in isolated rat parotid acinar cells. Ductal cells and some acinar cells were found to express NO and H2S synthases. Cevimeline, a muscarinic receptor agonist upregulated endothelial NO synthase in parotid tissue. NO and H2S donors increased the intracellular Ca2+ concentration ([Ca2+]i). This was not affected by inhibitors of phospholipase C and inositol 1,4,5-trisphosphate receptors, but was decreased by blockers of ryanodine receptors (RyRs), soluble guanylyl cyclase, and protein kinase G. The H2S donor evoked NO production, which was decreased by blockade of NO synthases or phosphoinositide 3-kinase or by hypotaurine, an H2S scavenger. The H2S donor-induced [Ca2+]i increase was diminished by a NO scavenger or the NO synthases blocker. These results suggest that NO and H2S play important roles in regulating [Ca2+]i via soluble guanylyl cyclase-cGMP-protein kinase G-RyRs, but not via inositol 1,4,5-trisphosphate receptors. The effect of H2S may be partially through NO produced via phosphoinositide 3-kinase-Akt-endothelial NO synthase. It was concluded that both gases regulate [Ca2+]i in a synergistic way, mainly via RyRs in rat parotid acinar cells.
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Guo, Bingbing, Songle Fan, Mingyang Liu, Hong Yang, Longjun Dai, and Lifeng Wang. "ATP Synthase Members of Chloroplasts and Mitochondria in Rubber Trees (Hevea brasiliensis) Response to Plant Hormones." Plants 14, no. 4 (February 17, 2025): 604. https://doi.org/10.3390/plants14040604.
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ATP synthase is a key enzyme in photophosphorylation in photosynthesis and oxidative phosphorylation in respiration, which can catalyze the synthesis of ATP and supply energy to organisms. ATP synthase has been well studied in many animal species but has been poorly characterized in plants. This research identified forty ATP synthase family members in the rubber tree, and the phylogenetic relationship, gene structure, cis-elements, and expression pattern were analyzed. These results indicated that the ATP synthase of mitochondria was divided into three subgroups and the ATP synthase of chloroplast was divided into two subgroups, respectively. ATP synthase in the same subgroup shared a similar gene structure. Evolutionary relationships were consistent with the introns and exons domains, which were highly conserved patterns. A large number of cis elements related to light, phytohormones and stress resistance were present in the promoters of ATP synthase genes in rubber trees, of which the light signal accounts for the most. Transcriptome and qRT-PCR analysis showed that HbATP synthases responded to cold stress and hormone stimulation, and the response to ethylene was most significant. HbMATPR3 was strongly induced by ethylene and salicylic acid, reaching 122-fold and 17-fold, respectively. HbMATP7-1 was 41 times higher than the control after induction by jasmonic acid. These results laid a foundation for further studies on the function of ATP synthase, especially in plant hormone signaling in rubber trees.
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Martín-Urdíroz, Magdalena, M. Isabel G. Roncero, José Antonio González-Reyes, and Carmen Ruiz-Roldán. "ChsVb, a Class VII Chitin Synthase Involved in Septation, Is Critical for Pathogenicity in Fusarium oxysporum." Eukaryotic Cell 7, no. 1 (November 9, 2007): 112–21. http://dx.doi.org/10.1128/ec.00347-07.
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ABSTRACT A new myosin motor-like chitin synthase gene, chsVb, has been identified in the vascular wilt fungus Fusarium oxysporum f. sp. lycopersici. Phylogenetic analysis of the deduced amino acid sequence of the chsVb chitin synthase 2 domain (CS2) revealed that ChsVb belongs to class VII chitin synthases. The ChsVb myosin motor-like domain (MMD) is shorter than the MMD of class V chitin synthases and does not contain typical ATP-binding motifs. Targeted disrupted single (ΔchsVb) and double (ΔchsV ΔchsVb) mutants were unable to infect and colonize tomato plants or grow invasively on tomato fruit tissue. These strains were hypersensitive to compounds that interfere with fungal cell wall assembly, produced lemon-like shaped conidia, and showed swollen balloon-like structures in hyphal subapical regions, thickened walls, aberrant septa, and intrahyphal hyphae. Our results suggest that the chsVb gene is likely to function in polarized growth and confirm the critical importance of cell wall integrity in the complex infection process of this fungus.
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Denny, William A. "Inhibitors of F1F0-ATP synthase enzymes for the treatment of tuberculosis and cancer." Future Medicinal Chemistry 13, no. 10 (May 2021): 911–26. http://dx.doi.org/10.4155/fmc-2021-0010.
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The spectacular success of the mycobacterial F1F0-ATP synthase inhibitor bedaquiline for the treatment of drug-resistant tuberculosis has generated wide interest in the development of other inhibitors of this enzyme. Work in this realm has included close analogues of bedaquiline with better safety profiles and ‘bedaquiline-like’ compounds, some of which show potent antibacterial activity in vitro although none have yet progressed to clinical trials. The search has lately extended to a range of new scaffolds as potential inhibitors, including squaramides, diaminoquinazolines, chloroquinolines, dihydropyrazolo[1,5-a]pyrazin-4-ones, thiazolidinediones, diaminopyrimidines and tetrahydroquinolines. Because of the ubiquitous expression of ATP synthase enzymes, there has also been interest in inhibitors of other bacterial ATP synthases, as well as inhibitors of human mitochondrial ATP synthase for cancer therapy. The latter encompass both complex natural products and simpler small molecules. The review seeks to demonstrate the breadth of the structural types of molecules able to effectively inhibit the function of variants of this intriguing enzyme.
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Mühleip, Alexander W., Friederike Joos, Christoph Wigge, Achilleas S. Frangakis, Werner Kühlbrandt, and Karen M. Davies. "Helical arrays of U-shaped ATP synthase dimers form tubular cristae in ciliate mitochondria." Proceedings of the National Academy of Sciences 113, no. 30 (July 11, 2016): 8442–47. http://dx.doi.org/10.1073/pnas.1525430113.
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F1Fo-ATP synthases are universal energy-converting membrane protein complexes that synthesize ATP from ADP and inorganic phosphate. In mitochondria of yeast and mammals, the ATP synthase forms V-shaped dimers, which assemble into rows along the highly curved ridges of lamellar cristae. Using electron cryotomography and subtomogram averaging, we have determined the in situ structure and organization of the mitochondrial ATP synthase dimer of the ciliate Paramecium tetraurelia. The ATP synthase forms U-shaped dimers with parallel monomers. Each complex has a prominent intracrista domain, which links the c-ring of one monomer to the peripheral stalk of the other. Close interaction of intracrista domains in adjacent dimers results in the formation of helical ATP synthase dimer arrays, which differ from the loose dimer rows in all other organisms observed so far. The parameters of the helical arrays match those of the cristae tubes, suggesting the unique features of the P. tetraurelia ATP synthase are directly responsible for generating the helical tubular cristae. We conclude that despite major structural differences between ATP synthase dimers of ciliates and other eukaryotes, the formation of ATP synthase dimer rows is a universal feature of mitochondria and a fundamental determinant of cristae morphology.
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Durkin, Colleen A., Thomas Mock, and E. Virginia Armbrust. "Chitin in Diatoms and Its Association with the Cell Wall." Eukaryotic Cell 8, no. 7 (May 8, 2009): 1038–50. http://dx.doi.org/10.1128/ec.00079-09.
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ABSTRACT Chitin is a globally abundant polymer widely distributed throughout eukaryotes that has been well characterized in only a few lineages. Diatoms are members of the eukaryotic lineage of stramenopiles. Of the hundreds of diatom genera, two produce long fibers of chitin that extrude through their cell walls of silica. We identify and describe here genes encoding putative chitin synthases in a variety of additional diatom genera, indicating that the ability to produce chitin is more widespread and likely plays a more central role in diatom biology than previously considered. Diatom chitin synthases fall into four phylogenetic clades. Protein domain predictions and differential gene expression patterns provide evidence that chitin synthases have multiple functions within a diatom cell. Thalassiosira pseudonana possesses six genes encoding three types of chitin synthases. Transcript abundance of the gene encoding one of these chitin synthase types increases when cells resume division after short-term silicic acid starvation and during short-term limitation by silicic acid or iron, two nutrient conditions connected in the environment and known to affect the cell wall. During long-term silicic acid starvation transcript abundance of this gene and one additional chitin synthase gene increased at the same time a chitin-binding lectin localized to the girdle band region of the cell wall. Together, these results suggest that the ability to produce chitin is more widespread in diatoms than previously thought and that a subset of the chitin produced by diatoms is associated with the cell wall.
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Hao, Jijun, Willie F. Vann, Stephan Hinderlich, and Munirathinam Sundaramoorthy. "Elimination of 2-keto-3-deoxy-D-glycero-D-galacto-nonulosonic acid 9-phosphate synthase activity from human N-acetylneuraminic acid 9-phosphate synthase by a single mutation." Biochemical Journal 397, no. 1 (June 14, 2006): 195–201. http://dx.doi.org/10.1042/bj20052034.
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The most commonly occurring sialic acid Neu5Ac (N-acetylneuraminic acid) and its deaminated form, KDN (2-keto-3-deoxy-D-glycero-D-galacto-nonulosonic acid), participate in many biological functions. The human Neu5Ac-9-P (Neu5Ac 9-phosphate) synthase has the unique ability to catalyse the synthesis of not only Neu5Ac-9-P but also KDN-9-P (KDN 9-phosphate). Both reactions are catalysed by the mechanism of aldol condensation of PEP (phosphoenolpyruvate) with sugar substrates, ManNAc-6-P (N-acetylmannosamine 6-phosphate) or Man-6-P (mannose 6-phosphate). Mouse and putative rat Neu5Ac-9-P synthases, however, do not show KDN-9-P synthase activity, despite sharing high sequence identity (>95%) with the human enzyme. Here, we demonstrate that a single mutation, M42T, in human Neu5Ac-9-P synthase can abolish the KDN-9-P synthase activity completely without compromising the Neu5Ac-9-P synthase activity. Saturation mutagenesis of Met42 of the human Neu5Ac-9-P synthase showed that the substitution with all amino acids except leucine retains only the Neu5Ac-9-P synthase activity at levels comparable with the wild-type enzyme. The M42L mutant, like the wild-type enzyme, showed the additional KDN-9-P synthase activity. In the homology model of human Neu5Ac-9-P synthase, Met42 is located 22 Å (1 Å=0.1 nm) away from the substrate-binding site and the impact of this distant residue on the enzyme functions is discussed.
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Daiyasu, Hiromi, Kei-Ichi Kuma, Toshiro Yokoi, Hiroyuki Morii, Yosuke Koga, and Hiroyuki Toh. "A study of archaeal enzymes involved in polar lipid synthesis linking amino acid sequence information, genomic contexts and lipid composition." Archaea 1, no. 6 (2005): 399–410. http://dx.doi.org/10.1155/2005/452563.
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Cellular membrane lipids, of which phospholipids are the major constituents, form one of the characteristic features that distinguish Archaea from other organisms. In this study, we focused on the steps in archaeal phospholipid synthetic pathways that generate polar lipids such as archaetidylserine, archaetidylglycerol, and archaetidylinositol. Only archaetidylserine synthase (ASS), fromMethanothermobacter thermautotrophicus, has been experimentally identified. Other enzymes have not been fully examined. Through database searching, we detected many archaeal hypothetical proteins that show sequence similarity to members of the CDP alcohol phosphatidyltransferase family, such as phosphatidylserine synthase (PSS), phosphatidylglycerol synthase (PGS) and phosphatidylinositol synthase (PIS) derived from Bacteria and Eukarya. The archaeal hypothetical proteins were classified into two groups, based on the sequence similarity. Members of the first group, including ASS fromM. thermautotrophicus, were closely related to PSS. The rough agreement between PSS homologue distribution within Archaea and the experimentally identified distribution of archaetidylserine suggested that the hypothetical proteins are ASSs. We found that an open reading frame (ORF) tends to be adjacent to that of ASS in the genome, and that the order of the two ORFs is conserved. The sequence similarity of phosphatidylserine decarboxylase to the product of the ORF next to the ASS gene, together with the genomic context conservation, suggests that the ORF encodes archaetidylserine decarboxylase, which may transform archaetidylserine to archaetidylethanolamine. The second group of archaeal hypothetical proteins was related to PGS and PIS. The members of this group were subjected to molecular phylogenetic analysis, together with PGSs and PISs and it was found that they formed two distinct clusters in the molecular phylogenetic tree. The distribution of members of each cluster within Archaea roughly corresponded to the experimentally identified distribution of archaetidylglycerol or archaetidylinositol. The molecular phylogenetic tree patterns and the correspondence to the membrane compositions suggest that the two clusters in this group correspond to archaetidylglycerol synthases and archaetidylinositol synthases. No archaeal hypothetical protein with sequence similarity to known phosphatidylcholine synthases was detected in this study.
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Mülsch, A., A. Vanin, P. Mordvintcev, S. Hauschildt, and R. Busse. "NO accounts completely for the oxygenated nitrogen species generated by enzymic l-arginine oxygenation." Biochemical Journal 288, no. 2 (December 1, 1992): 597–603. http://dx.doi.org/10.1042/bj2880597.
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We have assessed the stoichiometry of the nitric oxide (NO) synthase reaction by using a novel e.p.r. technique. NO generated by crude and partially purified NO synthase from endothelial cells and Escherichia coli-lipopolysaccharide-activated macrophages was trapped by a ferrous diethyldithiocarbamate complex dispersed in yeast. The paramagnetic ferrous mononitrosyl dithiocarbamate complex formed exhibited a characteristic e.p.r. signal at g perpendicular = 2.035 and g parallel = 2.02 with a triplet hyperfine structure (hfs) at g perpendicular. NO, 3-morpholinosydnonimine and S-nitroso-L-cysteine, but not nitrite or hydroxylamine, generated a similar e.p.r. signal. NO generated by NO synthase and by SIN-1 accumulated at a constant rate for 1 h, as measured by continuous e.p.r. registration at 37 degrees C. The formation of e.p.r.-detectable NO by NO synthases was inhibited by NG-nitro-L-arginine. Incubation with [15N]NG-L-arginine caused an e.p.r. signal with doublet hfs, indicating that the nitrosyl nitrogen derived exclusively from the guanidino nitrogen. The amount of NO generated by NO synthase as measured by e.p.r. technique was compared with formation of L-[3H]citrulline from L-[3H]arginine. NO and L-citrulline were detected at a 1:1 ratio with both NO synthase preparations. GSH and thiol depletion did not significantly affect NO synthase activity, excluding S-nitrosothiols as intermediates in the NO synthase reaction. We conclude that NO fully accounts for the immediate oxygenated nitrogen species derived from the enzymic oxygenation of L-arginine.
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Holtzapple, Erik, and Claudia Schmidt-Dannert. "Biosynthesis of Isoprenoid Wax Ester in Marinobacter hydrocarbonoclasticus DSM 8798: Identification and Characterization of Isoprenoid Coenzyme A Synthetase and Wax Ester Synthases." Journal of Bacteriology 189, no. 10 (March 9, 2007): 3804–12. http://dx.doi.org/10.1128/jb.01932-06.
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ABSTRACT Marinobacter hydrocarbonoclasticus DSM 8798 has been reported to synthesize isoprenoid wax ester storage compounds when grown on phytol as the sole carbon source under limiting nitrogen and/or phosphorous conditions. We hypothesized that isoprenoid wax ester synthesis involves (i) activation of an isoprenoid fatty acid by a coenzyme A (CoA) synthetase and (ii) ester bond formation between an isoprenoid alcohol and isoprenoyl-CoA catalyzed, most likely, by an isoprenoid wax ester synthase similar to an acyl wax ester synthase, wax ester synthase/diacylglycerol acyltransferase (WS/DGAT), recently described from Acinetobacter sp. strain ADP1. We used the recently released rough draft genome sequence of a closely related strain, M. aquaeolei VT8, to search for WS/DGAT and acyl-CoA synthetase candidate genes. The sequence information from putative WS/DGAT and acyl-CoA synthetase genes identified in this strain was used to clone homologues from the isoprenoid wax ester synthesizing Marinobacter strain. The activities of the recombinant enzymes were characterized, and two new isoprenoid wax ester synthases capable of synthesizing isoprenoid ester and acyl/isoprenoid hybrid ester in vitro were identified along with an isoprenoid-specific CoA synthetase. One of the Marinobacter wax ester synthases displays several orders of magnitude higher activity toward acyl substrates than any previously characterized acyl-WS and may reflect adaptations to available carbon sources in their environments.
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Loke, Paxton, and Tiow-Suan Sim. "Molecular cloning, heterologous expression, and functional characterisation of a malate synthase gene fromStreptomyces coelicolorA3(2)." Canadian Journal of Microbiology 46, no. 8 (August 1, 2000): 764–69. http://dx.doi.org/10.1139/w00-044.
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With the rapid generation of genetic information from the Streptomyces coelicolor genome project, deciphering the relevant gene products is critical for understanding the genetics of this model streptomycete. A putative malate synthase gene (aceB) from S. coelicolor A3(2) was identified by homology-based analysis, cloned by polymerase chain reaction, and fully sequenced on both strands. The putative malate synthase from S. coelicolor has an amino acid identity of 77% with the malate synthase of S. clavuligerus, and possesses an open reading frame which codes for a protein of 540 amino acids. In order to establish the identity of this gene, the putative aceB clones were subcloned into the expression vector pET24a, and heterologously expressed in Escherichia coli BL21(DE3). Soluble cell-free extracts containing the recombinant putative malate synthase exhibited a specific activity of 1623 (nmol·mg-1·min-1), which is an increment of 92-fold compared to the non-recombinant control. Thus, the gene product was confirmed to be a malate synthase. Interestingly, the specific activity of S. coelicolor malate synthase was found to be almost 8-fold higher than the specific activity of S. clavuligerus malate synthase under similar expression conditions. Furthermore, the genomic organisation of the three Streptomyces aceB genes cloned thus far is different from that of other bacterial malate synthases, and warrants further investigation.Key words: primary metabolism, polymerase chain reaction, glyoxylate pathway.
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Grabińska, Kariona A., Sudip K. Ghosh, Ziqiang Guan, Jike Cui, Christian R. H. Raetz, Phillips W. Robbins, and John Samuelson. "Dolichyl-Phosphate-Glucose Is Used To Make O-Glycans on Glycoproteins of Trichomonas vaginalis." Eukaryotic Cell 7, no. 8 (June 13, 2008): 1344–51. http://dx.doi.org/10.1128/ec.00061-08.
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ABSTRACT Trichomonas vaginalis, the protist that causes vaginal itching, has a huge genome with numerous gene duplications. Recently we found that Trichomonas has numerous genes encoding putative dolichyl-phosphate-glucose (Dol-P-Glc) synthases (encoded by ALG5 genes) despite the fact that Trichomonas lacks the glycosyltransferases (encoded by ALG6, ALG8, and ALG10 genes) that use Dol-P-Glc to glucosylate dolichyl-PP-linked glycans. In addition, Trichomonas does not have a canonical DPM1 gene, encoding a dolichyl-P-mannose (Dol-P-Man) synthase. Here we show Trichomonas membranes have roughly 300 times the Dol-P-Glc synthase activity of Saccharomyces cerevisiae membranes and about one-fifth the Dol-P-Man synthase activity of Saccharomyces membranes. Endogenous Dol-P-hexoses of Trichomonas are relatively abundant and contain 16 isoprene units. Five paralogous Trichomonas ALG5 gene products have Dol-P-Glc synthase activity when expressed as recombinant proteins, and these Trichomonas Alg5s correct a carboxypeptidase N glycosylation defect in a Saccharomyces alg5 mutant in vivo. A recombinant Trichomonas Dpm1, which is deeply divergent in its sequence, has Dol-P-Man synthase activity. When radiolabeled Dol-P-Glc is incubated with Trichomonas membranes, Glc is incorporated into reducing and nonreducing sugars of O-glycans of endogenous glycoproteins. To our knowledge, this is the first demonstration of Dol-P-Glc as a sugar donor for O-glycans on glycoproteins.
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Mohammadi, Malihe, Mona Atabakhshi Kashi, Shekufeh Zareian, Manoochehr Mirshahi, and Khosro Khajeh. "Remarkable Improvement of Methylglyoxal Synthase Thermostability by His–His Interaction." Applied Biochemistry and Biotechnology 172, no. 1 (September 21, 2013): 157–67. http://dx.doi.org/10.1007/s12010-013-0404-y.
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Kotowska, Magdalena, Krzysztof Pawlik, Aleksandra Smulczyk-Krawczyszyn, Hubert Bartosz-Bechowski, and Katarzyna Kuczek. "Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate." Applied and Environmental Microbiology 75, no. 4 (December 12, 2008): 887–96. http://dx.doi.org/10.1128/aem.01371-08.
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ABSTRACT Type II thioesterases (TE IIs) were shown to maintain the efficiency of polyketide synthases (PKSs) by removing acyl residues blocking extension modules. However, the substrate specificity and kinetic parameters of these enzymes differ, which may have significant consequences when they are included in engineered hybrid systems for the production of novel compounds. Here we show that thioesterase ScoT associated with polyketide synthase Cpk from Streptomyces coelicolor A3(2) is able to hydrolyze acetyl, propionyl, and butyryl residues, which is consistent with its editing function. This enzyme clearly prefers propionate, in contrast to the TE IIs tested previously, and this indicates that it may have a role in control of the starter unit. We also determined activities of ScoT mutants and concluded that this enzyme is an α/β hydrolase with Ser90 and His224 in its active site.
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Tracey, W. R., J. S. Pollock, F. Murad, M. Nakane, and U. Forstermann. "Identification of an endothelial-like type III NO synthase in LLC-PK1 kidney epithelial cells." American Journal of Physiology-Cell Physiology 266, no. 1 (January 1, 1994): C22—C28. http://dx.doi.org/10.1152/ajpcell.1994.266.1.c22.
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Porcine kidney tubular epithelial cells (LLC-PK1) produce nitric oxide or a related compound (e.g., a nitrosothiol) after stimulation with various agonists. We now report the identification and characterization of a constitutive, particulate nitric oxide (NO) synthase from LLC-PK1 cells. After partial purification on adenosine 2',5'-bisphosphate-Sepharose, the particulate NO synthase activity eluted anomalously from Superose 6 gel permeation columns near the total included volume, similar to that observed for the endothelial (type III) NO synthase. Substrate/cofactor requirements of the epithelial and endothelial NO synthases were identical, i.e., dependency on L-arginine, (6R)-5,6,7,8-tetrahydrobiopterin, FAD, calcium and calmodulin. The epithelial enzyme activity was inhibited by the arginine analogues, NG-methyl-L-arginine (100 microM) and NG-nitro-L-arginine (100 microM), as well as the calmodulin antagonists, trifluoperazine (100 microM) and calmidazolium (30 microM). Anti-type III (H32), but not anti-type I (brain, 6763-5) or anti-type II (macrophage, 8196) NO synthase antibodies, detected a single immunoreactive band in the LLC-PK1 particulate fraction of approximately 140 kDa by Western blot analysis. Finally, the presence of type III NO synthase mRNA in LLC-PK1 cells was demonstrated using the polymerase chain reaction. These data indicate that LLC-PK1 kidney epithelial cells contain type III NO synthase, which has been classically associated with the vascular endothelium.
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Sutton, Kristin A., Jennifer Breen, Thomas A. Russo, L. Wayne Schultz, and Timothy C. Umland. "Crystal structure of 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase from the ESKAPE pathogenAcinetobacter baumannii." Acta Crystallographica Section F Structural Biology Communications 72, no. 3 (February 16, 2016): 179–87. http://dx.doi.org/10.1107/s2053230x16001114.
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The enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase catalyzes the sixth step of the seven-step shikimate pathway. Chorismate, the product of the pathway, is a precursor for the biosynthesis of aromatic amino acids, siderophores and metabolites such as folate, ubiquinone and vitamin K. The shikimate pathway is present in bacteria, fungi, algae, plants and apicomplexan parasites, but is absent in humans. The EPSP synthase enzyme produces 5-enolpyruvylshikimate 3-phosphate and phosphate from phosphoenolpyruvate and shikimate 3-phosphateviaa transferase reaction, and is the target of the herbicide glyphosate. TheAcinetobacter baumanniigene encoding EPSP synthase,aroA, has previously been demonstrated to be essential during host infection for the growth and survival of this clinically important drug-resistant ESKAPE pathogen. Prephenate dehydrogenase is also encoded by the bifunctionalA. baumannii aroAgene, but its activity is dependent upon EPSP synthase since it operates downstream of the shikimate pathway. As part of an effort to evaluate new antimicrobial targets, recombinantA. baumanniiEPSP (AbEPSP) synthase, comprising residues Ala301–Gln756 of thearoAgene product, was overexpressed inEscherichia coli, purified and crystallized. The crystal structure, determined to 2.37 Å resolution, is described in the context of a potential antimicrobial target and in comparison to EPSP synthases that are resistant or sensitive to the herbicide glyphosate.
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Gojković, Zoran, Michael P. B. Sandrini, and Jure Piškur. "Eukaryotic β-Alanine Synthases Are Functionally Related but Have a High Degree of Structural Diversity." Genetics 158, no. 3 (July 1, 2001): 999–1011. http://dx.doi.org/10.1093/genetics/158.3.999.
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Abstract β-Alanine synthase (EC 3.5.1.6), which catalyzes the final step of pyrimidine catabolism, has only been characterized in mammals. A Saccharomyces kluyveri pyd3 mutant that is unable to grow on N-carbamyl-β-alanine as the sole nitrogen source and exhibits diminished β-alanine synthase activity was used to clone analogous genes from different eukaryotes. Putative PYD3 sequences from the yeast S. kluyveri, the slime mold Dictyostelium discoideum, and the fruit fly Drosophila melanogaster complemented the pyd3 defect. When the S. kluyveri PYD3 gene was expressed in S. cerevisiae, which has no pyrimidine catabolic pathway, it enabled growth on N-carbamyl-β-alanine as the sole nitrogen source. The D. discoideum and D. melanogaster PYD3 gene products are similar to mammalian β-alanine synthases. In contrast, the S. kluyveri protein is quite different from these and more similar to bacterial N-carbamyl amidohydrolases. All three β-alanine synthases are to some degree related to various aspartate transcarbamylases, which catalyze the second step of the de novo pyrimidine biosynthetic pathway. PYD3 expression in yeast seems to be inducible by dihydrouracil and N-carbamyl-β-alanine, but not by uracil. This work establishes S. kluyveri as a model organism for studying pyrimidine degradation and β-alanine production in eukaryotes.
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Cheuk, Anthony, and Thomas Meier. "Rotor subunits adaptations in ATP synthases from photosynthetic organisms." Biochemical Society Transactions 49, no. 2 (April 23, 2021): 541–50. http://dx.doi.org/10.1042/bst20190936.
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Driven by transmembrane electrochemical ion gradients, F-type ATP synthases are the primary source of the universal energy currency, adenosine triphosphate (ATP), throughout all domains of life. The ATP synthase found in the thylakoid membranes of photosynthetic organisms has some unique features not present in other bacterial or mitochondrial systems. Among these is a larger-than-average transmembrane rotor ring and a redox-regulated switch capable of inhibiting ATP hydrolysis activity in the dark by uniquely adapted rotor subunit modifications. Here, we review recent insights into the structure and mechanism of ATP synthases specifically involved in photosynthesis and explore the cellular physiological consequences of these adaptations at short and long time scales.
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Abu-Zaitoon, Yousef M., Ezz Al-Dein Muhammed Al-Ramamneh, Abdel Rahman Al Tawaha, Sulaiman M. Alnaimat, and Fouad A. Almomani. "Comparative Coexpression Analysis of Indole Synthase and Tryptophan Synthase A Reveals the Independent Production of Auxin via the Cytosolic Free Indole." Plants 12, no. 8 (April 18, 2023): 1687. http://dx.doi.org/10.3390/plants12081687.
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Indole synthase (INS), a homologous cytosolic enzyme of the plastidal tryptophan synthase A (TSA), has been reported as the first enzyme in the tryptophan-independent pathway of auxin synthesis. This suggestion was challenged as INS or its free indole product may interact with tryptophan synthase B (TSB) and, therefore, with the tryptophan-dependent pathway. Thus, the main aim of this research was to find out whether INS is involved in the tryptophan-dependent or independent pathway. The gene coexpression approach is widely recognized as an efficient tool to uncover functionally related genes. Coexpression data presented here were supported by both RNAseq and microarray platforms and, hence, considered reliable. Coexpression meta-analyses of Arabidopsis genome was implemented to compare between the coexpression of TSA and INS with all genes involved in the production of tryptophan via the chorismate pathway. Tryptophan synthase A was found to be coexpressed strongly with TSB1/2, anthranilate synthase A1/B1, phosphoribosyl anthranilate transferase1, as well as indole-3-glycerol phosphate synthase1. However, INS was not found to be coexpressed with any target genes suggesting that it may exclusively and independently be involved in the tryptophan-independent pathway. Additionally, annotation of examined genes as ubiquitous or differentially expressed were described and subunits-encoded genes available for the assembly of tryptophan and anthranilate synthase complex were suggested. The most probable TSB subunits expected to interact with TSA is TSB1 then TSB2. Whereas TSB3 is only used under limited hormone conditions to assemble tryptophan synthase complex, putative TSB4 is not expected to be involved in the plastidial synthesis of tryptophan in Arabidopsis.
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Goto, Saki, Yuki Miyahara, Seiichi Taguchi, Takeharu Tsuge, and Ayaka Hiroe. "Enhanced Production of (R)-3-Hydroxybutyrate Oligomers by Coexpression of Molecular Chaperones in Recombinant Escherichia coli Harboring a Polyhydroxyalkanoate Synthase Derived from Bacillus cereus YB-4." Microorganisms 10, no. 2 (February 16, 2022): 458. http://dx.doi.org/10.3390/microorganisms10020458.
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The biodegradable polyester poly-(R)-3-hydroxybutyrate [P(3HB)] is synthesized by a polymerizing enzyme called polyhydroxyalkanoate (PHA) synthase and accumulates in a wide variety of bacterial cells. Recently, we demonstrated the secretory production of a (R)-3HB oligomer (3HBO), a low-molecular-weight P(3HB), by using recombinant Escherichia coli expressing PHA synthases. The 3HBO has potential value as an antibacterial substance and as a building block for various polymers. In this study, to construct an efficient 3HBO production system, the coexpression of molecular chaperones and a PHA synthase derived from Bacillus cereus YB-4 (PhaRCYB4) was examined. First, genes encoding enzymes related to 3HBO biosynthesis (phaRCYB4, phaA and phaB derived from Ralstonia eutropha H16) and two types of molecular chaperones (groEL, groES, and tig) were introduced into the E. coli strains BW25113 and BW25113ΔadhE. As a result, coexpression of the chaperones promoted the enzyme activity of PHA synthase (approximately 2–3-fold) and 3HBO production (approximately 2-fold). The expression assay of each chaperone and PHA synthase subunit (PhaRYB4 and PhaCYB4) indicated that the combination of the two chaperone systems (GroEL-GroES and TF) supported the folding of PhaRYB4 and PhaCYB4. These results suggest that the utilization of chaperone proteins is a valuable approach to enhance the formation of active PHA synthase and the productivity of 3HBO.
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Sun, Lijing, Mengyun Hu, Jie Zhao, Liangjie Lv, Yelun Zhang, Qian Liu, Li Zhang, et al. "Molecular Characteristics, Synthase, and Food Application of Cereal β-Glucan." Journal of Food Quality 2021 (March 8, 2021): 1–8. http://dx.doi.org/10.1155/2021/6682014.
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Cereal β-glucan is a type of valuable dietary fiber that mainly exists in the aleurone, subaleurone, and endosperm of some cereal grains. β-Glucan is acknowledged as a functional food ingredient owing to its multiple health benefits, including the prevention of diabetes, reduction in the incidence of cardiovascular disease, antitumor effects, antioxidant activities, and immunostimulation. It is well documented that cellulose synthase-like CslF/H/J genes encode synthases responsible for β-glucan biosynthesis in cereal grains. Recently, β-glucan has been widely applied as an emulsion stabilizer, thickening agent, fat substitute, and bioactive ingredient in the food industry due to its water solubility, viscosity, gelation property, and health benefits. Therefore, the present paper aims to review the molecular characteristics, synthase gene family, and food application of cereal β-glucan in recent years.
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Ee, Su-Fang, Zeti-Azura Mohamed-Hussein, Roohaida Othman, Noor Azmi Shaharuddin, Ismanizan Ismail, and Zamri Zainal. "Functional Characterization of Sesquiterpene Synthase fromPolygonum minus." Scientific World Journal 2014 (2014): 1–11. http://dx.doi.org/10.1155/2014/840592.
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Polygonum minusis an aromatic plant, which contains high abundance of terpenoids, especially the sesquiterpenes C15H24. Sesquiterpenes were believed to contribute to the many useful biological properties in plants. This study aimed to functionally characterize a full length sesquiterpene synthase gene fromP. minus.P. minussesquiterpene synthase (PmSTS) has a complete open reading frame (ORF) of 1689 base pairs encoding a 562 amino acid protein. Similar to other sesquiterpene synthases, PmSTS has two large domains: the N-terminal domain and the C-terminal metal-binding domain. It also consists of three conserved motifs: the DDXXD, NSE/DTE, and RXR. A three-dimensional protein model for PmSTS built clearly distinguished the two main domains, where conserved motifs were highlighted. We also constructed a phylogenetic tree, which showed that PmSTS belongs to the angiosperm sesquiterpene synthase subfamily Tps-a. To examine the function ofPmSTS, we expressed this gene inArabidopsis thaliana. Two transgenic lines, designated asOE3andOE7, were further characterized, both molecularly and functionally. The transgenic plants demonstrated smaller basal rosette leaves, shorter and fewer flowering stems, and fewer seeds compared to wild type plants. Gas chromatography-mass spectrometry analysis of the transgenic plants showed that PmSTS was responsible for the production ofβ-sesquiphellandrene.
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Bimmer, Martin, Wolfgang Liebl, and Armin Ehrenreich. "Bakterielle Cellulose — ein Netzwerk gestaltet von drei Cellulosesynthasen." BIOspektrum 29, no. 2 (March 2023): 218–20. http://dx.doi.org/10.1007/s12268-023-1908-9.
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AbstractBesides plants also some bacterial genera are able to synthesize cellulose in remarkably high quantities. Bacterial cellulose from the acetic acid bacterium Komagataeibacter hansenii has a big advantage supporting its use as multifunctional and sustainable material — it is free of non-cellulosic components, unlike cellulose of plant origin. Based on marker-free in frame deletions, we propose a model where cellulose fibers released by the main cellulose synthase (BcsAB1) are modified by two additional cellulose synthases.
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Varghese, Febin, James N. Blaza, Andrew J. Y. Jones, Owen D. Jarman, and Judy Hirst. "Deleting the IF 1 -like ζ subunit from Paracoccus denitrificans ATP synthase is not sufficient to activate ATP hydrolysis." Open Biology 8, no. 1 (January 2018): 170206. http://dx.doi.org/10.1098/rsob.170206.
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In oxidative phosphorylation, ATP synthases interconvert two forms of free energy: they are driven by the proton-motive force across an energy-transducing membrane to synthesize ATP and displace the ADP/ATP ratio from equilibrium. For thermodynamically efficient energy conversion they must be reversible catalysts. However, in many species ATP synthases are unidirectional catalysts (their rates of ATP hydrolysis are negligible), and in others mechanisms have evolved to regulate or minimize hydrolysis. Unidirectional catalysis by Paracoccus denitrificans ATP synthase has been attributed to its unique ζ subunit, which is structurally analogous to the mammalian inhibitor protein IF 1 . Here, we used homologous recombination to delete the ζ subunit from the P. denitrificans genome, and compared ATP synthesis and hydrolysis by the wild-type and knockout enzymes in inverted membrane vesicles and the F 1 -ATPase subcomplex. ATP synthesis was not affected by loss of the ζ subunit, and the rate of ATP hydrolysis increased by less than twofold, remaining negligible in comparison with the rates of the Escherichia coli and mammalian enzymes. Therefore, deleting the P. denitrificans ζ subunit is not sufficient to activate ATP hydrolysis. We close by considering our conclusions in the light of reversible catalysis and regulation in ATP synthase enzymes.
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Caballero Cerbon, Daniel Alejandro, Jeremias Widmann, and Dirk Weuster-Botz. "Metabolic control analysis enabled the improvement of the L-cysteine production process with Escherichia coli." Applied Microbiology and Biotechnology 108, no. 1 (January 11, 2024): 1–13. http://dx.doi.org/10.1007/s00253-023-12928-z.
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Abstract L-cysteine is an amino acid with relevance to the pharmaceutical, food, feed, and cosmetic industry. The environmental and societal impact of its chemical production has led to the development of more sustainable fermentative L-cysteine production processes with engineered E. coli based on glucose and thiosulfate as sulphur source. Still, most of the published processes show low yields. For the identification of further metabolic engineering targets, engineered E. coli cells were withdrawn from a fed-batch production process, followed by in vivo metabolic control analysis (MCA) based on the data of short-term perturbation experiments, metabolomics (LC–MS), and thermodynamic flux analysis (TFA). In vivo MCA indicated that the activities of the L-cysteine synthases of the cells withdrawn from the production process might be limiting, and we hypothesised that the L-cysteine precursor O-acetylserine (OAS) might be exported from the cells faster than it took to transform OAS into L-cysteine. By increasing the expression of the L-cysteine synthases, either sulfocysteine synthase or L-cysteine synthase, which transform OAS into L-cysteine, an improvement of up to 70% in specific L-cysteine productivity and up to 47% in the final L-cysteine concentration was achieved in standardised fed-batch processes thereby increasing the yield on glucose by more than 85 to 9.2% (w/w). Key points • Metabolic control analysis was applied to analyse L-cysteine production with E. coli • OAS export was faster than its transformation to L-cysteine • Overexpression of L-cysteine synthases improved L-cysteine productivity and yield
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Ker, De-Sheng, Sze Lei Pang, Noor Farhan Othman, Sekar Kumaran, Ee Fun Tan, Thiba Krishnan, Kok Gan Chan, Roohaida Othman, Maizom Hassan, and Chyan Leong Ng. "Purification and biochemical characterization of recombinant Persicaria minor β-sesquiphellandrene synthase." PeerJ 5 (February 28, 2017): e2961. http://dx.doi.org/10.7717/peerj.2961.
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Background Sesquiterpenes are 15-carbon terpenes synthesized by sesquiterpene synthases using farnesyl diphosphate (FPP) as a substrate. Recently, a sesquiterpene synthase gene that encodes a 65 kDa protein was isolated from the aromatic plant Persicaria minor. Here, we report the expression, purification and characterization of recombinant P. minor sesquiterpene synthase protein (PmSTS). Insights into the catalytic active site were further provided by structural analysis guided by multiple sequence alignment. Methods The enzyme was purified in two steps using affinity and size exclusion chromatography. Enzyme assays were performed using the malachite green assay and enzymatic product was identified using gas chromatography-mass spectrometry (GC-MS) analysis. Sequence analysis of PmSTS was performed using multiple sequence alignment (MSA) against plant sesquiterpene synthase sequences. The homology model of PmSTS was generated using I-TASSER server. Results Our findings suggest that the recombinant PmSTS is mainly expressed as inclusion bodies and soluble aggregate in the E. coli protein expression system. However, addition of 15% (v/v) glycerol to the protein purification buffer and removal of N-terminal 24 amino acids of PmSTS helped to produce homogenous recombinant protein. Enzyme assay showed that recombinant PmSTS is active and specific to the C15 substrate FPP. The optimal temperature and pH for the recombinant PmSTS are 30 °C and pH 8.0, respectively. The GC-MS analysis further showed that PmSTS produces β-sesquiphellandrene as a major product and β-farnesene as a minor product. MSA analysis revealed that PmSTS adopts a modified conserved metal binding motif (NSE/DTE motif). Structural analysis suggests that PmSTS may binds to its substrate similarly to other plant sesquiterpene synthases. Discussion The study has revealed that homogenous PmSTS protein can be obtained with the addition of glycerol in the protein buffer. The N-terminal truncation dramatically improved the homogeneity of PmSTS during protein purification, suggesting that the disordered N-terminal region may have caused the formation of soluble aggregate. We further show that the removal of the N-terminus disordered region of PmSTS does not affect the product specificity. The optimal temperature, optimal pH, Km and kcat values of PmSTS suggests that PmSTS shares similar enzyme characteristics with other plant sesquiterpene synthases. The discovery of an altered conserved metal binding motif in PmSTS through MSA analysis shows that the NSE/DTE motif commonly found in terpene synthases is able to accommodate certain level of plasticity to accept variant amino acids. Finally, the homology structure of PmSTS that allows good fitting of substrate analog into the catalytic active site suggests that PmSTS may adopt a sesquiterpene biosynthesis mechanism similar to other plant sesquiterpene synthases.
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Krishnan, Hari B., Won-Seok Kim, Jeong Sun-Hyung, Kil Yong Kim, and Guoqiao Jiang. "Citrate Synthase Mutants of Sinorhizobium fredii USDA257 Form Ineffective Nodules with Aberrant Ultrastructure." Applied and Environmental Microbiology 69, no. 6 (June 2003): 3561–68. http://dx.doi.org/10.1128/aem.69.6.3561-3568.2003.
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ABSTRACT The tricarboxylic acid (TCA) cycle plays an important role in generating the energy required by bacteroids to fix atmospheric nitrogen. Citrate synthase is the first enzyme that controls the entry of carbon into the TCA cycle. We cloned and determined the nucleotide sequence of the gltA gene that encodes citrate synthase in Sinorhizobium fredii USDA257, a symbiont of soybeans (Glycine max [L.] Merr.) and several other legumes. The deduced citrate synthase protein has a molecular weight of 48,198 and exhibits sequence similarity to citrate synthases from several bacterial species, including Sinorhizobium meliloti and Rhizobium tropici. Southern blot analysis revealed that the fast-growing S. fredii strains and Rhizobium sp. strain NGR234 contained a single copy of the gene located in the bacterial chromosome. S. fredii USDA257 gltA mutant HBK-CS1, which had no detectable citrate synthase activity, had diminished nodulation capacity and produced ineffective nodules on soybean. Light and electron microscopy observations revealed that the nodules initiated by HBK-CS1 contained very few bacteroids. The infected cells contained large vacuoles and prominent starch grains. Within the vacuoles, membrane structures that appeared to be reminiscent of disintegrating bacteroids were detected. The citrate synthase mutant had altered cell surface characteristics and produced three times more exopolysaccarides than the wild type produced. A plasmid carrying the USDA257 gltA gene, when introduced into HBK-CS1, was able to restore all of the defects mentioned above. Our results demonstrate that a functional citrate synthase gene of S. fredii USDA257 is essential for efficient soybean nodulation and nitrogen fixation.
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Lien, Huang-Chun, Yi-Hsuan Lee, Yung-Ming Jeng, Ching-Hung Lin, Yen-Shen Lu, and Yu-Tung Yao. "Differential expression of hyaluronan synthase 2 in breast carcinoma and its biological significance." Histopathology 65, no. 3 (April 2, 2014): 328–39. http://dx.doi.org/10.1111/his.12390.
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Fujii, Nobuharu, Marni D. Boppart, Scott D. Dufresne, Patricia F. Crowley, Alison C. Jozsi, Kei Sakamoto, Haiyan Yu, et al. "Overexpression or ablation of JNK in skeletal muscle has no effect on glycogen synthase activity." American Journal of Physiology-Cell Physiology 287, no. 1 (July 2004): C200—C208. http://dx.doi.org/10.1152/ajpcell.00415.2003.
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c-Jun NH2-terminal kinase (JNK) is highly expressed in skeletal muscle and is robustly activated in response to muscle contraction. Little is known about the biological functions of JNK signaling in terminally differentiated muscle cells, although this protein has been proposed to regulate insulin-stimulated glycogen synthase activity in mouse skeletal muscle. To determine whether JNK signaling regulates contraction-stimulated glycogen synthase activation, we applied an electroporation technique to induce JNK overexpression (O/E) in mouse skeletal muscle. Ten days after electroporation, in situ muscle contraction increased JNK activity 2.6-fold in control muscles and 15-fold in the JNK O/E muscles. Despite the enormous activation of JNK activity in JNK O/E muscles, contraction resulted in similar increases in glycogen synthase activity in control and JNK O/E muscles. Consistent with these findings, basal and contraction-induced glycogen synthase activity was normal in muscles of both JNK1- and JNK2-deficient mice. JNK overexpression in muscle resulted in significant alterations in the basal phosphorylation state of several signaling proteins, such as extracellular signal-regulated kinase 1/2, p90 S6 kinase, glycogen synthase kinase 3, protein kinase B/Akt, and p70 S6 kinase, in the absence of changes in the expression of these proteins. These data suggest that JNK signaling regulates the phosphorylation state of several kinases in skeletal muscle. JNK activation is unlikely to be the major mechanism by which contractile activity increases glycogen synthase activity in skeletal muscle.
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Stahlberg, Henning, Daniel J. Müller, Kitaru Suda, Dimitrios Fotiadis, Andreas Engel, Thomas Meier, Ulrich Matthey, and Peter Dimroth. "Bacterial Na + ‐ATP synthase has an undecameric rotor." EMBO reports 2, no. 3 (March 2001): 229–33. http://dx.doi.org/10.1093/embo-reports/kve047.
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PUICĂ, Gina. "La peur de rater en tant que Roumain, un mobile de sa réussite littéraire." Synthesis 3, no. 3 (December 9, 2024): 115–24. https://doi.org/10.59277/synthe.2024.3.115.
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In this article, we attempt to analyse what seems to be a persistent obsession of Cioran’s over time: the fear of failing in life (and therefore also in his writing career) because of his Romanian origin. Cioran was convinced that Romanians were doomed to failure and that Romania embodied “the genius of failure”. To escape the fate reserved for writers from “small” countries and join the “World Republic of Letters” (Pascale Casanova), Cioran chose France as his country of residence, especially as he was fascinated by the country’s grandiose history. But despite having done everything to escape the “Romanian failure”, the “Romanian nothingness”, Cioran paradoxically remained attracted all his life by the philosophical and existential question of failure, and even seems to have despised his own success when success began to come. It is on these questions that our article will focus, drawing in particular on Cioran’s correspondence, a substantial selection of which has recently been published.