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  1 / 1777 MEDLINE  
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[PMID]:27773789
[Au] Autor:Mkrtchyan G; Graf A; Bettendorff L; Bunik V
[Ad] Endereço:Faculty of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Leninskije gori 1, 119992 Moscow, Russia.
[Ti] Título:Cellular thiamine status is coupled to function of mitochondrial 2-oxoglutarate dehydrogenase.
[So] Source:Neurochem Int;101:66-75, 2016 12.
[Is] ISSN:1872-9754
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Decreased thiamine and reduced activity of thiamine diphosphate (ThDP)-dependent 2-oxoglutarate dehydrogenase (OGDH) cause neurodegeneration. We hypothesized on concerted cell-specific regulation of the thiamine metabolism and ThDP-dependent reactions. We identified a smaller thiamine pool, a lower expression of the mitochondrial ThDP transporter, and a higher expression of OGDH in rat astrocytes versus neuroblastoma N2A. According to the data, the astrocytic OGDH may be up-regulated by an increase in intracellular ThDP, while the neuroblastomal OGDH functions at full ThDP saturation. Indeed, in rat astrocytes and brain cortex, OGDH inhibition by succinyl phosphonate (SP) enlarged the pool of thiamine compounds. Increased ThDP level in response to the OGDH inhibition presumably up-regulated the enzyme to compensate for a decrease in reducing power which occurred in SP-treated astrocytes. Under the same SP treatment of N2A cells, their thiamine pool and reducing power were unchanged, although SP action was evident from accumulation of glutamate. The presented data indicate that functional interplay between OGDH, other proteins of the tricarbocylic acid cycle and proteins of thiamine metabolism is an important determinant of physiology-specific networks and their homeostatic mechanisms.
[Mh] Termos MeSH primário: Córtex Cerebral/efeitos dos fármacos
Complexo Cetoglutarato Desidrogenase/metabolismo
Mitocôndrias/efeitos dos fármacos
Tiamina/metabolismo
[Mh] Termos MeSH secundário: Animais
Córtex Cerebral/metabolismo
Citoplasma/metabolismo
Ácido Glutâmico/metabolismo
Homeostase/efeitos dos fármacos
Homeostase/fisiologia
Camundongos
Mitocôndrias/metabolismo
Organofosfonatos/metabolismo
Organofosfonatos/farmacologia
Succinatos/metabolismo
Succinatos/farmacologia
Tiamina Pirofosfato/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Organophosphonates); 0 (Succinates); 0 (succinyl phosphonate); 3KX376GY7L (Glutamic Acid); EC 1.2.4.2 (Ketoglutarate Dehydrogenase Complex); Q57971654Y (Thiamine Pyrophosphate); X66NSO3N35 (Thiamine)
[Em] Mês de entrada:1801
[Cu] Atualização por classe:180228
[Lr] Data última revisão:
180228
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161105
[St] Status:MEDLINE


  2 / 1777 MEDLINE  
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[PMID]:28918750
[Au] Autor:Song Z; Pan J; Xie L; Gong G; Han S; Zhang W; Hu Y
[Ad] Endereço:China State Institute of Pharmaceutical Industry, Zhangjiang Institute, Shanghai, 201203, China. bebydou@hotmail.com.
[Ti] Título:Expression, Purification, and Activity of ActhiS, a Thiazole Biosynthesis Enzyme from Acremonium chrysogenum.
[So] Source:Biochemistry (Mosc);82(7):852-860, 2017 Jul.
[Is] ISSN:1608-3040
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Thiamine pyrophosphate is an essential coenzyme in all organisms. Its biosynthesis involves independent syntheses of the precursors, pyrimidine and thiazole, which are then coupled. In our previous study with overexpressed and silent mutants of ActhiS (thiazole biosynthesis enzyme from Acremonium chrysogenum), we found that the enzyme level correlated with intracellular thiamine content in A. chrysogenum. However, the exact structure and function of ActhiS remain unclear. In this study, the enzyme-bound ligand was characterized as the ADP adduct of 5-(2-hydroxyethyl)-4-methylthiazole-2-carboxylic acid (ADT) using HPLC and H NMR. The ligand-free ActhiS expressed in M9 minimal medium catalyzed conversion of NAD+ and glycine to ADT in the presence of iron. Furthermore, the C217 residue was identified as the sulfur donor for the thiazole moiety. These observations confirm that ActhiS is a thiazole biosynthesis enzyme in A. chrysogenum, and it serves as a sulfur source for the thiazole moiety.
[Mh] Termos MeSH primário: Acremonium/enzimologia
Proteínas Fúngicas/genética
Proteínas Fúngicas/metabolismo
Tiazóis/metabolismo
[Mh] Termos MeSH secundário: Difosfato de Adenosina/química
Difosfato de Adenosina/metabolismo
Sequência de Aminoácidos
Cromatografia Líquida de Alta Pressão
Proteínas Fúngicas/química
Proteínas Fúngicas/isolamento & purificação
Glicina/metabolismo
Ligantes
Espectroscopia de Ressonância Magnética
Espectrometria de Massas
Mutagênese Sítio-Dirigida
NAD/metabolismo
Proteínas Recombinantes/biossíntese
Proteínas Recombinantes/química
Proteínas Recombinantes/isolamento & purificação
Alinhamento de Sequência
Tiamina Pirofosfato/metabolismo
Tiazóis/química
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Fungal Proteins); 0 (Ligands); 0 (Recombinant Proteins); 0 (Thiazoles); 0U46U6E8UK (NAD); 61D2G4IYVH (Adenosine Diphosphate); Q57971654Y (Thiamine Pyrophosphate); TE7660XO1C (Glycine)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170926
[Lr] Data última revisão:
170926
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170919
[St] Status:MEDLINE
[do] DOI:10.1134/S0006297917070112


  3 / 1777 MEDLINE  
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[PMID]:28552632
[Au] Autor:Pasquini M; Fermani S; Tedesco D; Sciabolini C; Crozet P; Naldi M; Henri J; Vothknecht U; Bertucci C; Lemaire SD; Zaffagnini M; Francia F
[Ad] Endereço:Department of Pharmacy and Biotechnology, University of Bologna, Bologna, Italy.
[Ti] Título:Structural basis for the magnesium-dependent activation of transketolase from Chlamydomonas reinhardtii.
[So] Source:Biochim Biophys Acta;1861(8):2132-2145, 2017 08.
[Is] ISSN:0006-3002
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: In photosynthetic organisms, transketolase (TK) is involved in the Calvin-Benson cycle and participates to the regeneration of ribulose-5-phosphate. Previous studies demonstrated that TK catalysis is strictly dependent on thiamine pyrophosphate (TPP) and divalent ions such as Mg . METHODS: TK from the unicellular green alga Chlamydomonas reinhardtii (CrTK) was recombinantly produced and purified to homogeneity. Biochemical properties of the CrTK enzyme were delineated by activity assays and its structural features determined by CD analysis and X-ray crystallography. RESULTS: CrTK is homodimeric and its catalysis depends on the reconstitution of the holo-enzyme in the presence of both TPP and Mg . Activity measurements and CD analysis revealed that the formation of fully active holo-CrTK is Mg -dependent and proceeds with a slow kinetics. The 3D-structure of CrTK without cofactors (CrTK ) shows that two portions of the active site are flexible and disordered while they adopt an ordered conformation in the holo-form. Oxidative treatments revealed that Mg participates in the redox control of CrTK by changing its propensity to be inactivated by oxidation. Indeed, the activity of holo-form is unaffected by oxidation whereas CrTK in the apo-form or reconstituted with the sole TPP show a strong sensitivity to oxidative inactivation. CONCLUSION: These evidences indicate that Mg is fundamental to allow gradual conformational arrangements suited for optimal catalysis. Moreover, Mg is involved in the control of redox sensitivity of CrTK. GENERAL SIGNIFICANCE: The importance of Mg in the functionality and redox sensitivity of CrTK is correlated to light-dependent fluctuations of Mg in chloroplasts.
[Mh] Termos MeSH primário: Chlamydomonas reinhardtii/enzimologia
Magnésio/farmacologia
Transcetolase/química
[Mh] Termos MeSH secundário: Domínio Catalítico
Dicroísmo Circular
Cristalografia por Raios X
Oxirredução
Conformação Proteica
Tiamina Pirofosfato/farmacologia
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
EC 2.2.1.1 (Transketolase); I38ZP9992A (Magnesium); Q57971654Y (Thiamine Pyrophosphate)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171026
[Lr] Data última revisão:
171026
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170530
[St] Status:MEDLINE


  4 / 1777 MEDLINE  
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[PMID]:28520932
[Au] Autor:Bastet L; Chauvier A; Singh N; Lussier A; Lamontagne AM; Prévost K; Massé E; Wade JT; Lafontaine DA
[Ad] Endereço:Department of Biology, Faculty of Sciences, RNA Group, Université de Sherbrooke, Sherbrooke, Quebec J1K 2R1, Canada.
[Ti] Título:Translational control and Rho-dependent transcription termination are intimately linked in riboswitch regulation.
[So] Source:Nucleic Acids Res;45(12):7474-7486, 2017 Jul 07.
[Is] ISSN:1362-4962
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Riboswitches are regulatory elements that control gene expression by altering RNA structure upon the binding of specific metabolites. Although Bacillus subtilis riboswitches have been shown to control premature transcription termination, less is known about regulatory mechanisms employed by Escherichia coli riboswitches, which are predicted to regulate mostly at the level of translation initiation. Here, we present experimental evidence suggesting that the majority of known E. coli riboswitches control transcription termination by using the Rho transcription factor. In the case of the thiamin pyrophosphate-dependent thiM riboswitch, we find that Rho-dependent transcription termination is triggered as a consequence of translation repression. Using in vitro and in vivo assays, we show that the Rho-mediated regulation relies on RNA target elements located at the beginning of thiM coding region. Gene reporter assays indicate that relocating Rho target elements to a different gene induces transcription termination, demonstrating that such elements are modular domains controlling Rho. Our work provides strong evidence that translationally regulating riboswitches also regulate mRNA levels through an indirect control mechanism ensuring tight control of gene expression.
[Mh] Termos MeSH primário: Escherichia coli/genética
Regulação Bacteriana da Expressão Gênica
Biossíntese de Proteínas
Fator Rho/genética
Riboswitch
Terminação da Transcrição Genética
[Mh] Termos MeSH secundário: Sequência de Bases
Escherichia coli/metabolismo
Genes Reporter
Conformação de Ácido Nucleico
Proteínas Recombinantes de Fusão/genética
Proteínas Recombinantes de Fusão/metabolismo
Fator Rho/metabolismo
Tiamina Pirofosfato/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Recombinant Fusion Proteins); 0 (Rho Factor); 0 (Riboswitch); Q57971654Y (Thiamine Pyrophosphate)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171017
[Lr] Data última revisão:
171017
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170519
[St] Status:MEDLINE
[do] DOI:10.1093/nar/gkx434


  5 / 1777 MEDLINE  
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[PMID]:28490063
[Au] Autor:Wang B; Bai Y; Fan T; Zheng X; Cai Y
[Ad] Endereço:The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
[Ti] Título:Characterisation of a thiamine diphosphate-dependent alpha-keto acid decarboxylase from Proteus mirabilis JN458.
[So] Source:Food Chem;232:19-24, 2017 Oct 01.
[Is] ISSN:0308-8146
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:Alpha-keto acid decarboxylases can convert keto acids to their corresponding aldehydes, which are often volatile aroma compounds. The gene encoding α-keto acid decarboxylase in Proteus mirabilis JN458 was cloned, and the enzyme overexpressed in Escherichia coli BL21 (DE3), purified in high yield, and characterised. The molecular weight is 62.291kDa by MALDI-TOF MS, and optimum activity at pH 6.0 and 40-50°C. The enzyme is a typical decarboxylase, dependent on thiamine diphosphate and Mg as cofactors. For the decarboxylation reaction, the enzyme displayed a broad substrate range. Kinetic parameters were determined using 4-methyl-2-oxopentanoic acid, phenyl pyruvate and 3-methyl-2-oxopentanoic acid as substrates. K and k values for phenyl pyruvate were 0.62mM and 77.38s , respectively, and the k /K value was 124.81mM s . The enzyme properties suggest it may act effectively under cheese ripening conditions.
[Mh] Termos MeSH primário: 3-Metil-2-Oxobutanoato Desidrogenase (Lipoamida)/metabolismo
Proteínas de Bactérias/metabolismo
Proteus mirabilis/enzimologia
Tiamina Pirofosfato/metabolismo
[Mh] Termos MeSH secundário: 3-Metil-2-Oxobutanoato Desidrogenase (Lipoamida)/química
Proteínas de Bactérias/química
Ativação Enzimática
Cinética
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); EC 1.2.4.4 (3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)); Q57971654Y (Thiamine Pyrophosphate)
[Em] Mês de entrada:1708
[Cu] Atualização por classe:170816
[Lr] Data última revisão:
170816
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170512
[St] Status:MEDLINE


  6 / 1777 MEDLINE  
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[PMID]:28296385
[Au] Autor:Paulikat M; Wechsler C; Tittmann K; Mata RA
[Ad] Endereço:Institute of Physical Chemistry, University of Goettingen , Tammannstraße 6, D-37077 Göttingen, Germany.
[Ti] Título:Theoretical Studies of the Electronic Absorption Spectra of Thiamin Diphosphate in Pyruvate Decarboxylase.
[So] Source:Biochemistry;56(13):1854-1864, 2017 Apr 04.
[Is] ISSN:1520-4995
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Electronic absorption spectra are oftentimes used to identify reaction intermediates or substrates/products in enzymatic systems, as long as absorption bands can be unequivocally assigned to the species being studied. The latter task is far from trivial given the transient nature of some states and the complexity of the surrounding environment around the active site. To identify unique spectral fingerprints, controlled experiments with model compounds have been used in the past, but even these can sometimes be unreliable. Circular dichroism (CD) and ultraviolet-visible spectra have been tools of choice in the study of the rich chemistry of thiamin diphosphate-dependent enzymes. In this study, we focus on the Zymomonas mobilis pyruvate decarboxylase, and mutant analogues thereof, as a prototypical representative of the thiamin diphosphate (ThDP) enzyme superfamily. Through the use of electronic structure methods, we analyze the nature of electronic excitations in the cofactor. We find that all the determining CD bands around the 280-340 nm spectral range correspond to charge-transfer excitations between the pyrimidine and thiazolium rings of ThDP, which, most likely, is a general property of related ThDP-dependent enzymes. While we can confirm the assignments of previously proposed bands to chemical states, our calculations further suggest that a hitherto unassigned band of enzyme-bound ThDP reports on the ionization state of the canonical glutamate that is required for cofactor activation. This finding expands the spectroscopic "library" of chemical states of ThDP enzymes, permitting a simultaneous assignment of both the cofactor ThDP and the activating glutamate. We anticipate this finding to be helpful for mechanistic analyses of related ThDP enzymes.
[Mh] Termos MeSH primário: Proteínas de Bactérias/química
Coenzimas/química
Ácido Glutâmico/química
Piruvato Descarboxilase/química
Tiamina Pirofosfato/química
Zymomonas/química
[Mh] Termos MeSH secundário: Proteínas de Bactérias/genética
Proteínas de Bactérias/metabolismo
Domínio Catalítico
Coenzimas/metabolismo
Transporte de Elétrons
Expressão Gênica
Ácido Glutâmico/metabolismo
Cinética
Simulação de Dinâmica Molecular
Mutação
Estrutura Secundária de Proteína
Pirimidinas/química
Piruvato Descarboxilase/genética
Piruvato Descarboxilase/metabolismo
Eletricidade Estática
Termodinâmica
Tiamina Pirofosfato/metabolismo
Zymomonas/enzimologia
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (Coenzymes); 0 (Pyrimidines); 3KX376GY7L (Glutamic Acid); EC 4.1.1.1 (Pyruvate Decarboxylase); K8CXK5Q32L (pyrimidine); Q57971654Y (Thiamine Pyrophosphate)
[Em] Mês de entrada:1705
[Cu] Atualização por classe:170515
[Lr] Data última revisão:
170515
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170316
[St] Status:MEDLINE
[do] DOI:10.1021/acs.biochem.6b00984


  7 / 1777 MEDLINE  
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[PMID]:27920295
[Au] Autor:Rodionov DA; Leyn SA; Li X; Rodionova IA
[Ad] Endereço:A. A. Kharkevich Institute for Information Transmission Problems, Russian Academy of Sciences, Moscow, Russia rodionov@burnham.org.
[Ti] Título:A Novel Transcriptional Regulator Related to Thiamine Phosphate Synthase Controls Thiamine Metabolism Genes in Archaea.
[So] Source:J Bacteriol;199(4), 2017 Feb 15.
[Is] ISSN:1098-5530
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Thiamine (vitamin B ) is a precursor of thiamine pyrophosphate (TPP), an essential coenzyme in the central metabolism of all living organisms. Bacterial thiamine biosynthesis and salvage genes are controlled at the RNA level by TPP-responsive riboswitches. In Archaea, TPP riboswitches are restricted to the Thermoplasmatales order. Mechanisms of transcriptional control of thiamine genes in other archaeal lineages remain unknown. Using the comparative genomics approach, we identified a novel family of transcriptional regulators (named ThiR) controlling thiamine biosynthesis and transport genes in diverse lineages in the Crenarchaeota phylum as well as in the Halobacteria and Thermococci classes of the Euryarchaeota ThiR regulators are composed of an N-terminal DNA-binding domain and a C-terminal ligand-binding domain, which is similar to the archaeal thiamine phosphate synthase ThiN. By using comparative genomics, we predicted ThiR-binding DNA motifs and reconstructed ThiR regulons in 67 genomes representing all above-mentioned lineages. The predicted ThiR-binding motifs are characterized by palindromic symmetry with several distinct lineage-specific consensus sequences. In addition to thiamine biosynthesis genes, the reconstructed ThiR regulons include various transporters for thiamine and its precursors. Bioinformatics predictions were experimentally validated by in vitro DNA-binding assays with the recombinant ThiR protein from the hyperthermophilic archaeon Metallosphaera yellowstonensis MK1. Thiamine phosphate and, to some extent, TPP and hydroxyethylthiazole phosphate were required for the binding of ThiR to its DNA targets, suggesting that ThiR is derepressed by limitation of thiamine phosphates. The thiamine phosphate-binding residues previously identified in ThiN are highly conserved in ThiR regulators, suggesting a conserved mechanism for effector recognition. IMPORTANCE: Thiamine pyrophosphate is a cofactor for many essential enzymes for glucose and energy metabolism. Thiamine or vitamin B biosynthesis and its transcriptional regulation in Archaea are poorly understood. We applied the comparative genomics approach to identify a novel family of regulators for the transcriptional control of thiamine metabolism genes in Archaea and reconstructed the respective regulons. The predicted ThiR regulons in archaeal genomes control the majority of thiamine biosynthesis genes. The reconstructed regulon content suggests that numerous uptake transporters for thiamine and/or its precursors are encoded in archaeal genomes. The ThiR regulon was experimentally validated by DNA-binding assays with Metallosphaera spp. These discoveries contribute to our understanding of metabolic and regulatory networks involved in vitamin homeostasis in diverse lineages of Archaea.
[Mh] Termos MeSH primário: Alquil e Aril Transferases/metabolismo
Archaea/enzimologia
Regulação da Expressão Gênica em Archaea/fisiologia
Regulação Enzimológica da Expressão Gênica/fisiologia
Tiamina Pirofosfato/metabolismo
Tiamina/metabolismo
[Mh] Termos MeSH secundário: Alquil e Aril Transferases/genética
Archaea/genética
Archaea/metabolismo
Biologia Computacional
Genoma Arqueal/genética
Genômica
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
EC 2.5.- (Alkyl and Aryl Transferases); Q57971654Y (Thiamine Pyrophosphate); X66NSO3N35 (Thiamine)
[Em] Mês de entrada:1706
[Cu] Atualização por classe:170815
[Lr] Data última revisão:
170815
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161207
[St] Status:MEDLINE


  8 / 1777 MEDLINE  
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[PMID]:27657847
[Au] Autor:Halma M; Doumèche B; Hecquet L; Prévot V; Mousty C; Charmantray F
[Ad] Endereço:Clermont Université, Université Blaise Pascal, Institut de Chimie de Clermont-Ferrand, BP 10448, F-63000 Clermont-Ferrand, France; CNRS, UMR 6296, ICCF, F-63171 Aubière, France.
[Ti] Título:Thiamine biosensor based on oxidative trapping of enzyme-substrate intermediate.
[So] Source:Biosens Bioelectron;87:850-857, 2017 Jan 15.
[Is] ISSN:1873-4235
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:In the present work, we describe a new thiamine amperometric biosensor based on thiamine pyrophosphate (ThDP)-dependent transketolase (TK)-catalyzed reaction, followed by the oxidative trapping of TK intermediate α,ß-dihydroxyethylthiamine diphosphate (DHEThDP) within the enzymatic active site. For the biosensor design purpose, TK from Escherichia coli (TKec) was immobilized in Mg Al-NO Layered Double Hydroxides (LDH) and the electrochemical detection was achieved with the TKec/LDH modified glassy carbon electrode (GCE). The transduction process was based on the ability of Fe(CN) to oxidize DHEThDP to glycolic acid along with ThDP regeneration. The released Fe(CN) was re-oxidized at +0.5V vs Ag-AgCl and the reaction was followed by chronoamperometry. The TKec/LDH/GCE biosensor was optimized using the best TK donor substrates, namely l-erythrulose and d-fructose-6-phosphate. ThDP was assayed with great sensitivity (3831mAM cm ) over 20-400nM linear range.
[Mh] Termos MeSH primário: Técnicas Biossensoriais/métodos
Escherichia coli/enzimologia
Tiamina Pirofosfato/análise
Tiamina/análise
Transcetolase/metabolismo
[Mh] Termos MeSH secundário: Técnicas Eletroquímicas/métodos
Enzimas Imobilizadas/metabolismo
Ferrocianetos/metabolismo
Hidróxidos/metabolismo
Oxirredução
Tiamina/metabolismo
Tiamina Pirofosfato/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Enzymes, Immobilized); 0 (Ferrocyanides); 0 (Hydroxides); EC 2.2.1.1 (Transketolase); FLX0VIC39Y (hexacyanoferrate II); Q57971654Y (Thiamine Pyrophosphate); X66NSO3N35 (Thiamine)
[Em] Mês de entrada:1702
[Cu] Atualização por classe:170816
[Lr] Data última revisão:
170816
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160923
[St] Status:MEDLINE


  9 / 1777 MEDLINE  
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[PMID]:27694800
[Au] Autor:Genee HJ; Bali AP; Petersen SD; Siedler S; Bonde MT; Gronenberg LS; Kristensen M; Harrison SJ; Sommer MO
[Ad] Endereço:The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kongens Lyngby, Denmark.
[Ti] Título:Functional mining of transporters using synthetic selections.
[So] Source:Nat Chem Biol;12(12):1015-1022, 2016 Dec.
[Is] ISSN:1552-4469
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Only 25% of bacterial membrane transporters have functional annotation owing to the difficulty of experimental study and of accurate prediction of their function. Here we report a sequence-independent method for high-throughput mining of novel transporters. The method is based on ligand-responsive biosensor systems that enable selective growth of cells only if they encode a ligand-specific importer. We developed such a synthetic selection system for thiamine pyrophosphate and mined soil and gut metagenomes for thiamine-uptake functions. We identified several members of a novel class of thiamine transporters, PnuT, which is widely distributed across multiple bacterial phyla. We demonstrate that with modular replacement of the biosensor, we could expand our method to xanthine and identify xanthine permeases from gut and soil metagenomes. Our results demonstrate how synthetic-biology approaches can effectively be deployed to functionally mine metagenomes and elucidate sequence-function relationships of small-molecule transport systems in bacteria.
[Mh] Termos MeSH primário: Técnicas Biossensoriais/métodos
Proteínas de Membrana Transportadoras/isolamento & purificação
Proteínas de Membrana Transportadoras/metabolismo
Metagenoma
Tiamina Pirofosfato/metabolismo
Xantinas/metabolismo
[Mh] Termos MeSH secundário: Bactérias/enzimologia
Bactérias/metabolismo
Microbioma Gastrointestinal
Ensaios de Triagem em Larga Escala
Ligantes
Microbiologia do Solo
Biologia Sintética/métodos
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Ligands); 0 (Membrane Transport Proteins); 0 (Xanthines); Q57971654Y (Thiamine Pyrophosphate)
[Em] Mês de entrada:1705
[Cu] Atualização por classe:170509
[Lr] Data última revisão:
170509
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161004
[St] Status:MEDLINE
[do] DOI:10.1038/nchembio.2189


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Fotocópia
[PMID]:27677881
[Au] Autor:Mimura M; Zallot R; Niehaus TD; Hasnain G; Gidda SK; Nguyen TN; Anderson EM; Mullen RT; Brown G; Yakunin AF; de Crécy-Lagard V; Gregory JF; McCarty DR; Hanson AD
[Ad] Endereço:Horticultural Sciences Department, University of Florida, Gainesville, Florida 32611.
[Ti] Título:Arabidopsis TH2 Encodes the Orphan Enzyme Thiamin Monophosphate Phosphatase.
[So] Source:Plant Cell;28(10):2683-2696, 2016 Oct.
[Is] ISSN:1532-298X
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:To synthesize the cofactor thiamin diphosphate (ThDP), plants must first hydrolyze thiamin monophosphate (ThMP) to thiamin, but dedicated enzymes for this hydrolysis step were unknown and widely doubted to exist. The classical thiamin-requiring th2-1 mutation in Arabidopsis thaliana was shown to reduce ThDP levels by half and to increase ThMP levels 5-fold, implying that the THIAMIN REQUIRING2 (TH2) gene product could be a dedicated ThMP phosphatase. Genomic and transcriptomic data indicated that TH2 corresponds to At5g32470, encoding a HAD (haloacid dehalogenase) family phosphatase fused to a TenA (thiamin salvage) family protein. Like the th2-1 mutant, an insertional mutant of At5g32470 accumulated ThMP, and the thiamin requirement of the th2-1 mutant was complemented by wild-type At5g32470 Complementation tests in Escherichia coli and enzyme assays with recombinant proteins confirmed that At5g32470 and its maize (Zea mays) orthologs GRMZM2G148896 and GRMZM2G078283 are ThMP-selective phosphatases whose activity resides in the HAD domain and that the At5g32470 TenA domain has the expected thiamin salvage activity. In vitro and in vivo experiments showed that alternative translation start sites direct the At5g32470 protein to the cytosol and potentially also to mitochondria. Our findings establish that plants have a dedicated ThMP phosphatase and indicate that modest (50%) ThDP depletion can produce severe deficiency symptoms.
[Mh] Termos MeSH primário: Proteínas de Arabidopsis/metabolismo
Arabidopsis/metabolismo
Tiamina Pirofosfato/metabolismo
[Mh] Termos MeSH secundário: Arabidopsis/enzimologia
Proteínas de Arabidopsis/genética
Escherichia coli/genética
Escherichia coli/metabolismo
Monoéster Fosfórico Hidrolases/genética
Monoéster Fosfórico Hidrolases/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Arabidopsis Proteins); EC 3.1.3.2 (Phosphoric Monoester Hydrolases); Q57971654Y (Thiamine Pyrophosphate)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171107
[Lr] Data última revisão:
171107
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160929
[St] Status:MEDLINE



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