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[PMID]:28465208
[Au] Autor:Lee YJ; Park MK; Park GS; Lee SJ; Lee SJ; Kim BS; Shin JH; Lee DW
[Ad] Endereço:School of Applied Biosciences, Kyungpook National University, Daegu 41566, South Korea.
[Ti] Título:Complete genome sequence of the thermophilic bacterium Geobacillus subterraneus KCTC 3922 as a potential denitrifier.
[So] Source:J Biotechnol;251:141-144, 2017 Jun 10.
[Is] ISSN:1873-4863
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:Denitrification is a crucial process for the global nitrogen cycle through the reduction of nitrates by heterotrophic bacteria. Denitrifying microorganisms play an important role in eliminating fixed nitrogen pollutants from the ecosystem, concomitant with N O emission. Although many microbial denitrifiers have been identified, little is known about the denitrifying ability of the genus Geobacillus. Here, we report the first complete genome sequences of Geobacillus subterraneus KCTC 3922 , isolated from Liaohe oil field in China, and G. thermodenitrificans KCTC 3902 . The strain KCTC 3922 contains a complete set of genes involved in denitrification, cofactor biogenesis, and transport systems, which is consistent with a denitrifying activity. On the other hand, G. thermodenitrificans KCTC 3902 exhibited no denitrifying activity probably due to the lack of molybdnumtransferase (moeA) and nitrite transporter (nirC) genes. Therefore, comparative genome analysis of Geobacillus strains highlights the potential impact on treatment of nitrate-contaminated environments.
[Mh] Termos MeSH primário: Desnitrificação/genética
Genoma Bacteriano
Geobacillus/genética
Geobacillus/metabolismo
[Mh] Termos MeSH secundário: Sequência de Bases
DNA Bacteriano/genética
Nitratos/metabolismo
Análise de Sequência de DNA
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (DNA, Bacterial); 0 (Nitrates)
[Em] Mês de entrada:1802
[Cu] Atualização por classe:180205
[Lr] Data última revisão:
180205
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170504
[St] Status:MEDLINE


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[PMID]:28934494
[Au] Autor:Oscorbin IP; Belousova EA; Boyarskikh UA; Zakabunin AI; Khrapov EA; Filipenko ML
[Ad] Endereço:Laboratory of Pharmacogenomics, Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, Lavrentiev Avenue 8, Novosibirsk 630090, Russian Federation.
[Ti] Título:Derivatives of Bst-like Gss-polymerase with improved processivity and inhibitor tolerance.
[So] Source:Nucleic Acids Res;45(16):9595-9610, 2017 Sep 19.
[Is] ISSN:1362-4962
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:At the moment, one of the actual trends in medical diagnostics is a development of methods for practical applications such as point-of-care testing, POCT or research tools, for example, whole genome amplification, WGA. All the techniques are based on using of specific DNA polymerases having strand displacement activity, high synthetic processivity, fidelity and, most significantly, tolerance to contaminants, appearing from analysed biological samples or collected under purification procedures. Here, we have designed a set of fusion enzymes based on catalytic domain of DNA polymerase I from Geobacillus sp. 777 with DNA-binding domain of DNA ligase Pyrococcus abyssi and Sto7d protein from Sulfolobus tokodaii, analogue of Sso7d. Designed chimeric DNA polymerases DBD-Gss, Sto-Gss and Gss-Sto exhibited the same level of thermal stability, thermal transferase activity and fidelity as native Gss; however, the processivity was increased up to 3-fold, leading to about 4-fold of DNA product in WGA which is much more exiting. The attachment of DNA-binding proteins enhanced the inhibitor tolerance of chimeric polymerases in loop-mediated isothermal amplification to several of the most common DNA sample contaminants-urea and whole blood, heparin, ethylenediaminetetraacetic acid, NaCl, ethanol. Therefore, chimeric Bst-like Gss-polymerase will be promising tool for both WGA and POCT due to increased processivity and inhibitor tolerance.
[Mh] Termos MeSH primário: DNA Polimerase I/metabolismo
Geobacillus/enzimologia
Engenharia de Proteínas/métodos
Proteínas Recombinantes de Fusão/genética
[Mh] Termos MeSH secundário: Domínio Catalítico
Clonagem Molecular
DNA/metabolismo
DNA Polimerase I/antagonistas & inibidores
DNA Polimerase I/genética
Inibidores Enzimáticos/farmacologia
Genoma Humano
Geobacillus/genética
Geobacillus stearothermophilus/enzimologia
Geobacillus stearothermophilus/genética
Seres Humanos
Técnicas de Amplificação de Ácido Nucleico/métodos
Estabilidade Proteica
Pyrococcus abyssi/genética
Proteínas Recombinantes de Fusão/metabolismo
Sulfolobus/genética
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Enzyme Inhibitors); 0 (Recombinant Fusion Proteins); 9007-49-2 (DNA); EC 2.7.7.- (DNA Polymerase I)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171024
[Lr] Data última revisão:
171024
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170922
[St] Status:MEDLINE
[do] DOI:10.1093/nar/gkx645


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[PMID]:28842992
[Au] Autor:Kinfu BM; Jahnke M; Janus M; Besirlioglu V; Roggenbuck M; Meurer R; Vojcic L; Borchert M; Schwaneberg U; Chow J; Streit WR
[Ad] Endereço:Microbiology and Biotechnology, Biocenter Klein Flottbek, University of Hamburg, Ohnhorststr, Hamburg, Germany.
[Ti] Título:Recombinant RNA Polymerase from Geobacillus sp. GHH01 as tool for rapid generation of metagenomic RNAs using in vitro technologies.
[So] Source:Biotechnol Bioeng;114(12):2739-2752, 2017 Dec.
[Is] ISSN:1097-0290
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The exciting promises of functional metagenomics for the efficient discovery of novel biomolecules from nature are often hindered by factors associated with expression hosts. Aiming to shift functional metagenomics to a host independent innovative system, we here report on the cloning, heterologous expression, and reconstitution of an RNA polymerase (RNAP) from the thermophilic Geobacillus sp. GHH01 and in vitro transcription thereafter. The five genes coding for RNAP subunits, a house keeping sigma factor and two transcription elongation factors were cloned and over expressed as His -tagged and/ or tag-free proteins. Purified subunits were reconstituted into a functional polymerase through either the classical method of denaturation and subsequent renaturation or through a new resource and time efficient thermo-reconstitution method which takes advantage of the subunits' temperature stability. Additionally, all subunits were cloned into a single vector system for a co-expression and in vivo reconstitution to the RNAP core enzyme. Both the core and holoenzyme form of the RNAP exhibited a robust transcription activity and were stable up to a temperature of 55°C close to their fullest activity. The Geobacillus RNAP showed a remarkable in vitro transcription profile recognizing DNA template sequences of diverse bacteria and archaea as well as metagenomic samples. Coupled with a subsequent in vitro translation step, this recombinant transcription system could allow a new, clone-free, and functional metagenomic screening approach.
[Mh] Termos MeSH primário: RNA Polimerases Dirigidas por DNA/genética
Melhoramento Genético/métodos
Geobacillus/genética
Metagenoma/genética
RNA/biossíntese
Proteínas Recombinantes/genética
[Mh] Termos MeSH secundário: Regulação Bacteriana da Expressão Gênica/genética
Regulação Enzimológica da Expressão Gênica/genética
RNA/genética
Proteínas Recombinantes/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Recombinant Proteins); 63231-63-0 (RNA); EC 2.7.7.6 (DNA-Directed RNA Polymerases)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171113
[Lr] Data última revisão:
171113
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170827
[St] Status:MEDLINE
[do] DOI:10.1002/bit.26436


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[PMID]:28587636
[Au] Autor:Pei S; Slinger BL; Meyer MM
[Ad] Endereço:Boston College, 140 Commonwealth Ave., 02467, Chestnut Hill, USA.
[Ti] Título:Recognizing RNA structural motifs in HT-SELEX data for ribosomal protein S15.
[So] Source:BMC Bioinformatics;18(1):298, 2017 Jun 06.
[Is] ISSN:1471-2105
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: Proteins recognize many different aspects of RNA ranging from single stranded regions to discrete secondary or tertiary structures. High-throughput sequencing (HTS) of in vitro selected populations offers a large scale method to study RNA-proteins interactions. However, most existing analysis methods require that the binding motifs are enriched in the population relative to earlier rounds, and that motifs are found in a loop or single stranded region of the potential RNA secondary structure. Such methods do not generalize to all RNA-protein interaction as some RNA binding proteins specifically recognize more complex structures such as double stranded RNA. RESULTS: In this study, we use HT-SELEX derived populations to study the landscape of RNAs that interact with Geobacillus kaustophilus ribosomal protein S15. Our data show high sequence and structure diversity and proved intractable to existing methods. Conventional programs identified some sequence motifs, but these are found in less than 5-10% of the total sequence pool. Therefore, we developed a novel framework to analyze HT-SELEX data. Our process accounts for both sequence and structure components by abstracting the overall secondary structure into smaller substructures composed of a single base-pair stack, which allows us to leverage existing approaches already used in k-mer analysis to identify enriched motifs. By focusing on secondary structure motifs composed of specific two base-pair stacks, we identified significantly enriched or depleted structure motifs relative to earlier rounds. CONCLUSIONS: Discrete substructures are likely to be important to RNA-protein interactions, but they are difficult to elucidate. Substructures can help make highly diverse sequence data more tractable. The structure motifs provide limited accuracy in predicting enrichment suggesting that G. kaustophilus S15 can either recognize many different secondary structure motifs or some aspects of the interaction are not captured by the analysis. This highlights the importance of considering secondary and tertiary structure elements and their role in RNA-protein interactions.
[Mh] Termos MeSH primário: Algoritmos
Proteínas Ribossômicas/metabolismo
[Mh] Termos MeSH secundário: Pareamento de Bases
Sequência de Bases
Análise por Conglomerados
Geobacillus/genética
Sequenciamento de Nucleotídeos em Larga Escala
Modelos Logísticos
Conformação de Ácido Nucleico
Ligação Proteica
Proteínas de Ligação a RNA/metabolismo
Proteínas Ribossômicas/química
Proteínas Ribossômicas/genética
Análise de Sequência de RNA
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (RNA-Binding Proteins); 0 (Ribosomal Proteins); 0 (ribosomal protein S15)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171102
[Lr] Data última revisão:
171102
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170608
[St] Status:MEDLINE
[do] DOI:10.1186/s12859-017-1704-y


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[PMID]:28574263
[Au] Autor:Miller EK; Trivelas NE; Maugeri PT; Blaesi EJ; Shafaat HS
[Ti] Título:Time-Resolved Investigations of Heterobimetallic Cofactor Assembly in R2lox Reveal Distinct Mn/Fe Intermediates.
[So] Source:Biochemistry;56(26):3369-3379, 2017 Jul 05.
[Is] ISSN:1520-4995
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The assembly mechanism of the Mn/Fe ligand-binding oxidases (R2lox), a family of proteins that are homologous to the nonheme diiron carboxylate enzymes, has been investigated using time-resolved techniques. Multiple heterobimetallic intermediates that exhibit unique spectral features, including visible absorption bands and exceptionally broad electron paramagnetic resonance signatures, are observed through optical and magnetic resonance spectroscopies. On the basis of comparison to known diiron species and model compounds, the spectra have been attributed to (µ-peroxo)-Mn /Fe and high-valent Mn/Fe species. Global spectral analysis coupled with isotopic substitution and kinetic modeling reveals elementary rate constants for the assembly of Mn/Fe R2lox under aerobic conditions. A complete reaction mechanism for cofactor maturation that is consistent with experimental data has been developed. These results suggest that the Mn/Fe cofactor can perform direct C-H bond abstraction, demonstrating the potential for potent chemical reactivity that remains unexplored.
[Mh] Termos MeSH primário: Proteínas de Bactérias/metabolismo
Coenzimas/metabolismo
Geobacillus/enzimologia
Ferro/metabolismo
Manganês/metabolismo
Modelos Moleculares
Oxirredutases/metabolismo
[Mh] Termos MeSH secundário: Algoritmos
Apoenzimas/química
Apoenzimas/genética
Apoenzimas/metabolismo
Proteínas de Bactérias/química
Proteínas de Bactérias/genética
Domínio Catalítico
Coenzimas/química
Medição da Troca de Deutério
Espectroscopia de Ressonância de Spin Eletrônica
Ativação Enzimática
Estabilidade Enzimática
Ferro/química
Isótopos de Ferro
Cinética
Ligantes
Manganês/química
Oxirredutases/química
Oxirredutases/genética
Proteínas Recombinantes/química
Proteínas Recombinantes/metabolismo
Espectrofotometria
[Pt] Tipo de publicação:COMPARATIVE STUDY; JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Apoenzymes); 0 (Bacterial Proteins); 0 (Coenzymes); 0 (Iron Isotopes); 0 (Ligands); 0 (Recombinant Proteins); 42Z2K6ZL8P (Manganese); E1UOL152H7 (Iron); EC 1.- (Oxidoreductases)
[Em] Mês de entrada:1707
[Cu] Atualização por classe:171111
[Lr] Data última revisão:
171111
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170603
[St] Status:MEDLINE
[do] DOI:10.1021/acs.biochem.7b00403


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[PMID]:28557456
[Au] Autor:Liu Y; Ban X; Li C; Gu Z; Cheng L; Hong Y; Li Z
[Ad] Endereço:School of Food Science and Technology, Jiangnan University , Wuxi 214122, China.
[Ti] Título:Met349 Mutations Enhance the Activity of 1,4-α-Glucan Branching Enzyme from Geobacillus thermoglucosidans STB02.
[So] Source:J Agric Food Chem;65(28):5674-5680, 2017 Jul 19.
[Is] ISSN:1520-5118
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:1,4-α-Glucan branching enzyme (GBE, EC 2.4.1.18) is used to increase the number of α-1,6 branch points in starch and glycogen. On the basis of a multiple sequence alignment of the GBEs from a variety of bacteria, residue 349 (Geobacillus thermoglucosidans STB02 numbering) in region III is generally methionine in bacteria with higher identity, while it is threonine or serine in bacteria with lower identity. Four mutants (M349T, M349S, M349H, and M349Y) were constructed by site-directed mutagenesis and characterized. M349T and M349S showed 24.5% and 21.1% increases in specific activity compared with that of wild-type GBE, respectively. In addition, M349T and M349S displayed 24.2% and 17.6% enhancements in the α-1,6-glycosidic linkage ratio of potato starch samples, respectively. However, M349Y displayed a significant reduction in activity. Moreover, the mutations at M349 have a negligible effect on substrate specificity. Thus, M349T and M349S are more suitable for industrial applications than wild-type GBE.
[Mh] Termos MeSH primário: Enzima Ramificadora de 1,4-alfa-Glucana/genética
Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo
Proteínas de Bactérias/genética
Proteínas de Bactérias/metabolismo
Geobacillus/enzimologia
[Mh] Termos MeSH secundário: Proteínas de Bactérias/química
Geobacillus/química
Geobacillus/genética
Glicogênio/metabolismo
Mutagênese Sítio-Dirigida
Mutação
Mutação de Sentido Incorreto
Especificidade por Substrato
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 9005-79-2 (Glycogen); EC 2.4.1.18 (1,4-alpha-Glucan Branching Enzyme)
[Em] Mês de entrada:1707
[Cu] Atualização por classe:170728
[Lr] Data última revisão:
170728
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170531
[St] Status:MEDLINE
[do] DOI:10.1021/acs.jafc.7b01227


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[PMID]:28422446
[Au] Autor:Jia X; Guo Y; Lin X; You M; Lin C; Chen L; Chen J
[Ad] Endereço:Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou, China.
[Ti] Título:Fusion of a family 20 carbohydrate-binding module (CBM20) with cyclodextrin glycosyltransferase of Geobacillus sp. CHB1 improves catalytic efficiency.
[So] Source:J Basic Microbiol;57(6):471-480, 2017 Jun.
[Is] ISSN:1521-4028
[Cp] País de publicação:Germany
[La] Idioma:eng
[Ab] Resumo:Cyclodextrin glycosyltransferase (CGTase) is an important industrial enzyme for production of cyclodextrins (CDs) from starch by intramolecular transglycosylation. CGTase consists of five domains labeled A to E. For optimizing catalytic activity of CGTase, CGTase of Geobacillus sp. was fused with the family 20 carbohydrate-binding module (CBM) of the Bacillus circulans strain 251 CGTase. The CBM that has a low binding free energy with maltohexaose, was selected by in silico design. Then the fusion enzyme, CGTΔE-CBM was constructed by fusing the CBM to the C-terminal region of CGTΔE. The fusion enzyme displayed an even greater enhancement of total α-cyclization activity (40.2%) and γ-cyclization activity (181.58%). Optimal reaction pH range was wilder and the thermal stability was better under 50 and 60 °C. Compared to the wild-type CGTase, the fusion enzyme showed a remarkable decrease in Km and a slight alteration in Vmax. The enhancement of soluble starch catalytic efficiency might be due to the changes of substrate binding ability in the critical substrate binding sites between the CBM and starch granule.
[Mh] Termos MeSH primário: Geobacillus/enzimologia
Glucosiltransferases/genética
Glucosiltransferases/metabolismo
Amido/metabolismo
[Mh] Termos MeSH secundário: Bacillus/enzimologia
Bacillus/genética
Sítios de Ligação
Biocatálise
Carboidratos
Simulação por Computador
Ciclodextrinas/metabolismo
Geobacillus/genética
Glucosiltransferases/química
Concentração de Íons de Hidrogênio
Cinética
Modelos Moleculares
Mutagênese Sítio-Dirigida
Oligossacarídeos/metabolismo
Conformação Proteica
Proteínas Recombinantes de Fusão/química
Proteínas Recombinantes de Fusão/metabolismo
Amido/química
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Carbohydrates); 0 (Cyclodextrins); 0 (Oligosaccharides); 0 (Recombinant Fusion Proteins); 34620-77-4 (maltohexaose); 9005-25-8 (Starch); EC 2.4.1.- (Glucosyltransferases); EC 2.4.1.19 (cyclomaltodextrin glucanotransferase)
[Em] Mês de entrada:1711
[Cu] Atualização por classe:171109
[Lr] Data última revisão:
171109
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170420
[St] Status:MEDLINE
[do] DOI:10.1002/jobm.201600628


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[PMID]:28406925
[Au] Autor:Potprommanee L; Wang XQ; Han YJ; Nyobe D; Peng YP; Huang Q; Liu JY; Liao YL; Chang KL
[Ad] Endereço:School of Environmental Science and Engineering and Institute of Environmental Health and Pollution Control, Guangdong University of Technology, Guangzhou, China.
[Ti] Título:Characterization of a thermophilic cellulase from Geobacillus sp. HTA426, an efficient cellulase-producer on alkali pretreated of lignocellulosic biomass.
[So] Source:PLoS One;12(4):e0175004, 2017.
[Is] ISSN:1932-6203
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:A themophilic cellulase-producing bacterium was isolated from a hot spring district and identified as Geobacillus sp. HTA426. The cellulase enzyme produced by the Geobacillus sp. HTA426 was purified through ammonium sulfate precipitation and ion exchange chromatography, with the recovery yield and fold purification of 10.14% and 5.12, respectively. The purified cellulase has a molecular weight of 40 kDa. The optimum temperature and pH for carboxymethyl cellulase (CMCase) activity of the purified cellulase were 60°C and pH 7.0, respectively. The enzyme was also stable over a wide temperature range of 50°C to 70°C after 5 h of incubation. Moreover, the strain HTA426 was able to grow and produce cellulase on alkali-treated sugarcane bagasse, rice straw and water hyacinth as carbon sources. Enzymatic hydrolysis of sugarcane bagasse, which was regarded as the most effective carbon source for cellulase production (CMCase activity = 103.67 U/mL), followed by rice straw (74.70 U/mL) and water hyacinth (51.10 U/mL). This strain producing an efficient thermostable cellulose is a potential candidate for developing a more efficient and cost-effective process for converting lignocellulosic biomass into biofuel and other industrial process.
[Mh] Termos MeSH primário: Biomassa
Celulase
Celulose/química
Geobacillus/enzimologia
Temperatura Alta
Lignina/química
[Mh] Termos MeSH secundário: Proteínas de Bactérias/biossíntese
Proteínas de Bactérias/química
Proteínas de Bactérias/isolamento & purificação
Celulase/biossíntese
Celulase/química
Celulase/isolamento & purificação
Estabilidade Enzimática
Concentração de Íons de Hidrogênio
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Bacterial Proteins); 11132-73-3 (lignocellulose); 9004-34-6 (Cellulose); 9005-53-2 (Lignin); 9006-97-7 (bagasse); EC 3.2.1.4 (Cellulase)
[Em] Mês de entrada:1704
[Cu] Atualização por classe:170504
[Lr] Data última revisão:
170504
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170414
[St] Status:MEDLINE
[do] DOI:10.1371/journal.pone.0175004


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[PMID]:28393686
[Au] Autor:Moisan JK; Meddeb-Mouelhi F; Charbonneau DM; Beauregard M
[Ad] Endereço:Centre de Recherche sur les Matériaux Lignocellulosiques, Université du Québec à Trois-Rivières, Trois-Rivières (Québec) G9A 5H7, Canada.
[Ti] Título:Impact of Salt Concentration and pH on Surface Charged Residues: Controlling Protein Association Pathways in Carboxylesterase EstGtA2.
[So] Source:Protein Pept Lett;24(6):561-572, 2017.
[Is] ISSN:1875-5305
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: Understanding the relationship between enzymatic stability and the amino acid sequence encoding carboxylesterases is of utmost importance. OBJECTIVES: Here we thoroughly characterized the behavior of the carboxylesterase EstGtA2 from Geobacillus thermodenitrificans during thermal denaturation at different pH with various salt concentrations. METHOD: EstGtA2 was characterized by circular dichroism regarding conformation and thermal stability, by dynamic light scattering for detection of association/aggregation, by enzymatic assays for activity and by monitoring the impact of heat treatments on activity. RESULTS: Our investigation revealed a particular dependence between aggregation/association and preservation of secondary structures upon heating in EstGtA2. At pH 7, 8 and 9, depending on salt concentration, a folded but non-native associated state characterised by an apparent particle size of 300 nm resisted secondary structure unfolding up to 95°C. CONCLUSION: The paths leading to various aggregative states were found to be controlled by pH (depending on proximity to pI) and to a lesser extent, ionic strength, suggesting that ionic interactions at the surface of the protein are responsible for behavior of EstGtA2. The various paths available to EstGtA2 could be important for protection of Geobacillus termodenitrificans when exposed to heat stress. The understanding and/or control of these paths would allow for optimal use of EstGtA2 in industrial processes.
[Mh] Termos MeSH primário: Carboxilesterase/química
Estabilidade Enzimática
Geobacillus/enzimologia
[Mh] Termos MeSH secundário: Sequência de Aminoácidos/genética
Carboxilesterase/genética
Carboxilesterase/metabolismo
Dicroísmo Circular
Temperatura Alta
Concentração de Íons de Hidrogênio
Concentração Osmolar
Conformação Proteica
Desnaturação Proteica
Dobramento de Proteína
Estrutura Secundária de Proteína
Cloreto de Sódio/química
Propriedades de Superfície
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
451W47IQ8X (Sodium Chloride); EC 3.1.1.1 (Carboxylesterase)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171016
[Lr] Data última revisão:
171016
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170411
[St] Status:MEDLINE
[do] DOI:10.2174/0929866524666170404162730


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[PMID]:28381218
[Au] Autor:Bacon LF; Hamley-Bennett C; Danson MJ; Leak DJ
[Ad] Endereço:Department of Biology and Biochemistry, University of Bath, Bath, BA2 7AY, UK.
[Ti] Título:Development of an efficient technique for gene deletion and allelic exchange in Geobacillus spp.
[So] Source:Microb Cell Fact;16(1):58, 2017 Apr 05.
[Is] ISSN:1475-2859
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: Geobacillus thermoglucosidasius is a thermophilic, natural ethanol producer and a potential candidate for commercial bioethanol production. Previously, G. thermoglucosidasius has been genetically modified to create an industrially-relevant ethanol production strain. However, creating chromosomal integrations and deletions in Geobacillus spp. is laborious. Here we describe a new technique to create marker-less mutations in Geobacillus utilising a novel homologous recombination process. RESULTS: Our technique incorporates counter-selection using ß-glucosidase and the synthetic substrate X-Glu, in combination with a two-step homologous recombination process where the first step is a selectable double-crossover event that deletes the target gene. We demonstrate how we have utilised this technique to delete two components of the proteinaceous shell of the Geobacillus propanediol-utilization microcompartment, and simultaneously introduce an oxygen-sensitive promoter in front of the remaining shell-component genes and confirm its functional incorporation. CONCLUSION: The selectable deletion of the target gene in the first step of our process prevents re-creation of wild-type which can occur in most homologous recombination techniques, circumventing the need for PCR screening to identify mutants. Our new technique therefore offers a faster, more efficient method of creating mutants in Geobacillus.
[Mh] Termos MeSH primário: Alelos
Deleção de Genes
Engenharia Genética/métodos
Geobacillus/genética
Recombinação Homóloga
Mutação
[Mh] Termos MeSH secundário: Etanol/metabolismo
Reação em Cadeia da Polimerase
Regiões Promotoras Genéticas
Deleção de Sequência
beta-Glucosidase/metabolismo
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
3K9958V90M (Ethanol); EC 3.2.1.21 (beta-Glucosidase)
[Em] Mês de entrada:1704
[Cu] Atualização por classe:170424
[Lr] Data última revisão:
170424
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170407
[St] Status:MEDLINE
[do] DOI:10.1186/s12934-017-0670-4



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