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1.
J Agric Food Chem ; 67(47): 13033-13039, 2019 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-31730339

RESUMO

Three new macrocyclic trichothecenes possessing rare 6'-ketal moieties, roridoxins A-C (1-3), and five known compounds (4-8) were isolated from the insect-associated fungus Myrothecium roridum. Their structures were confirmed by a combination of NMR and HRESIMS data, while their absolute configurations were unambiguously determined by single-crystal X-ray analysis and electronic circular dichroism experiments. Trichothecenes 1 and 3 showed potent antifungal activities against four strains of phytopathogenic fungi. In addition, 1, 3, 5, and 6 were found to significantly inhibit the cell growth of Candida albicans with minimal inhibitory concentration values from 8.8 to 18.5 µg/mL. Moreover, they were able to inhibit the biofilm formation of C. albicans better than the positive control.


Assuntos
Antifúngicos/química , Antifúngicos/farmacologia , Hypocreales/química , Insetos/microbiologia , Tricotecenos/química , Tricotecenos/farmacologia , Animais , Antifúngicos/metabolismo , Candida albicans/efeitos dos fármacos , Candida albicans/crescimento & desenvolvimento , Hypocreales/isolamento & purificação , Hypocreales/metabolismo , Testes de Sensibilidade Microbiana , Estrutura Molecular , Tricotecenos/metabolismo
2.
Food Chem ; 230: 291-294, 2017 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-28407913

RESUMO

A bioassay-guided fractionation of extract from Gluconobacter oxydans fermentation broth afforded Compound 1, which was identified as pyrroloquinoline quinone (PQQ) by spectroscopic methods. PQQ has been shown to enhance the superoxide anion-scavenging capacity significantly for Cu/Zn-SOD. To illustrate the mechanism, the interaction between PQQ and Cu/Zn-SOD was investigated. The multiple binding sites involving hydrogen bonds and van der Waals force between PQQ and Cu/Zn-SOD were revealed by isothermal titration calorimetry. The α-helix content was increased in the Cu/Zn-SOD structure with the addition of PQQ into the solution through ultraviolet (UV) spectroscopy. These results indicated that PQQ could change the conformation of Cu/Zn-SOD through interaction, which could enhance its superoxide anion-scavenging capacity. Therefore, PQQ is a potential natural antioxidant.


Assuntos
Gluconobacter oxydans/química , Cofator PQQ/química , Superóxido Dismutase/química , Superóxidos/química , Zinco/química , Animais , Fermentação , Espécies Reativas de Oxigênio
3.
Biotechnol Appl Biochem ; 64(4): 525-531, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27222303

RESUMO

A glycoside hydrolase from Penicillium oxalicum BL 3005 was purified to apparent homogeneity. Its molecular mass was estimated to be 90 kDa by SDS-PAGE. The enzyme was identified to be a new member of family-3 by peptide sequence. High transglycosylation activity was found in the hydrolytic reaction of cellobiose. In the reaction, salidroside (4-hydroxyphenethyl O-ß-d-glucopyranoside) was formed by adding tyrosol as the glycosyl acceptor. The optimum reaction pH and temperature were pH 6.5 and 55 °C, respectively. The maximum yield of salidroside was almost 20 g/L. These results indicated that the ß-glucosidase of P. oxalicum can be considered as a very promising catalyst for the synthesis of salidroside.


Assuntos
Biocatálise , Glucosídeos/biossíntese , Glicosídeo Hidrolases/metabolismo , Penicillium/enzimologia , Álcool Feniletílico/análogos & derivados , Eletroforese em Gel de Poliacrilamida , Glucosídeos/química , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/isolamento & purificação , Glicosilação , Peso Molecular , Fenóis/química , Álcool Feniletílico/química , Álcool Feniletílico/metabolismo
4.
Science ; 354(6310): 339-342, 2016 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-27846569

RESUMO

Methyl-coenzyme M reductase (MCR) is the key enzyme of methanogenesis and anaerobic methane oxidation. The activity of MCR is dependent on the unique nickel-containing tetrapyrrole known as coenzyme F430. We used comparative genomics to identify the coenzyme F430 biosynthesis (cfb) genes and characterized the encoded enzymes from Methanosarcina acetivorans C2A. The pathway involves nickelochelation by a nickel-specific chelatase, followed by amidation to form Ni-sirohydrochlorin a,c-diamide. Next, a primitive homolog of nitrogenase mediates a six-electron reduction and γ-lactamization reaction before a Mur ligase homolog forms the six-membered carbocyclic ring in the final step of the pathway. These data show that coenzyme F430 can be synthesized from sirohydrochlorin using Cfb enzymes produced heterologously in a nonmethanogen host and identify several targets for inhibitors of biological methane formation.


Assuntos
Proteínas Arqueais/metabolismo , Metaloporfirinas/metabolismo , Metano/metabolismo , Methanosarcina/enzimologia , Oxirredutases/metabolismo , Uroporfirinas/metabolismo , Amidas/metabolismo , Proteínas Arqueais/genética , Vias Biossintéticas , Genes Arqueais , Loci Gênicos , Genômica , Metaloporfirinas/genética , Methanosarcina/genética , Níquel/metabolismo , Oxirredutases/genética
5.
Biotechnol Lett ; 37(8): 1687-92, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26067662

RESUMO

OBJECTIVES: To investigate the conversion of lutein, a carotenoid, to aroma compounds by Pantoea dispersa Y08, a lutein-degrading bacterium isolated from marigold flower residue. Bioconversion conditions, including substrate concentration, applied co-solvent and reaction time, were optimized. RESULTS: A maximum biodegradation yield of 80 % for lutein at 10 g/l was achieved. The intermediate, 3-hydroxy-ß-ionone, and final ß-ionone products were revealed by GC-MS. A bioconversion pathway of lutein is proposed to involve cleavage at the 9-10 double bond position, followed by de-hydroxylation at the 3-hydroxy position. CONCLUSIONS: This is the first report of the ability of a bacterium, P. dispersa, to sequentially convert lutein to 3-hydroxy-ß-ionone and then ß-ionone.


Assuntos
Luteína/metabolismo , Norisoprenoides/metabolismo , Pantoea/metabolismo , Compostos Orgânicos Voláteis/metabolismo , Biotransformação , Cromatografia Gasosa-Espectrometria de Massas , Redes e Vias Metabólicas
6.
J Ind Microbiol Biotechnol ; 37(6): 575-80, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-20213113

RESUMO

We have expressed the pqqABCDE gene cluster from Gluconobacter oxydans, which is involved in pyrroloquinoline quinone (PQQ) biosynthesis, in Escherichia coli, resulting in PQQ accumulation in the medium. Since the gene cluster does not include the tldD gene needed for PQQ production, this result suggests that the E. coli tldD gene, which shows high homology to the G. oxydans tldD gene, carries out that function. The synthesis of PQQ activated d-glucose dehydrogenase in E. coli and the growth of the recombinant was improved. In an attempt to increase the production of PQQ, which acts as a vitamin or growth factor, we transformed E. coli with various recombinant plasmids, resulting in the overproduction of the PQQ synthesis enzymes and, consequently, PQQ accumulation--up to 6 mM--in the medium. This yield is 21.5-fold higher than that obtained in previous studies.


Assuntos
Clonagem Molecular/métodos , Escherichia coli/genética , Gluconobacter oxydans/genética , Cofator PQQ/biossíntese , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Genes Bacterianos , Vetores Genéticos , Gluconobacter oxydans/metabolismo , Glucose 1-Desidrogenase/metabolismo , Família Multigênica , Cofator PQQ/genética , Plasmídeos
7.
Appl Environ Microbiol ; 74(16): 5250-3, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18502922

RESUMO

A membrane-bound protein purified from Gluconobacter oxydans M5 was confirmed to be a pyrroloquinoline quinone-dependent D-sorbitol dehydrogenase. Gene disruption and complementation experiments demonstrated that this enzyme is responsible for the oxidation of 1-(2-hydroxyethyl) amino-1-deoxy-D-sorbitol (1NSL) to 6-(2-hydroxyethyl) amino-6-deoxy-L-sorbose (6NSE), which is the precursor of an antidiabetic drug, miglitol.


Assuntos
Proteínas de Bactérias/metabolismo , Gluconobacter oxydans/enzimologia , L-Iditol 2-Desidrogenase/metabolismo , Cofator PQQ/metabolismo , Sorbose/análogos & derivados , Sorbose/biossíntese , Sequência de Aminoácidos , Proteínas de Bactérias/isolamento & purificação , Teste de Complementação Genética , Vetores Genéticos , Gluconobacter oxydans/genética , L-Iditol 2-Desidrogenase/isolamento & purificação , Proteínas de Membrana/metabolismo , Oxirredução , Plasmídeos , Sorbitol/metabolismo
8.
Arch Biochem Biophys ; 477(2): 206-10, 2008 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-18407824

RESUMO

A membrane-bound pyrroloquinoline quinine (PQQ)-dependent D-sorbitol dehydrogenase (mSLDH) in Gluconobacter oxydans participates in the oxidation of D-sorbitol to L-sorbose by transferring electrons to ubiquinone which links to the respiratory chain. To elucidate the kinetic mechanism, the enzyme purified was subjected to two-substrate steady-state kinetic analysis, product and substrate inhibition studies. These kinetic data indicate that the catalytic reaction follows an ordered Bi Bi mechanism, where the substrates bind to the enzyme in a defined order (first ubiquinone followed by D-sorbitol), while products are released in sequence (first L-sorbose followed by ubiquinol). From these findings, we proposed that the native mSLDH bears two different substrate-binding sites, one for ubiquinone and the other for D-sorbitol, in addition to PQQ-binding and Mg(2+)-binding sites in the catalytic center.


Assuntos
Membrana Celular/enzimologia , Gluconobacter oxydans/enzimologia , L-Iditol 2-Desidrogenase/química , Cofator PQQ/química , Sorbitol/química , Ubiquinona/química , Catálise , Ativação Enzimática , Estabilidade Enzimática , Cinética , Especificidade por Substrato
9.
Bioprocess Biosyst Eng ; 29(5-6): 379-83, 2006 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17082915

RESUMO

Several ionic liquids were used as reaction media for penicillin G acylase catalysis. In all the assayed ionic liquids, [bmim]PF6 proved good media for PGA-catalyzed hydrolysis. A novel [bmim]PF6/water two-phase system is provided for 6-aminopenicillanic acid (APA) production, which will be more benefical than aquous batch systems used widely in industrial production of APA.


Assuntos
Escherichia coli/enzimologia , Líquidos Iônicos/química , Penicilina Amidase/química , Penicilina G/química , Catálise , Ativação Enzimática , Estabilidade Enzimática , Hidrólise , Especificidade por Substrato
10.
Sheng Wu Gong Cheng Xue Bao ; 21(1): 84-91, 2005 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-15859334

RESUMO

Beta-glycosidase (Tngly) from the thermophilic eubacterium Thermus nonproteolyticus HG102, which is a thermostable monomeric protein and adopts the (beta/alpha)8 barrel fold, is an excellent model system to be investigated for the thermostable mechanism, activity and substrate specificity. Here, based on the analysis of structural basis for thermostability of Tngly (Wang et al, 2003) and comparison of other proteins structure of homofamily, Glu164 and Glu338 may act as proton donor and nucleophile in the hydrolysis reaction respectively; proline located at N1 of alpha-helix and arginine which can form ion link may contribute to the thermostability. We aim to further identify the critical sites and the amino acid residue(s) responsible for the activity, the thermal stability and the substrate specificity. Mutations had been constructed by site-directed mutagenesis. They are Glu164Gln, Glu338Ala, Pro316Gly, Arg325Leu, Pro344Phe, Pro356Ala and Pro316Gly/Pro356Ala. All mutant proteins were purified to SDS-PAGE purity. Changes in the conformations were examined by means of CD. The Glu338Ala mutant showed no detectable hydrolysis activity, but can synthesize oligosaccharides, as expected for the residue acting as the nucleophile of the reaction. The Glu164 acts as the general acid/base catalyst in the hydrolysis reaction. Changes in stabilities of mutants compared with wild-type were determined by means of heat inactivity experiment. These results indicate that the amino acid residue of proline that is located at N1 positions of alpha-helix, and Arg325 that form salt bridge between alpha-helices 5 and alpha-helices 6, are the critical sites to protein thermostabilization.


Assuntos
Proteínas de Bactérias/metabolismo , Temperatura Alta , Mutação , Thermus/enzimologia , beta-Glucosidase/metabolismo , Proteínas de Bactérias/genética , Estabilidade Enzimática , Hidrólise , Mutagênese Sítio-Dirigida , Relação Estrutura-Atividade , Thermus/genética , beta-Glucosidase/genética
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