Purification and Characterization of a Fibrinolytic Protease from a Culture Supernatant of<i>Flammulina velutipes</i>Mycelia

Park, Se-Eun; Li, Meihong; Kim, Jae-Sung; Sapkota, Kumar; Kim, Jieun; Choi, Bong-Suk; Yoon, Yeon-Hee; Lee, Jin-Cheol; Lee, Hyun-Hwa; Kim, Chun-Sung; Kim, Sungjun

doi:10.1271/bbb.70193

Cited by 59 publications

(41 citation statements)

References 41 publications

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“…The fibrinolytic protease from Flammulina velutipes hydrolyzed fibrin digesting α-chain over β-and γ-γ-chain. This hydrolysis pattern is similar to that of plasmin [22]. This paper emphasizes the production of the enzyme with good yield, extracellular in nature for simple recovery and enzyme characterization, and potent in nature with respect to ex vivo clot lysis.…”

Section: Discussionmentioning

confidence: 60%

Production, Purification, and Biochemical Characterization of a Fibrinolytic Enzyme from Thermophilic Streptomyces sp. MCMB-379

Chitte

Deshmukh

Kanekar

2011

Appl Biochem Biotechnol

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Production of the fibrinolytic enzyme was carried out using 2.5-L glass fermentor, culture of thermophilic Streptomyces sp., and glucose yeast extract peptone medium of pH 8.0. Five successive batches were carried out under controlled fermentation conditions viz., agitation 140 rpm, aeration 0.5 vvm, 55 °C, and 18 h. The total protein extracellularly produced in the cell-free broth was ~300-500 mg/L. The enzyme belongs to serine endopeptidase type. Studies on the fibrin degradation indicate that the enzyme degrades the fibrin into small molecular weight products as seen from HPLC profile. Phase-contrast microscopic structure of fibrin showed that enzyme cleaves the fibrin filaments. The ex vivo activity of the actinokinase was compared with 500 IU of urokinase and 350 IU of streptokinase. The ex vivo clot lysis was found to be faster as compared to the commercial available enzymes.

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Section: Discussionmentioning

confidence: 60%

Production, Purification, and Biochemical Characterization of a Fibrinolytic Enzyme from Thermophilic Streptomyces sp. MCMB-379

Chitte

Deshmukh

Kanekar

2011

Appl Biochem Biotechnol

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“…Além disso, muitos produtos farmacêuticos de ação fibrinolítica utilizados atualmente possuem efeitos colaterais indesejáveis, tais como hemorragia intestinal no tratamento via oral e baixa especificidade à fibrina, além de custos relativamente altos. Dessa forma, a atividade fibrinolítica de proteases produzidas por micro-organismos vem despertando maior interesse médico e comercial 5 .…”

Section: Introductionunclassified

Produção de Protease e Lacase por Basidiomicetos

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Bueno

Nichida

et al. 2013

BBR - Biochem. Biotechnol.

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RESUMOExiste um crescente interesse pelas proteases devido à possibilidade de aplicação industrial das mesmas. Nas indústrias, as proteases contribuem para o desenvolvimento de processos ou produtos de alto valor agregado. Além disso, muitos produtos de ação fibrinolítica utilizados atualmente possuem efeitos colaterais indesejáveis. Este trabalho teve como objetivo estudar a produção de protease e lacase por basidiomicetos cultivados em resíduos agroindustriais. Além disso, foi avaliada a atividade fibrinolítica de um extrato enzimático bruto através de metodologia em placa. P. pulmonarius, G. lucidum e Trametes sp. produziram 161,8±6,2; 175,6±2,0 e 192,5±2,6 U/mL, respectivamente. O extrato bruto de G. lucidum apresentou atividade fibrinolítica, com formação de halo de hidrólise inclusive mais expressivo que na placa controle, com plasmina. Os dados obtidos devem ser considerados significantes pelo fato de que trabalhos recentes da literatura têm enfatizado tal capacidade proteolítica e fibrinolítica a partir de extratos enzimáticos purificados.Palavras-chave: enzimas, Ganoderma, atividade fibrinolítica, basidiomicetos, protease. INTRODUÇÃOA proteólise é um processo essencial para todos os organismos vivos. Crescimento e morte celular, coagulação sanguínea e defesa imunológica são alguns exemplos da importância das proteases na manutenção da homeostase nos indivíduos 1 . As proteases são enzimas que catalisam reações hidrolíticas, onde as moléculas de proteínas são degradadas em peptídios e aminoácidos 2 . Existe um crescente interesse por tais enzimas devido à possibilidade de aplicação industrial das mesmas. Suas propriedades, como especificidade por substrato, temperatura e pH ótimos de atividade, mecanismo catalítico e estabilidade, diferem muito devido a esse grupo ser grande e complexo 3 . Das enzimas industriais, 75% são hidrolases, sendo que as proteases representam aproximadamente 40% do total das hidrolases utilizadas na indústria e são responsáveis por aproximadamente 60% da venda total de enzimas no mundo 2 . Nas indústrias, as proteases contribuem para o desenvolvimento de processos ou produtos de alto valor agregado 4 . Além disso, muitos produtos farmacêuticos de ação fibrinolítica utilizados atualmente possuem efeitos colaterais indesejáveis, tais como hemorragia intestinal no tratamento via oral e baixa especificidade à fibrina, além de custos relativamente altos. Dessa forma, a atividade fibrinolítica de proteases produzidas por micro-organismos vem despertando maior interesse médico e comercial 5 .

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“…They are commonly used as food, food flavoring, and also in traditional Chinese medicines (Park et al 2007). Eight million tons a year are consumed.…”

Section: Introductionmentioning

confidence: 99%

A survey of proteases in edible mushrooms with synthetic peptides as substrates

2011

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The protease activities in six edible mushrooms were surveyed using synthetic fluorogenic substrates that have different specificities for each protease group. The activity was determined by measuring the fluorogenic intensity of the 7-amino-4-methylcoumarin (AMC) liberated by an enzyme. Various types of activities were found in all mushrooms, and their activities depended largely on the mushroom species, but also on the pH and localization. Flammulina velutipes and Pleurotus eryngii had the widest and highest proteolytic activities among the six mushrooms examined. The proteasome-like protease activities were generally much higher than those of other proteases. High caspase activities, which occur during apoptosis in cells, were detected in two mushrooms, F. velutipes and Hypsizigus marmoreus. The pH optima of the proteolytic activities were largely divided into two groups, acidic pH 5-6 for caspases and neutral to alkaline (pH 6.5-11) for the others. In F. velutipes, higher proteolytic activity was observed in the basement of the stem than in the cap and stem. Purification and characterization of protease were also carried out to identify a protease from Grifola frondosa using t-butyloxycarbonyl-Leu-Arg-Arg-4-methylcoumaryl-7-amide (Boc-LRR-MCA) as the substrate.

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Purification and Characterization of a Fibrinolytic Protease from a Culture Supernatant ofFlammulina velutipesMycelia

Cited by 59 publications

References 41 publications

Production, Purification, and Biochemical Characterization of a Fibrinolytic Enzyme from Thermophilic Streptomyces sp. MCMB-379

Production, Purification, and Biochemical Characterization of a Fibrinolytic Enzyme from Thermophilic Streptomyces sp. MCMB-379

Produção de Protease e Lacase por Basidiomicetos

A survey of proteases in edible mushrooms with synthetic peptides as substrates

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