Screening of Inhibitors Using Enzymes Entrapped in Sol−Gel-Derived Materials

Besanger, Travis R.; Chen, Yang; Deisingh, Anil K.; Hodgson, Richard; Jin, Wen; Mayer, Stanislas; Brook, Michael A.; Brennan, John D.

doi:10.1021/ac026370i

Cited by 96 publications

(117 citation statements)

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“…A porosidade da matriz deve evitar a perda de enzima e, ao mesmo tempo, permitir o livre movimento do substrato e do produto. 38,119,120 O método de sol-gel biocompatível para a imobilização, por aprisionamento, tem sido aplicado por diferentes grupos de pesquisa em uma grande variedade de proteínas solúveis, 121,122 proteínas de membrana 29,123 e, também, no preparo de colunas de bioafinidade. 98 No processo de preparo das colunas, tanto a formulação do gel quanto as condições de processamento devem ser ajustadas para produzir materiais com diâmetros de mesoporos adequados que proporcionem o aumento da quantidade de proteínas imobilizadas, além de uma considerável melhora nas propriedades de vazão.…”

Section: Métodos De Imobilizaçãounclassified

“…98 No processo de preparo das colunas, tanto a formulação do gel quanto as condições de processamento devem ser ajustadas para produzir materiais com diâmetros de mesoporos adequados que proporcionem o aumento da quantidade de proteínas imobilizadas, além de uma considerável melhora nas propriedades de vazão. 122,124 A imobilização em microcápsula pode ser obtida pelo envolvimento das enzimas por membranas semipermeáveis, ou por micelas reversas formadas por surfactantes. As enzimas permanecem livres em solução, mas em um espaço restrito.…”

Section: Métodos De Imobilizaçãounclassified

“…A razão entre essas atividades é chamada de eficiência de imobilização (η). 29 onde: AEi = atividade da enzima imobilizada; AEs = atividade da enzima em solução; η = eficiência de imobilização, geralmente é menor que 1.…”

Section: Métodos De Imobilizaçãounclassified

“…20 Nessa segunda classe, os IMER têm sido amplamente utilizados para separação e identificação de metabólitos, 21 nos estudos de metabolismo dos fármacos, 22,23 para análise e síntese enantiosseletiva, 2,24-26 na digestão proteolítica de proteínas on-line, 27 no controle biotecnoló-gico de fármacos, 28 e para a identificação de substratos e/ou inibidores, na busca por novos ligantes. [29][30][31][32][33][34][35] Também merece destaque o uso de enzimas imobilizadas como biossensores em aplicações biomédicas 36 ou como órgãos artificiais. 37 Várias revisões na literatura têm abordado os diferentes métodos de imobilização enzimática, 15 suas aplicações industriais na química analítica 38 e na medicina, 6 abordando principalmente os aspectos analíticos das interações enzima-substrato.…”

Section: Introductionunclassified

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Imobilização de enzimas em suportes cromatográficos: uma ferramenta na busca por substâncias bioativas

Cardoso¹,

Moraes²,

Cass³

2009

Quím. Nova

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Recebido em 5/10/07; aceito em 11/7/08; publicado na web em 2/12/08 IMMOBILIZATION OF THE ENZYMES ON CHROMATOGRAPHIC SUPPORTS: A TOOL TO RESEARCH OF INHIBITOR COMPOUNDS. The development and characterization of bioreactors or IMER (immobilized enzyme reactors) as research tools are important in the scope of medicinal chemistry and constitute an alternative for the rational development of drugs. This approach does not require highly purified enzymes or a great amount of protein, but increase the enzymatic stability against heat, organic solvents and pH, without too much loss of catalyst activity. Immobilized enzyme reactors (IMER) can be used for the accomplishment of high efficiency screening on-line and, thus inhibitors can be quickly identified. Here, we emphasize the development of IMER by use of different methods of immobilization and chromatographic supports. Their applications, in different areas of research, are also fully discussed.

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Section: Métodos De Imobilizaçãounclassified

Section: Introductionunclassified

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Imobilização de enzimas em suportes cromatográficos: uma ferramenta na busca por substâncias bioativas

Cardoso¹,

Moraes²,

Cass³

2009

Quím. Nova

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“…We do so by using the sol-gel materials methodology for enzyme entrapment [1]. Sol-gel entrapped enzymes [2-4] have already been shown to provide significant thermal stability [5][6][7][8], stability to extreme pH values [9][10][11], and stability to non-native environments [12][13][14]. In these earlier studies the protectability was attributed to the encaging itself [15]; here we extend these observations by showing protection against an oxidant due to active chemical scavenging of the destructive chemical by the matrix.…”

Section: Introductionmentioning

confidence: 99%

Erratum to: Preserving the activity of enzymes under harsh oxidizing conditions: sol–gel entrapped alkaline phosphatase exposed to bromine

Frenkel-Mullerad

Ben-Knaz

Avnir

2014

J Sol-Gel Sci Technol

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Chemically reactive sol-gel matrices hold the ability of protecting entrapped enzymes from destruction by external harsh chemicals. We show this concept by exposing alkaline phosphatase (AlP) to a strong oxidizing agent-bromine. In solution, AlP is immediately destroyed by this oxidant. When AlP was entrapped in hybrid silica sol-gel materials carrying double bonds, the reactivity of AlP was preserved after exposure to bromine under conditions which totally destroy it in solution. The matrices studied were vinylated and allylated silicas, and their protectability was compared to n-alkylated silicas and to silica itself. For instance, the reactivity of AlP entrapped in allylated silica after exposure to 25.6 mM bromine solution is 40 times higher than its reactivity when entrapped in pure silica; and in solution the enzyme is totally destroyed at this concentration. Molecular level mechanisms for these observations are proposed.Keywords Sol-gel Á Sol-gel preserved enzymes Á Oxidizing agent Á Ormosil BackgroundAlkaline phosphatase (AlP), like most other enzymes, is totally destructed when exposed to a strong oxidant such as bromine. Here we show how to protect an enzyme when exposed to such conditions. We do so by using the sol-gel materials methodology for enzyme entrapment [1]. Sol-gel entrapped enzymes [2-4] have already been shown to provide significant thermal stability [5][6][7][8], stability to extreme pH values [9][10][11], and stability to non-native environments [12][13][14]. In these earlier studies the protectability was attributed to the encaging itself [15]; here we extend these observations by showing protection against an oxidant due to active chemical scavenging of the destructive chemical by the matrix. Thus when the enzyme is entrapped within sol-gel derived olefinated silicas, and then exposed to bromine solution, the double bonds react with the bromine, and protect the enzyme. By comparing this mode of protection to entrapments in non-reactive matrices-sol-gel derived silicas and sol-gel-derived alkylated silicas-we were able to evaluate the weights of the bromine scavenging and of the physical entrapment to the protection of the enzyme; we find that this relative weight depends on the alkyl and alkenyl chain. Thus, the hybrid sol-gel material we used was based on vinyl (CH 2 =CH-Si; VTS; here and below we use the name of the monomer as the name of the material), or allyl (CH 2 =CH-CH 2 -Si; ATS). In earlier studies we showed these olefinated sol-gel materials are highly reactive towards bromine, undergoing in water an hydrobromination reaction [16,17] which proceeds according to Markovnikov's rule [18], as shown in Scheme 1, resulting in a bromohydrin and HBr. The formation of an acid is an environmental change that the entrapped enzyme can withstand. We recall that we showed that sol-gel entrapped alkaline phosphataseThe online version of the original article can be found under

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Enzyme Immobilization by Microencapsulation: Methods, Materials, and Technological Applications

Rother

Nidetzky

2014

Encyclopedia of Industrial Biotechnology

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Microencapsulation as a major technique of enzyme immobilization is reviewed. Fabrication of enzyme‐containing microcapsules is broadly categorized according to whether a semipermeable membrane coating or a porous polymeric network structure is primarily responsible for enclosure of enzymes within the internal microcapsule phase. Microcapsules are mostly spherical with diameters usually in the micro‐ to millimeter range and exhibit core‐shell structures of variable complexity. Liposomes and vesicles formed from amphiphilic (co)‐polymers (polymersomes) are widely used to prepare membrane‐coated microcapsules. Hydrogels, sol–gels and other organic–inorganic hybrid materials, or layer‐by‐layer structures made through controlled assembly of polyelectrolytes are used to prepare microcapsules composed of internal polymeric networks. Method combination where polymeric network microcapsules are coated with a suitable membrane to generate tailored core‐shell structures is also used for enzyme encapsulation. Advances in the material sciences contribute to the development of microcapsules with improved properties such as enhanced morphological stability, reduced enzyme leakage, and designed physicochemical permeability and enzyme biocompatibility. Soluble enzymes, but also enzyme aggregates and other insoluble enzyme preparations, are encapsulated. Multienzyme microcapsules are interesting small‐scale bioreactors for confined and even compartmentalized cascade biotransformation. Techniques of microcapsule preparation are described, and applications of microencapsulated enzymes in biotechnology and biomedicine are discussed.

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Screening of Inhibitors Using Enzymes Entrapped in Sol−Gel-Derived Materials

Cited by 96 publications

References 61 publications

Imobilização de enzimas em suportes cromatográficos: uma ferramenta na busca por substâncias bioativas

Imobilização de enzimas em suportes cromatográficos: uma ferramenta na busca por substâncias bioativas

Erratum to: Preserving the activity of enzymes under harsh oxidizing conditions: sol–gel entrapped alkaline phosphatase exposed to bromine

Enzyme Immobilization by Microencapsulation: Methods, Materials, and Technological Applications

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