Immobilized enzyme reactor chromatography: Optimization of protein retention and enzyme activity in monolithic silica stationary phases

Besanger, Travis R.; Hodgson, Richard; Green, James R. A.; Brennan, John D.

doi:10.1016/j.aca.2005.12.066

Cited by 48 publications

(54 citation statements)

References 67 publications

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“…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. 122,124 A imobilização em microcápsula pode ser obtida pelo envolvimento das enzimas por membranas semipermeáveis, ou por micelas reversas formadas por surfactantes.…”

Section: Métodos De Imobilizaçãounclassified

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

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

Cardoso¹,

Moraes²,

Cass³

2009

Quím. Nova

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“…[58][59][60][61] The silicabased monolithic column material showed good chromatographic properties of dual pore size distribution; large macropores and thin, highly porous skeleton. In addition, this material offers high permeability, high mechanical strength, and good organic solvent tolerance.…”

Section: 47mentioning

confidence: 99%

Protein Analysis Using a Combination of an Online Monolithic Trypsin Immobilized Enzyme Reactor and Collisionally-Activated Dissociation/Electron Transfer Dissociation Dual Tandem Mass Spectrometry

Hwang¹,

Cho²,

Kim³

et al. 2012

Bulletin of the Korean Chemical Society

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We demonstrated the combined applications of online protein digestion using trypsin immobilized enzyme reactor (IMER) and dual tandem mass spectrometry with collisionally activated dissociation (CAD) and electron transfer dissociation (ETD) for tryptic peptides eluted through the trypsin-IMER. For the trypsin-IMER, the organic and inorganic hybrid monolithic material was used. By employing the trypsin-IMER, the long digestion time could be saved with little or no sacrifice of the digestion efficiency, which was demonstrated for standard protein samples. For three model proteins (cytochrome c, carbonic anhydrase, and bovine serum albumin), the tryptic peptides digested by the IMER were analyzed using LC-MS/MS with the dual application of CAD and ETD. As previously shown by others, the dual application of CAD and ETD increased the sequence coverage in comparison with CAD application only. In particular, ETD was very useful for the analysis of highly-protontated peptide cations, e.g., ≥ 3+. The combination approach provided the advantages of both trypsin-IMER and CAD/ETD dual tandem mass spectrometry applications, which are rapid digestion (i.e., 10 min), good digestion efficiency, online coupling of trypsin-IMER and liquid chromatography, and high sequence coverage.

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“…[35,36] Silica as an enzyme-immobilization support for example, reported a mesoporous silica monolith that generates a back pressure of >3500 psi at flow rates of less than 1 µl/min, which excludes further integration with pressure-driven liquid chromatography. [19] The authors attempted to address the limitation using sol-gel synthesis; using glyceroxysilanes to create mesoporous materials that combines high protein retention of α-glutamyl transpeptidase (loading efficiency >80%) with enhanced flow characteristics. One limitation of this approach was long preparation times of several days, due to multiple aging steps.…”

Section: Immobilized Enzyme Reactorsmentioning

confidence: 99%

“…[17,18] The limitation has been resolved for some enzymes by fabrication of support matrices with pore sizes that are specifically tailored to allow substrate flow but prevent enzyme leaching. [19] Biosilicification is the formation of nano-patterned siliceous structures in biological systems and is mediated by proteins such as silaffins and silicateins, found in marine diatoms and sponges, respectively. [20][21][22] Using biological molecules as scaffold templates, biomimetic silicification reactions have been shown to provide a simple and effective alternative for enzyme encapsulation.…”

Section: Enzyme Immobilizationmentioning

confidence: 99%

“…The apparent catalytic activity of the immobilized enzyme, however, was comparable to soluble enzyme for the conversion of L-glutamic acid pnitroanilide to p-nitroaniline. [19] The sol-gel method can be used to produce an aggregate of spherical particles that occurs due to phase separation during sol-gel transition, resulting in a coarse porous matrix with good flow characteristics when packed into a flow-through column.…”

Section: Immobilized Enzyme Reactorsmentioning

confidence: 99%

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Silica-Immobilized Enzyme Reactors

Luckarift¹

2008

Journal of Liquid Chromatography & Related Technologies

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Standard Form 298 (Rev. 8/98) REPORT DOCUMENTATION PAGEPrescribed by ANSI Std. Z39.18 Form Approved OMB No. 0704-0188The public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing the collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing the burden, The effect of various silica matrices and immobilization techniques upon the enzymatic properties and stability of the biocatalysts is considered. In addition, reports in which the carrier matrix is biologically-derived silica are highlighted as an alternative and versatile technique that provides advantageous recovery, reuse and reproducibility. PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) 8. PERFORMING ORGANIZATION REPORT NUMBER SPONSORING/MONITORING AGENCY NAME(S) AND ADDRESS(ES) 10. SPONSOR/MONITOR'S ACRONYM(S) SPONSOR/MONITOR'S REPORT NUMBER(S)

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Immobilized enzyme reactor chromatography: Optimization of protein retention and enzyme activity in monolithic silica stationary phases

Cited by 48 publications

References 67 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

Protein Analysis Using a Combination of an Online Monolithic Trypsin Immobilized Enzyme Reactor and Collisionally-Activated Dissociation/Electron Transfer Dissociation Dual Tandem Mass Spectrometry

Silica-Immobilized Enzyme Reactors

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