Immobilization of glucoamylase by adsorption on carbon supports and its application for heterogeneous hydrolysis of dextrin

Коваленко, Г. А.; Perminova, L. V.

doi:10.1016/j.carres.2008.02.006

Cited by 18 publications

(19 citation statements)

References 10 publications

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“…The amount of GA adsorbed onto B 203 was 171 GA U per gram of support and the expressed activity 34 GA U per gram of support, which was 20% of the total immobilized activity. Immobilization efficiency (20%) was similar to that reported for GA adsorption on synthetic carbon material (Kovalenko and Perminova 2008). Both values (adsorbed and expressed activity) were lower than those attained, at laboratory scale, for tests performed to determine maximal support capacity in which adsorbed GA varied from 226 to 258 GA U per gram of support and expressed activity between 49 and 52 GA U per gram of support (Table 5).…”

Section: Bench-scale Ga Immobilization Onto Chicken Bone Particlessupporting

confidence: 63%

“…3 Effect of adsorption temperature: 20°C (filled diamonds) and 51.5°C (empty squares) on the immobilized activity of GA onto bone (d p , 203 µm) free and immobilized GA followed Michaelis-Menten kinetics; however, adsorption onto bone support increased the K M about nine times (from 0.08 to 0.71 mg soluble starch ml −1 ). Different K M variations ranging from a 10% reduction to a 69% increment were reported for biocatalysts produced by GA adsorption onto active carbon (Rani et al 2000), bone particles of 600-2,000 µm (Schafhauser and Storey 1992b), gelatinized corn starch plus subsequent trapping in alginate fibers (Tanriseven et al 2002), and synthetic carbon material Sibunit® (Kovalenko and Perminova 2008). The K M increment determined in this study could be due to changes in GA three-dimensional conformation produced by immobilization and to the limited access of the macromolecular substrate (maltodextrin) to the active site of the enzyme (Bahar and Çelebi 1998;Brena 1996;Trevan 1980).…”

Section: Properties Of Ga Bone Derivativementioning

confidence: 97%

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Bone-Bound Glucoamylase as a Biocatalyst in Bench-Scale Production of Glucose Syrups from Liquefied Cassava Starch

Carpio

Escobar

Batista‐Viera

et al. 2008

Food Bioprocess Technol

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This research assesses the bench-scale application of a non-conventional support, bone particles, for glucoamylase (GA) immobilization and its subsequent use in cassava starch hydrolysis. Upon determining the appropriate conditions to immobilize GA onto chicken bone particles, such as pH, ionic strength, particle size, and enzyme load, bench-scale immobilization of commercial GA without further purification was performed. Under the selected conditions, 270 GA units per gram of support were adsorbed. Optimal temperature and thermal stability of immobilized GA were only slightly different from those of the free enzyme, while optimal pH became more acidic by about one unit. The feasibility of the use of this immobilized biocatalyst for high glucose syrup production from liquefied cassava starch, at bench scale in batch process using a stirred-tank reactor, was demonstrated. Repeated use of the GA-bone derivative showed that similar conversions to those achieved with soluble enzyme (dextrose equivalent=98) were reached until the third batch and over 90% until the 25th batch.

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Section: Bench-scale Ga Immobilization Onto Chicken Bone Particlessupporting

confidence: 63%

Section: Properties Of Ga Bone Derivativementioning

confidence: 97%

Bone-Bound Glucoamylase as a Biocatalyst in Bench-Scale Production of Glucose Syrups from Liquefied Cassava Starch

Carpio

Escobar

Batista‐Viera

et al. 2008

Food Bioprocess Technol

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“…The obtained specific activity of CLEA-D (dextrin) reached a maximum of 102% at a dextrin concentration of 2.0 wt %, which was approximately 27% of the value previously enhanced with 0.5 wt % protection. This can be explained by the fact that immobilization interferes with the enzyme catalytic site or that diffusional problems are generally produced when immobilized enzymes are acting on macromolecular substrates [14]. Dextrin as a substrate can effectively protect active sites or maintain the enzyme in a catalytically active conformation.…”

Section: Protecting Agents Concentrationmentioning

confidence: 99%

“…Currently, a large variety of matrixes have been used in immobilized glucoamylase [7,[12][13][14]. The enzymes are linked to an insoluble matrix by chemical bonds, which generally produce very stable derivatives in which enzyme leakage is prevented.…”

Section: Introductionmentioning

confidence: 99%

Preparation of Cross-Linked Glucoamylase Aggregates Immobilization by Using Dextrin and Xanthan Gum as Protecting Agents

Jia

et al. 2016

Catalysts

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Abstract:In this paper glucoamylase from Aspergillus niger was immobilized by using a modified version of cross-linked enzyme aggregates (CLEA). The co-aggregates were cross-linked with glutaraldehyde; meanwhile dextrin and xanthan gum as protecting agents were added, which provides high affinity with the enzyme molecules. The immobilized glucoamylase was stable over a broad range of pH (3.0-8.0) and temperature (55-75˝C); dependence shows more catalytic activity than a free enzyme. The thermostability, kinetic behavior, and first-order inactivation rate constant (k i ) were investigated. The two types of protector made the immobilized glucoamylase more robust than the free form. Both of the immobilized enzymes have excellent recyclability, retaining over 45% of the relative activity after 24 runs. In addition, immobilized enzymes reduced only 40% of the initial activity after three months by the storability measure, indicating high activity.

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“…Possible explanations for the improvement of the MS analysis include an increased activity of the endoprotease in the presence of carbon as reported previously for other enzymes (49) and/or the prevention of aggregation of adsorbed, partly digested proteins in which the more hydrophobic protein core has been exposed upon endoproteolytic digestion. In a solution, such partly digested particles may be prone to form aggregates due to the (partial) uncovering of hydrophobic residues.…”

Section: Discussionmentioning

confidence: 69%

Merging Molecular Electron Microscopy and Mass Spectrometry by Carbon Film-assisted Endoproteinase Digestion

Richter

Sander

Golas

et al. 2010

Molecular & Cellular Proteomics

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Many fundamental processes in the cell are performed by complex macromolecular assemblies that comprise a large number of proteins. Numerous macromolecular assemblies are structurally rather fragile and may suffer during purification, resulting in the partial dissociation of the complexes. These limitations can be overcome by chemical fixation of the assemblies, and recently introduced protocols such as gradient fixation during ultracentrifugation (GraFix) offer advantages for the analysis of fragile macromolecular assemblies. The irreversible fixation, however, is thought to render macromolecular samples useless for studying their protein composition. We therefore developed a novel approach that possesses the advantages of fixation for structure determination by single particle electron microscopy while still allowing a correlative compositional analysis by mass spectrometry. In this method, which we call "electron microscopy carbon film-assisted digestion", macromolecular assemblies are chemically fixed and then adsorbed onto electron microscopical carbon films. Parallel, identically prepared specimens are then subjected to structural investigation by electron microscopy and proteomics analysis by mass spectrometry of the digested sample. As identical sample preparation protocols are used for electron microscopy and mass spectrometry, the results of both methods can directly be correlated. In addition, we demonstrate improved sensitivity and reproducibility of electron microscopy carbon film-assisted digestion as compared with standard protocols. We show that sample amounts of as low as 50 fmol are sufficient to obtain a comprehensive protein composition of two model complexes. We suggest our approach to be an optimization technique for the compositional analysis of macromolecules by mass spectrometry in general. Molecular & Cellular Proteomics 9:1729 -1741, 2010.An increasing number of proteins are nowadays recognized to fulfill their cellular tasks as part of macromolecular machines (1). Recent advances in the isolation procedures allow purification of many of these assemblies for further biochemical and structural analysis (2-4). Thereby, MS of peptides derived by digestion of the proteins of the assemblies (5, 6) or MS of intact assemblies (7, 8) is used to determine the protein composition, whereas electron microscopy (EM) 1 can be used to provide information about the structure of the assembly using computational averaging and reconstruction techniques (9). Available protocols to combine information derived by MS and EM therefore either analyze the endoproteolytic digestion products (10 -13) or the intact assembly (14). However, the sample preparation protocols typically differ between EM and MS. Although for MS the sample is either directly digested in solution (6), subsequent to gel electrophoretic separation of the sample (15-17) or used in an intact form (7,8), EM typically requires the adsorption of the sample to carbon film (13, 18 -20). In the case of sample heterogeneity, the utilization of thes...

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Immobilization of glucoamylase by adsorption on carbon supports and its application for heterogeneous hydrolysis of dextrin

Cited by 18 publications

References 10 publications

Bone-Bound Glucoamylase as a Biocatalyst in Bench-Scale Production of Glucose Syrups from Liquefied Cassava Starch

Bone-Bound Glucoamylase as a Biocatalyst in Bench-Scale Production of Glucose Syrups from Liquefied Cassava Starch

Preparation of Cross-Linked Glucoamylase Aggregates Immobilization by Using Dextrin and Xanthan Gum as Protecting Agents

Merging Molecular Electron Microscopy and Mass Spectrometry by Carbon Film-assisted Endoproteinase Digestion

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