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High enzyme deactivation

One method is to run the reaction in an aqueous buffer/organic solvent biphasic system. This makes it possible to work at high substrate and product concentrations and at the pH-optimum of the enzyme. In addition, in water-immiscible solvents the non-enzymahc addition of HCN to the carbonyl group is non-existent or extremely slow. Possible disadvantages are enzyme deactivation at the interface and the presence of organic solvent dissolved in the aqueous phase [15, 17, 18]. [Pg.213]

Ionic liquids are generally regarded as highly stable, and the widely used dial-kylimidazolium ionic liquids are indeed thermostable up to 300 °C [4]. The propensity of the [BF4] and [PF6] anions to hydrolyze with liberation of HF [37], which deactivates many enzymes, has already been mentioned. The [TfO] and [ Tf2N] anions, in contrast, are hydrolytically stable. Dialkylimidazolium cations have a tendency to deprotonate at C-2, with ylide (heterocarbene) formation. Such ylides are strong nucleophiles and have been used as transesterification catalysts, for example [38]. These could cause enzyme deactivation as well as background transesterification when formed in small amounts from anhydrous ionic liquids and basic buffer salts, for example. [Pg.229]

In order to improve this reaction, a proper understanding of all parameters affecting product yield is desired. Clearly, the high enzyme consumption is a major obstacle for an efficient and economically feasible process. A likely cause of the inefficient use of DERA in this conversion is enzyme deactivation resulting from a reaction of the substrates and (by-) products with the enzyme. In general, aldehydes and (z-halo carbonyls tend to denature enzymes because of irreversible reactions with amino acid residues, especially lysine residues. From the three-dimensional structure it is known that DERA contains several solvent-accessible lysine residues [25]. Moreover, the complicated reaction profile as shown in Scheme 6.5 indicates the potential pitfalls of this reaction. [Pg.135]

Optimization of this reaction is a delicate balance between minimizing enzyme deactivation by keeping the concentration of reactants low and a high enzyme activity and productivity by adding high amounts of substrate. In order to increase the concentration of the lactol 1 at the end of the reaction the initial substrate concentration was increased in a range of 100-600 mM ClAA. At the same time... [Pg.137]

High enzyme concentration. The reaction rate and s.t.y. can be enhanced by increasing the catalyst concentration [E] in practice, however, in contrast to the formalism of Eq. (2.3), owing to an either excessive viscosity increase or excess of deactivated protein in the reactor, a maximum limit of enzyme concentration is reached. [Pg.36]

Nonaqueous enzymatic redox reactions have been limitedby stability owing to solvents and highly reactive substrates (H202). Here we have shown evidence of methods to alleviate these concerns for reactions with CPO. In experimental systems, the in situ production of H202 by GOx was shown to function equally well and more reproducibly than added H202. In situ production is experimentally easier and prevents enzyme deactivation owing to high peroxide levels. GOx was more solvent stable than CPO therefore, the GOx system may be useful for this and other redox systems. [Pg.283]

Although there are a significant number of publications reporting enzyme deactivation due to parameters such as pH and temperature, much less is known about the cause of enzyme deactivation due to hydrodynamic shear stress. Elias and Joshi [52] studied the damage produced by shear forces in enzymes and concluded that (1) some proteins deactivate only due to hydrodynamic shear (2) for those proteins, the rate of deactivation increases in the presence of gas-liquid interface (3) some proteins do not get deactivated no matter how high the applied hydrodynamic shear is, in the absence of gas-liquid interface and (4) for the proteins that need gas-liquid interface for deactivation, the rate of deactivation increases with an increase in the hydrodynamic shear. [Pg.250]

High conversion with an optimal reaction rate [7, 11, 75, 95], increase of the turnover numbers, i.e., the moles of substrate converted per mole of enzyme deactivated [3, 75, 95], and high stereospecificity of the compound of interest are targets of particular interest in the operation of these batch reactors [10,11,48, 77]. The achievement of these goals requires the study of different variables type and concentration of peroxide, substrates and cofactors, enzyme activity and purity, composition of the reaction medium, pH, temperature, or agitation. Such optimization requires a deep knowledge of the system and a mathematical model that represents it satisfactorily. The kinetic model obtained in batch experiments is the... [Pg.254]

Fig. 25.2. Analysis of the catalytic activity and the inactivation of a-chymotrypsin at the single-molecule level, (a) Detection of single enzymatic turnover events of a-chymotrpysin. The fluorogenic substrate (suc-AAPF)2-rhodamine 110 is hydrolyzed by a-chymotrypsin, yielding the highly fluorescent dye rhodamine 110. (b) Representative intensity time trace for an individual a-chymotrypsin molecule undergoing spontaneous inactivation imder reaction conditions, (c) Inactivation trace for the intensity time transient in (b), obtained by counting the amount of turnover peaks in (b) in 10 s intervals. After approximately 1000 s, the enzyme deactivates through a transient phase with discrete active and inactive states, (d) Proposed model for the inactivation process. An initial active state is in equilibrium with an inactive state. This inactive state converts to another inactive state irreversibly whereby the corresponding active state has a lower activity than the previous one. All the transitions involved have energy barriers that can be overcome spontaneously at room temperature... Fig. 25.2. Analysis of the catalytic activity and the inactivation of a-chymotrypsin at the single-molecule level, (a) Detection of single enzymatic turnover events of a-chymotrpysin. The fluorogenic substrate (suc-AAPF)2-rhodamine 110 is hydrolyzed by a-chymotrypsin, yielding the highly fluorescent dye rhodamine 110. (b) Representative intensity time trace for an individual a-chymotrypsin molecule undergoing spontaneous inactivation imder reaction conditions, (c) Inactivation trace for the intensity time transient in (b), obtained by counting the amount of turnover peaks in (b) in 10 s intervals. After approximately 1000 s, the enzyme deactivates through a transient phase with discrete active and inactive states, (d) Proposed model for the inactivation process. An initial active state is in equilibrium with an inactive state. This inactive state converts to another inactive state irreversibly whereby the corresponding active state has a lower activity than the previous one. All the transitions involved have energy barriers that can be overcome spontaneously at room temperature...
Enzyme deactivation was a key factor in determining the overall cost of the process. In a period of nine days without adding fresh d-LDH and FDH, the rate of reaction decreases slowly and the enzymes lose their activity at a rate of only about 1% per day (Fig. 11). During actual production, the reactor was charged with new enzymes periodically to get a high conversion of above 90%. [Pg.329]

Other mechanisms for enzyme denaturation in the presence of surfactants have also been proposed. One hypothesis is that the high charge densities of ionic surfactants increase the probability of them binding strongly to protein sites. This causes conformational changes of the enzyme which subsequently leads to further enzyme deactivation [99,103],... [Pg.273]

From the kinetics of the enzymatic and the non-enzymatic reactions (Fig. 7-13) it is concluded that the side-reaction is suppressed very effectively by working with high enzyme concentrations and at a low benzaldehyde concentration. Benzalde-hyde may react with amino functions of the enzyme to form Schiffbases resulting in deactivation of oxynitrilase, so low stationary benzaldehyde concentrations are also necessary with respect to enzyme stability. [Pg.246]

Enzyme deactivation is frequently encountered when highly reactive carbonyl species such as acetaldehyde or cyclohexenone are involved in the recycling process. [Pg.141]

As an example the deactivation of immobilised Pen G acylase, which catalyses the reaction of Pen G to 6-Aminopenicillanic acid and Phenylacetic acid, was studied. This enzyme was covalently bound on an ion-exchanger and cross-linked by glutaric aldehyde. To maintain a high reaction velocity, a neutral pH value (removal of Phenylacetic acid) and therefore the supply of NaOH and stirring for distribution of the base are required. [Pg.78]


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See also in sourсe #XX -- [ Pg.542 ]




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Enzymes deactivation

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