Deacylation of benzylpenicillin by immobilized penicillin acylase in a continuous four‐stage stirred‐tank reactor

Carleysmith, S. W.; Lilly, M. D.

doi:10.1002/bit.260210610

Cited by 41 publications

(8 citation statements)

References 9 publications

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“…The operational stability of XAD7-PA in a reactor is described elsewhere. 8 The adsorption of challenge protein by CMC ceased after about 10 min, by which time PA had probably become adsorbed throughout the support particles. However, the amount of protein adsorbed to glutaraldehyde-activated XAD7 increased almost linearly with the logarithm of the adsorption time and was lower for larger particles (Fig.…”

Section: Discussionmentioning

confidence: 99%

Kinetic behavior of immobilized Penicillin acylase

Carleysmith

Dunnill

Lilly

1980

Biotech & Bioengineering

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SummaryPenicillin acylase has been immobilized to carboxymethylcellulose and to the resin Amberlite XAM. The reaction kinetics of the enzyme were affected by both intrinsic (molecular) and microenvironmental effects. The Michaelis constant for the enzyme increased after immobiiiation as a result of an intrinsic effect of the reagent, glutaraldehyde, used for enzyme immobilization. Microenvironmental effects were of two types: diffusional limitation of access of substrate and a reaction-generated pH depression in the support particles. This depression of internal pH was observed in all the preparations and could be reduced by addition of pH buffering salts to the reactor. An adsorbed pH-indicating dye was used to determine the surface and internal pH of particles of XAW-penicillin acylase under various reaction conditions. The extent of diffusional rate limitation in XAD7-penicillin acylase was related to the penetration depth of protein into the porous support particles. The penetration depth of protein and thus the diffusional limitation of the reaction rate could be controlled by the conditions of preparation of the immobilized enzyme. A staining technique was used to observe the location of the protein.

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Section: Discussionmentioning

confidence: 99%

Kinetic behavior of immobilized Penicillin acylase

Carleysmith

Dunnill

Lilly

1980

Biotech & Bioengineering

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“…The reaction conditions, i.e., no added buffer and the addition of 5N NaOH to maintain the pH constant, were designed to cause a reasonably fast activity loss of the immobilized penicillin acylase preparation containing no BSA. 4 The second preparation with an outer layer of BSA had a greater operational stability, probably because the enzyme was more protected from the localized regions of high pH that resulted when alkali was added. Although the location of stained immobilized protein is limited to particles that can be observed under a light microscope, it does allow reasonable measurements to be made on particles of 250 pm diam or larger.…”

Section: Resultsmentioning

confidence: 99%

Staining method for determination of the penetration of immobilized enzyme into a porous support

Carleysmith

Eames

Lilly

1980

Biotech & Bioengineering

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“…Biocatalysts for selective group reactions of molecules have already been used since the 1960s and 1970s (Tosa et al 1969;Carleysmith and Lilly 1979;Hummel et al 1981). These studies applied either enzymes or cells of microbial, plant, or animal origin.…”

Section: Introductionmentioning

confidence: 98%

Enzyme-based glucose delivery as a high content screening tool in yeast-based whole-cell biocatalysis

Grimm

Grimm²,

Klat³

et al. 2012

Appl Microbiol Biotechnol

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The influence of glucose release on growth and biotransformation of yeasts was examined by using the medium EnBase® Flo in shake flasks. The medium contains a polysaccharide acting as substrate, which is degraded to glucose by the addition of an enzyme. In the present paper, this medium was adapted for the cultivation of yeasts by increasing the complex components (booster) and the enzyme concentrations to guarantee a higher glucose release rate. Important changes were an increase of the complex component booster to 10-15% and an increased glucose release by increasing the enzyme content to 15 U L(-1). The 20 yeasts investigated in the present work showed an improvement of growth and biomass production when cultivated with the EnBase medium in comparison to yeast extract dextrose (YED) medium. Values of optical densities (OD(600)) of approximately 40 AU (corresponding to over 60 g L(-1) wet cell weight) were achieved for all 20 yeast strains tested. During the following screening of the yeasts in whole-cell biotransformation, an improvement of the conversion for 19 out of the 20 yeasts cultivated with the EnBase Flo medium could be observed. The biomass from the EnBase Flo cultivation showed a higher conversion activity in the reduction of 2-butanone to (R/S)-2-butanol. The enantioselectivity (ee) of 15 yeast strains showed an improvement by using the EnBase medium. The number of yeasts with an ee >97% increased from zero with YED to six with EnBase medium. Thus, the use of a glucose release cultivation strategy in the screening process for transformation approaches provides significant benefits compared to standard batch approaches.

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Deacylation of benzylpenicillin by immobilized penicillin acylase in a continuous four‐stage stirred‐tank reactor

Cited by 41 publications

References 9 publications

Kinetic behavior of immobilized Penicillin acylase

Kinetic behavior of immobilized Penicillin acylase

Staining method for determination of the penetration of immobilized enzyme into a porous support

Enzyme-based glucose delivery as a high content screening tool in yeast-based whole-cell biocatalysis

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