Journal ArticleDOI
The effect of Hofmeister anions and protein concentration on the activity and stability of some immobilized NAD‐dependent dehydrogenases
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TLDR
It was found that 1M phosphate and 0.5M sulfate dramatically stabilize both free and immobilized enzymes against inactivation by temperature and urea.Abstract:
The effect of several factors on the activity and stability of alcohol dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, and 20-beta-hydroxysteroid dehydrogenase, both free and immobilized on CNBr-activated Sepharose 4B, was investigated. Enzymes were immobilized under different conditions including various degrees of matrix activation, variable amounts of protein, in the presence, or in the absence of, additives (coenzymes, dithiothreitol, salts). Activity recovery was in general satisfactorily high with 20-beta-hydroxysteroid dehydrogenase, low with glyceraldehyde-3-phosphate dehydrogenase, and markedly linked to the concentration of immobilized protein with alcohol dehydrogenase. In the latter case the advantageous stabilizing effect of high enzyme concentrations was notably diminished by the paralled decrease of the effectiveness factor. The effect of high concentrations of anions of the Hofmeister series was examined. It was found that 1M phosphate and 0.5M sulfate dramatically stabilize both free and immobilized enzymes against inactivation by temperature and urea. Km values of apolar substrates were considerably lowered by the two anions while Km values of polar substrates were not affected. In some cases Vmax values also were influenced by high concentrations of these anions. The present results appear of interest particularly in view of enzyme utilization for analytical as well as for preparative purposes. (Refs. 13).read more
Citations
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Journal ArticleDOI
Categorization of enzyme deactivations using a series-type mechanism
James P. Henley,Ajit Sadana +1 more
TL;DR: A series-type enzyme deactivation model involving an active enzyme precursor and a final enzyme state with possible non-zero activity is proposed to categorize enzyme de activation curves.
Journal ArticleDOI
Use of isolated cyclohexanone monooxygenase from recombinant Escherichia coli as a biocatalyst for Baeyer-Villiger and sulfide oxidations.
TL;DR: The performance, in Baeyer-Villiger and heteroatom oxidations, of a partially purified preparation of cyclohexanone monooxygenase obtained from an Escherichia coli strain in which the gene of the enzyme was cloned and overexpressed was investigated.
Journal ArticleDOI
Analysis of the mechanism and kinetics of thermal inactivation of enzymes: Critical assessment of isothermal inactivation experiments
Milan Polakovič,Peter Vrábel +1 more
TL;DR: Carrying out inactivation experiments at several temperatures was shown to be very effective in checking the consistency of kinetic equations obtained by isothermal evaluation, and no consequent conclusions on inactivation mechanism could be drawn.
Journal ArticleDOI
A mathematical analysis of enzyme stabilization by a series‐type mechanism: Influence of chemical modifiers
James P. Henley,Ajit Sadana +1 more
TL;DR: A series‐type enzyme deactivation model is utilized to theoretically analyze and to quantify the effect of chemical modifier concentration on the eventual level of enzyme activity stabilization, α2.
Journal ArticleDOI
Structure-function relationships in the inorganic salt-induced precipitation of α-chymotrypsin
TL;DR: In this paper, the effects of salt-induced precipitation on protein conformation were investigated using spin-labeled α-Chymotrypsin (αCT) precipitates.
References
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Book
Methods of Enzymatic Analysis
TL;DR: Methods of enzymatic analysis, Methods of enzymes analysis, the authors, Methods of enzyme analysis, enzymatics, methods of enzymes, and methods of analysis, method of enzymes.
Journal ArticleDOI
Chemical Coupling of Peptides and Proteins to Polysaccharides by Means of Cyanogen Halides
TL;DR: Biologically active proteins and polypeptides can be coupled by various means to cellulose, starch and cross-linked polysaccharide gels such as ‘Sephadex’.
Journal ArticleDOI
Effect of neutral salts on enzyme activity and structure.
James C. Warren,Sarah G. Cheatum +1 more
Journal ArticleDOI
Enzymatic preparation of 20 beta-hydroxysteroids in a two-phase system.
TL;DR: The behavior of 20β–hydroxysteroid dehydrogenase in a two–phase system consisting of an emulsion of water with an immiscible organic solvent was investigated and the most suitable system for carrying out the enzymatic reaction proved to be water–butyl acetate.
Journal ArticleDOI
Immobilized hydroxysteroid dehydrogenases for the transformation of steroids in water–organic solvent systems
TL;DR: Immobilized hydroxysteroid dehydrogenases were shown to be suitable for preparative transformation of steroids in water–organic solvent systems and lyophilization in the presence of sucrose had full activity restored even after several months storage at room temperature.
Related Papers (5)
Series-type enzyme deactivations: Influence of intermediate activity on deactivation kinetics
James P. Henley,Ajit Sadana +1 more