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Results for “Enzyme Stability”

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At least 19 recordsLinked to original sources
PubMed1978

[Chemical modification of lysine epsilon-NH2-groups in horseradish peroxidase. Its effect on enzyme stability. Temperature dependence of thermo-inactivation constants for native and modified peroxidase].

Thermostability of horseradish peroxidase modified by acetic, propionic, butyric, valeric and succinic anhydrides and trinitrobenzolsulfonic acid (TNBS) is studied within the temperature range of 56-80 degrees C. Acylation of 4 amino groups and arylation of 3 amino groups with TNBS are found to stabilize the enzyme, while modification of 6 groups decreases the enzyme stability. Chemical modification of peroxidase does not change its pH-dependence with respect to enzyme thermostability. Thermodynamic activation parameters of irreversible thermoinactivation are determined for native and modified peroxidase. Native peroxidase has deltaH not equal to = 30+/-1 kcal/mole and deltaS not equal to = 14 e. e.; modified by acid anhydrides peroxidase has deltaH not equal to within 64-87 kcal/mole and deltaS not equal to within 110-178 e. e. depending on the nature of a modifying agent. The effect of the structure of a radical introduced into the enzyme molecule, and of a number of modified epsilon-amino groups on thermoinactivation deltaH not equal to and deltaS not equal to values is discussed.

Acylation↗
PubMed1979

Principles of enzyme stabilization. V. The possibility of enzyme selfstabilization under the action of potentially reversible intramolecular cross-linkages of different length.

The denaturing action of guanidine . HCl on modified alpha-chymotrypsin (EC 3.4.21.1) preparations has been studied. The consecutive treatment of alpha-chymotrypsin with N-acetyl-homocysteine thiolactone, 5,5'-dithio-bis-(2-nitrobenzoic acid) and dithiols of HS-(CH2)n-SH type, with n ranging from 4 to 10, leads to enzyme stabilization as a result of protein modification. A greater stabilization effect can be achieved by enriching the protein molecule with groups reactive towards dithiols, after first modifying carboxygroups. In this case dithiol with n=5 forms an intramolecular cross-linkage. If an equimolecular mixture of different dithiols is used for enzyme modification, the enzyme gradually 'selects' 1,5-dithiol for the formation of an intramolecular cross-linkage instead of the initial one-point modification. The use of potentially reversible cross-linkages may be generally employed for the preparation of stabilized water-soluble enzymes via the mechanism of selfstabilization.

Chymotrypsin↗
PubMed1991

Enzyme stabilization: state of the art.

'Enzyme stabilization' is one of the most important fields in basic and applied enzymology. In basic enzymology, it is of particular relevance to understand enzyme stabilization principles first elucidating how and why the enzymes lose their biological activity and then deriving structure-stability relationships existing in enzymatic molecules. In applied enzymology, the most significant goal is to achieve useful compounds by biocatalysis. Enzymes are good catalysts in terms of high catalytic and specific activity with ability to function under mild conditions. However, they are not always ideal catalysts for practical applications because they are generally unstable and they inactivate rapidly through several mechanisms. In order to enhance enzyme stability, many strategies have been pursued in recent years. The present article is an attempt to provide detailed information about these strategies.

Animals↗
PubMedDate not supplied

Stability of firefly luciferase in tricine buffer and in a commercial enzyme stabilizer.

A solution of firefly luciferase in AuthentiZyme Enzyme Stabilizer retains full activity when stored in an ice bath (0.5 degrees C) during one day. These solutions have the advantage that no additional protein (other than the luciferase) is present, which is desirable for proteolytic digestion and protein derivatization experiments. For longer-term experiments, firefly luciferase solutions in 0.05 mol/l Tricine buffer at pH 7.8, 10 mmol/l MgSO4, 1 mmol/l EDTA, and 1 mmol/l DTT which contain 100 micrograms/ml of bovine serum albumin are stable for 6 weeks if frozen and thawed only once.

Animals↗
PubMed1986

Variation in alpha-L-fucosidase properties among 28 inbred mouse strains: six strains have high enzyme activity and heat-stabile enzyme with a variant pH-activity curve; twenty-two strains have low activity and heat-labile enzyme.

Alpha-L-fucosidase in tissues of 28 inbred mouse strains varied with respect to three properties: high or low heat stability, a pH-activity curve with high or low relative activity at pH 2.8, and high or low activity. Alpha-L-fucosidase from six strains (A/J, BDP/J, LP/J, P/J, SEA/GNJ, and 129/J) had high heat stability, high pH 2.8 relative activity, and high activity, whereas the other 22 strains all had low heat stability, low pH 2.8 relative activity, and low activity. The heat-stability difference was seen in all organs tested (brain, liver, kidney, spleen, heart, skeletal muscle, lung, and testis) for two heat-stabile strains (P/J and 129/J) and four heat-labile strains (C57BL/6J, C3H/HeJ, DBA/2J, and BALB/cJ) studied in detail. The findings suggested that two structural variants of alpha-L-fucosidase, probably genetically determined, exist in these 28 inbred mouse strains, although the presence of linkage disequilibrium between alleles of tightly linked structural and regulatory genes could not be excluded.

Animals↗
PubMed1972

Glycerol as an enzyme-stabilizing agent: effects on aldehyde dehydrogenase.

The potassium-dependent aldehyde dehydrogenase (EC 1.2.1.3), from yeast is markedly altered by the addition of high concentrations of glycerol or other polyhydric alcohols to aqueous buffers. Several lines of evidence suggest that the three-dimensional structure near the active site is involved: (i) The stability of the enzyme when stored at 2 degrees , or when subjected to repeated freezing and thawing, depends upon the presence of at least 30% (v/v) glycerol. (ii) In the same solvent, the K(m) value for DPN and the binding constant for benzaldehyde decrease by 3- and 10-fold, respectively, compared with the values obtained for these substrates in fully aqueous media. (iii) Competitive inhibition by trivalent arsenicals with respect to DPN is no longer observed in glycerol; the inhibition becomes mixed and the K(i) values increase by 5- and 50-fold, respectively, with arsenite and Mapharsen. (iv) Essential sulfhydryl groups, which are easily carboxymethylated in aqueous buffers, are not readily available in either glycerol or mannitol. The data are consistent with a change in topography induced by polyhydric alcohols in which sulfhydryl groups near the DPN-binding site are displaced to a more protected environment, where their reactivity is reduced. Since the stabilizing effects of such alcohols are frequently encountered, these results may have application to other enzymes.

Aldehydes↗
PubMed1979

Kinetics of digestive enzyme stability in solid state I: application of Weibull distribution function to solid-state enzyme inactivation.

The Weibull distribution function was applied to solid-state enzyme inactivation. On Weibull probability paper (within a narrow range), the plots of the accumulated inactivation ratio of each enzyme versus time regressed to a straight line. The parameters m and k, which correspond to the type and rate of the type and rate of the inactivation, were characteristic of each enzyme. The effect of temperature and parameter reproducibility are discussed.

Drug Stability↗
PubMed1989

Formation of the 1-(S-glutathionyl)-2,4,6-trinitrocyclohexadienate anion at the active site of glutathione S-transferase: evidence for enzymic stabilization of sigma-complex intermediates in nucleophilic aromatic substitution reactions.

Formation of the Meisenheimer complex or sigma-complex [1-(S-glutathionyl)-2,4,6-trinitrocyclohexadienate] between glutathione (GSH) and 1,3,5-trinitrobenzene (TNB) can be observed at the active sites of isoenzymes 3-3 and 4-4 of rat liver GSH transferase. The spectroscopic properties (UV-visible and CD) of the enzyme-bound sigma-complex are consistent with a 1:1 complex in an asymmetric environment. Competitive inhibitors which occupy the GSH binding site (e.g., gamma-L-glutamyl-D,L-2-aminomalonylglycine) inhibit sigma-complex formation. The apparent formation constants of the sigma-complex (M) with enzyme-bound GSH (E.GS- + TNB in equilibrium E.M) at pH 7.5 are 5 x 10(4) M-1 and 7 x 10(2) M-1 for isoenzymes 3-3 and 4-4, respectively. Both values are much greater than that in aqueous solution (GS- + TNB in equilibrium M), where Kf = 28 M-1. Isoenzyme 3-3 is roughly an order of magnitude more efficient than 4-4 in catalyzing nucleophilic aromatic substitutions, a fact that appears to correlate with the ability of each enzyme to stabilize the sigma-complex. The pH dependence of Kf(app) for isoenzyme 3-3 is used to probe the ionization behavior of enzyme-bound GSH. The results are consistent with a double-ionization scheme (e.g., H+E.GSH in equilibrium H+E.GS- in equilibrium E.GS-) with pK's of 5.7 and 7.6, which are assigned to the thiol pK and the pK of a protonated base in the active site, respectively. Formation of the sigma-complex is also observed in single crystals of isoenzyme 3-3, providing a clear demonstration of the chemical competence of the crystallized enzyme. The results are discussed with respect to catalytic efficiency and the ability of the enzyme to stabilize sigma-complex intermediates in nucleophilic aromatic substitution reactions.

Animals↗