The principles of enzyme stabilization

Martínek, Karel; Klibanov, Alexander M.; Goldmacher, Victor S.; Tchernysheva, A.V.; Mozhaev, Vadim V.; Березин, И.В.; Glotov, Boris O.

doi:10.1016/0005-2744(77)90189-9

Cited by 104 publications

(43 citation statements)

References 21 publications

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“…a-Chymotrypsin was covalently immobilized in a polyacrylamide gel and heated in the presence and absence of KSCN, a typical salting-in agent. It is known that rnulti-point attachment to a polymeric matrix preserves the enzyme from significant conformational changes of the unfolding type [5,291; immobilization should also arrest the transformation of a-chymotrypsin into form E,. If the stabilization mechanism shown in Fig.…”

Section: Mechanisms Of the Inactivation Of Conformations E And Ementioning

confidence: 99%

“…In the presence of salting-out solutes, form E, of the enzyme becomes more rigid [5]. Consequently, in solutions of salting-out compounds, the stability of form El should be higher with respect to the conformational processes responsible for a rapid inactivation of a-chymotrypsin according to the 'incorrect refolding' mechanism [20,25, 331. Owing to this fact, the values of k, and, consequently, k,, are slightly lower in the presence of salting-out additives than in non-saline media.…”

Section: Small Stabilization Effects Observed In the Solutions Of Salmentioning

confidence: 99%

“…a-Chymotrypsin is one of the favorite subjects for studies of the mechanisms of protein stability [l-41 and stabilization [5] (and references therein). Nevertheless, the influence of low-molecular-mass compounds on irreversible thermoinactivation of this enzyme (long-term stability) has not been studied at high temperatures (60-80°C).…”

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confidence: 99%

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Correlation of high‐temperature stability of α‐chymotrypsin with ‘salting‐in’ properties of solution

Levitsky¹,

Panova²,

Mozhaev³

1994

European Journal of Biochemistry

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A correlation between the stability of a-chymotrypsin against irreversible thermal inactivation at high temperatures (long-term stability) and the coefficient of Setchenov equation as a measure of salting-idout efficiency of solutes in the Hofmeister series has been found. An increase in the concentration of salting-in solutes (KSCN, urea, guanidinium chloride, formamide) leads to a manyfold decrease of the inactivation rate of the enzyme. In contrast, addition of salting-out solutes has a small effect on the long-term stability of a-chymotrypsin at high temperatures. The effects of solutes are additive with respect to their salting-idout capacities ; the stabilizing action of the solutes is determined by the calculated Setchenov coefficient of solution. The correlation is explained by a solute-driven shift of the conformational equilibrium between the 'low-temperature' native and the 'high-temperature' denatured forms of the enzyme within the range of the kinetic scheme put forward in the preceding paper in this journal : irreversible inactivation of the high-temperature form proceeds much more slowly compared with the low-temperature form.a-Chymotrypsin is one of the favorite subjects for studies of the mechanisms of protein stability [l-41 and stabilization [5] (and references therein). Nevertheless, the influence of low-molecular-mass compounds on irreversible thermoinactivation of this enzyme (long-term stability) has not been studied at high temperatures (60-80°C). Such studies may provide a better practical application of enzymes because it is often necessary to preserve an active enzyme from inactivation during treatment at high temperatures. The preparation of protease-containing detergents, sterilization of medicines and some other biotechnological processes which include prolonged heating can be mentioned as examples [6]. On the other hand, even at lower temperatures (4O-5O0C), the effects on proteins of urea and guanidinium chloride (GdmC1) [7-91, salts [lo-121, organic solvents [13-151 and other solutes are very different and cannot be described easily by a general theory (for one of the best recent attempts, see [16]). Thus, a better understanding of denaturation at high temperatures may be welcomed from a theoretical point of view [17].The principal possibility of regulating long-term stability by solutes may be based on the idea that enzyme inactivation is a multi-step process often involving the reversible unfolding of the protein molecule [18-201. If in the folded native state and in the unfolded reversibly denatured state an enzyme is inactivated at different rates, one may try to control inactivation by shifting the conformational equilibrium between these states. The equilibrium constant that governs the reversible unfolding of proteins is influenced significantly by low-molecular-mass additives [14, 211. An ability of ions to favor either the folded or the unfolded state is determined qualitatively by their position in the Hofmeister series, on condition that ions do not preferentially ...

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Section: Mechanisms Of the Inactivation Of Conformations E And Ementioning

confidence: 99%

Section: Small Stabilization Effects Observed In the Solutions Of Salmentioning

confidence: 99%

See 1 more Smart Citation

Correlation of high‐temperature stability of α‐chymotrypsin with ‘salting‐in’ properties of solution

Levitsky¹,

Panova²,

Mozhaev³

1994

European Journal of Biochemistry

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show abstract

“…with the boundary condition of dy/d~ 0 at On the other hand, fI, the mean sodium ion concentration, is given by Boltzmann's equation. +E(h, cpo)} (6) …”

Section: Appendix __ _mentioning

confidence: 99%

“…fllcd/n=(2/.j7i){.jr"-li~ sin ,pg/tan <Po (6) where 0 is the surface charge density, ii, the mean sodium ion concentration, K the reciprocal of the ion atmosphere radius defined by .j8rre 2 n/ckT, )' a dimensionless quantity defined by…”

Section: Effect Of Polyelectrolytes On the Thermostabilitymentioning

confidence: 99%