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. 1990 Jun 15;268(3):679–684. doi: 10.1042/bj2680679

The effect of substrate partitioning on the kinetics of enzymes acting in reverse micelles.

R Bru 1, A Sánchez-Ferrer 1, F García-Carmona 1
PMCID: PMC1131493  PMID: 2114098

Abstract

A theoretical model for the expression of enzymic activity in reverse micelles previously developed [Bru. Sánchez-Ferrer & García-Carmona (1989) Biochem. J. 259, 355-361] was extended in the present work. The substrate concentration in each reverse-micelle phase (free water, bound water and surfactant apolar tails) and the organic solvent was expressed as a function of the total substrate concentration, taking into account its partition coefficients, that is, partitioning of the substrate in a multiphasic system. In each phase the enzyme expresses a catalytic constant and a Km. Thus the whole reaction rate is the addition of the particular rates expressed in each domain. This model was compared with that developed for a biphasic system [Levashov, Klyachko, Pantin, Khmelnitski & Martinek (1980) Bioorg. Khim. 6, 929-943] by fitting the experimental results obtained with mushroom tyrosinase (working on both 4-t-butylcatechol and 4-methylcatechol) to the two models. The parameters which characterize reverse micelles, omega 0 (water/surfactant molar ratio) and theta (fraction of water) were investigated. The omega 0 profile was shown to be hyperbolic for both substrates. Activity towards 4-t-butylcatechol decreases as theta increases, this observation being attributable to a dilution of the substrate. A Km of 7.8 M for 4-t-butylcatechol could be calculated on the basis of the biphasic model, whereas it was 13.5 mM when calculating on the basis of our model. A new parameter, rho (= [substrate]/theta), was defined to characterize those substrates that mainly solubilize in the reverse micelle ('micellar substrates').

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Selected References

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  1. Bru R., Sánchez-Ferrer A., Garcia-Carmona F. A theoretical study on the expression of enzymic activity in reverse micelles. Biochem J. 1989 Apr 15;259(2):355–361. doi: 10.1042/bj2590355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cabanes J., García-Cánovas F., Lozano J. A., García-Carmona F. A kinetic study of the melanization pathway between L-tyrosine and dopachrome. Biochim Biophys Acta. 1987 Feb 20;923(2):187–195. doi: 10.1016/0304-4165(87)90003-1. [DOI] [PubMed] [Google Scholar]
  3. Dekker M., Hilhorst R., Laane C. Isolating enzymes by reversed micelles. Anal Biochem. 1989 May 1;178(2):217–226. doi: 10.1016/0003-2697(89)90628-3. [DOI] [PubMed] [Google Scholar]
  4. Hilhorst R., Spruijt R., Laane C., Veeger C. Rules for the regulation of enzyme activity in reserved micelles as illustrated by the conversion of apolar steroids by 20 beta-hydroxysteroid dehydrogenase. Eur J Biochem. 1984 Nov 2;144(3):459–466. doi: 10.1111/j.1432-1033.1984.tb08488.x. [DOI] [PubMed] [Google Scholar]
  5. Jimenez M., Garcia-Carmona F., Garcia-Canovas F., Iborra J. L., Lozano J. A., Martinez F. Chemical intermediates in dopamine oxidation by tyrosinase, and kinetic studies of the process. Arch Biochem Biophys. 1984 Dec;235(2):438–448. doi: 10.1016/0003-9861(84)90217-0. [DOI] [PubMed] [Google Scholar]
  6. Luisi P. L., Giomini M., Pileni M. P., Robinson B. H. Reverse micelles as hosts for proteins and small molecules. Biochim Biophys Acta. 1988 Feb 24;947(1):209–246. doi: 10.1016/0304-4157(88)90025-1. [DOI] [PubMed] [Google Scholar]
  7. Luisi P. L., Magid L. J. Solubilization of enzymes and nucleic acids in hydrocarbon micellar solutions. CRC Crit Rev Biochem. 1986;20(4):409–474. doi: 10.3109/10409238609081999. [DOI] [PubMed] [Google Scholar]
  8. Malakhova E. A., Kurganov B. I., Levashov A. V., Berezin I. V., Martinek K. Novyi podkhod k izucheniiu fermentativnykh reaktsii s uchastiem nerastvorimykh v vode substratov. Pankreaticheskaia lipaza, vkliuchennaia v obrashchennye mitselly poverkhnostno-aktivnogo veshchestva v organicheskom rastvoritele. Dokl Akad Nauk SSSR. 1983;270(2):474–477. [PubMed] [Google Scholar]
  9. Martinek K., Khmel'nitskii Iu L., Levashov A. V., Berezin I. V. Substratnaia spetsifichnost' alkogol'degidrogenazy v kolloidnom rastvore vody v organicheskom rastvoritele. Dokl Akad Nauk SSSR. 1982;263(3):737–741. [PubMed] [Google Scholar]
  10. Martinek K., Kliachko N. L., Levashov A. V., Berezin I. V. Mitselliarnaia énzimologiia. Kataliticheskaia aktivnost' peroksidazy v kolloidnom rastvore vody v organicheskom rastvoritele. Dokl Akad Nauk SSSR. 1983;269(2):491–493. [PubMed] [Google Scholar]
  11. Martinek K., Levashov A. V., Klyachko N. L., Pantin V. I., Berezin I. V. The principles of enzyme stabilization. VI. Catalysis by water-soluble enzymes entrapped into reversed micelles of surfactants in organic solvents. Biochim Biophys Acta. 1981 Jan 15;657(1):277–294. doi: 10.1016/0005-2744(81)90151-0. [DOI] [PubMed] [Google Scholar]
  12. Martinek K., Levashov A. V., Klyachko N., Khmelnitski Y. L., Berezin I. V. Micellar enzymology. Eur J Biochem. 1986 Mar 17;155(3):453–468. doi: 10.1111/j.1432-1033.1986.tb09512.x. [DOI] [PubMed] [Google Scholar]
  13. Walde P., Peng Q., Fadnavis N. W., Battistel E., Luisi P. L. Structure and activity of trypsin in reverse micelles. Eur J Biochem. 1988 Apr 15;173(2):401–409. doi: 10.1111/j.1432-1033.1988.tb14013.x. [DOI] [PubMed] [Google Scholar]

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