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Title: Effect of crowding by dextrans on the hydrolysis of N-Succinyl-L-phenyl-Ala-p-nitroanilide catalyzed by α-chymotrypsin
Author: Pastor, Isabel
Vilaseca i Font, Eudald
Madurga Díez, Sergio
Garcés, Josep Lluís
Cascante i Serratosa, Marta
Mas i Pujadas, Francesc
Keywords: Cinètica enzimàtica
Enzyme kinetics
Issue Date: 2011
Publisher: American Chemical Society
Abstract: Traditionally, studies on the diffusion-controlled reaction of biological macromolecules have been carried out in dilute solutions (in vitro). However, in an intracellular environment (in vivo), there is a high concentration of macromolecules, which results in nonspecific interactions (macromolecular crowding). This affects the kinetics and thermodynamics of the reactions that occur in these systems. In this paper, we study the crowding effect of large macromolecules on the reaction rates of the hydrolysis of N-succinyl-L-phenyl-Ala-p-nitroanilide catalyzed by R-chymo- trypsin, by adding dextrans of various molecular weights to the reaction solutions. The results indicate that the volume occupied by the crowding agent, but not its size, plays an important role in the rate of this reaction. A vmax decay and a Km increase were obtained when the dextran concentration in the sample was increased. The increase in Km can be attributed to the slowing of protein diffusion, due to the presence of crowding. Whereas the decrease in vmax could be explained by the effect of mixed inhibition by product, which is enhanced in crowded media. As far as we know, this is the first reported experiment on the crowding effect in an enzymatic reaction with a mixed inhibition by product.
Note: Versió postprint del document publicat a:
It is part of: Journal of Physical Chemistry B, 2011, vol. 115, num. 5, p. 1115-1121
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ISSN: 1520-6106
Appears in Collections:Articles publicats en revistes (Ciència dels Materials i Química Física)

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