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Kinetic studies of the urease-catalyzed hydrolysis of urea in a buffer-free system

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Abstract

The kinetics of urea hydrolysis catalyzed by urease, mainly in the absence of buffers by use of the self-buffer effect of the products, was investigated. The effect of pH, temperature, and concentration of enzyme, substrate, product, salt ions, and buffers on the kinetic behavior of urease was examined. A kinetic model of a modified Michaelis-Menten form, incorporating substrate and product inhibition, pH dependence, and temperature effect, was developed to describe the reaction rate. Experimental data indicated that urease in a buffer-free solution was less susceptible to the inhibition of substrate product. The Michaelis constant keeps almost constant with the variation of pH and temperature, and increases with the addition of buffers and salts. The data also suggested that the noncompetitive pattern of the product inhibition, which is not significantly affected by temperature, increases gently with increasing pH. A Monod form rate expression was proposed to analyze the pH effect on the maximum rate. The proposed kinetic model was also examined by the long-time experiments in which pH, substrate, and product concentration varied obviously during the reaction course.

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Abbreviations

CNH3 :

initial ammonia concentration in the reaction solution, M

Curea :

urea concentration in the reaction solution, M

Curease :

urease concentation in the reaction solution, mg/L

Ea :

activation energy, kJ/mol

[H+]:

hydrogen ion concentration, M

k 0 :

frequency factor, mol NH3/s/g urease

Kes,1, Kes,2 :

empirical parameters in Eq. (4) and Eq. (10),M

K M :

Michaelis constant,M

K p :

product inhibition constant,M

K s :

substrate inhibition constant,M

P :

product concentration,M

S :

substrate concentration,M

T :

temperature, K or °C

t :

time, s

V :

reaction rate, mol NH3/s/g urease

V m :

maximum reaction rate, mol NH3/s/g urease

\(V_{m_0 } \) :

maximum reaction rate without pH inhibition, mol NH3/s/g urease

V m,p :

maximum reaction rate at product concentration P, mol NH3/s/g urease

V m(pH):

maximum reaction rate at pH, mol NH3/s/g urease

α :

empirical exponent in Eq. (4) and Eq. (10)

β :

empirical exponent in Eq. (4) and Eq. (10)

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Authors and Affiliations

  1. Laborat’orio de Engenharia Bioqu’imica, Institute Superior T’ecnico, 1000, Lisboa, Portugal

    Yingjie Qin & Joaquim M. S. Cabral

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  1. Yingjie Qin
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Qin, Y., Cabral, J.M.S. Kinetic studies of the urease-catalyzed hydrolysis of urea in a buffer-free system. Appl Biochem Biotechnol 49, 217–240 (1994). https://doi.org/10.1007/BF02783059

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  • DOI: https://doi.org/10.1007/BF02783059

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