Open Access Highly Accessed Research article

Extraction, purification, kinetic and thermodynamic properties of urease from germinating Pisum Sativum L. seeds

Mohamed E EL-Hefnawy13*, Mohamed Sakran23, Ali I Ismail1 and Eman Fahmy Aboelfetoh3

Author Affiliations

1 Department of Chemistry, Rabigh College of Science and Arts, King Abdulaziz University, P.O. Box 344, Rabigh 21911, Saudi Arabia

2 Department of Biochemistry, Faculty of Science, Tabuk University, Tabuk, Saudi Arabia

3 Department of Chemistry, Faculty of Science, Tanta University, Tanta 31527, Egypt

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BMC Biochemistry 2014, 15:15  doi:10.1186/1471-2091-15-15

Published: 28 July 2014

Abstract

Background

Urease, one of the highly efficient known enzymes, catalyzes the hydrolysis of urea into ammonia and carbon dioxide. The present study aimed to extract urease from pea seeds (Pisum Sativum L). The enzyme was then purified in three consequence steps: acetone precipitation, DEAE-cellulose ion-exchange chromatography, and gel filtration chromatography (Sephacryl S-200 column).

Results

The purification fold was 12.85 with a yield of 40%. The molecular weight of the isolated urease was estimated by chromatography to be 269,000 Daltons. Maximum urease activity (190 U/g) was achieved at the optimum conditions of 40°C and pH of 7.5 after 5 min of incubation. The kinetic parameters, Km and Vmax, were estimated by Lineweaver-Burk fits and found to be 500 mM and 333.3 U/g, respectively. The thermodynamic constants of activation, ΔH, Ea, and ΔS, were determined using Arrhenius plot and found to be 21.20 kJ/mol, 23.7 kJ/mol, and 1.18 kJ/mol/K, respectively.

Conclusions

Urease was purified from germinating Pisum Sativum L. seeds. The purification fold, yield, and molecular weight were determined. The effects of pH, concentration of enzyme, temperature, concentration of substrate, and storage period on urease activity were examined. This may provide an insight on the various aspects of the property of the enzyme. The significance of extracting urease from different sources could play a good role in understanding the metabolism of urea in plants.

Keywords:
Urease; Enzyme activity; Enzyme purification; Pisum Sativum L; Pea seeds