Production, Isolation, and Purification of L-Asparaginase from Pseudomonas Aeruginosa 50071 Using Solid-state Fermentation

  • El-Bessoumy, Ashraf A. ;
  • Sarhan, Mohamed ;
  • Mansour, Jehan
  • Published : 2004.07.31


The L-asparaginase (E. C. 3. 5. 1. 1) enzyme was purified to homogeneity from Pseudomonas aeruginosa 50071 cells that were grown on solid-state fermentation. Different purification steps (including ammonium sulfate fractionation followed by separation on Sephadex G-100 gel filtration and CM-Sephadex C50) were applied to the crude culture filtrate to obtain a pure enzyme preparation. The enzyme was purified 106-fold and showed a final specific activity of 1900 IU/mg with a 43% yield. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the purified enzyme revealed it was one peptide chain with $M_r$ of 160 kDa. A Lineweaver-Burk analysis showed a $K_m$ value of 0.147 mM and $V_{max}$ of 35.7 IU. The enzyme showed maximum activity at pH 9 when incubated at $37^{\circ}C$ for 30 min. The amino acid composition of the purified enzyme was also determined.


Amino acid composition;L-asparaginase;Production;Pseudomonas aeruginosa 50071;Purification


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