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Purification, characterization, cytotoxicity and anticancer activities of L-asparaginase, anti-colon cancer protein, from the newly isolated alkaliphilic Streptomyces fradiae NEAE-82

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ABSTRACT

L-asparaginase is an important enzyme as therapeutic agents used in combination with other drugs in the treatment of acute lymphoblastic leukemia. A newly isolated actinomycetes strain, Streptomyces sp. NEAE-82, was potentially producing extracellular L-asparaginase, it was identified as Streptomyces fradiae NEAE-82, sequencing product was deposited in the GenBank database under accession number KJ467538. L-asparaginase was purified from the crude enzyme using ammonium sulfate precipitation, dialysis and ion exchange chromatography using DEAE Sepharose CL-6B. Further the kinetic studies of purified enzyme were carried out. The optimum pH, temperature and incubation time for maximum L-asparaginase activity were found to be 8.5, 40 °C and 30 min, respectively. The optimum substrate concentration was found to be 0.06 M. The Km and Vmax of the enzyme were 0.01007 M and 95.08 Uml−1min−1, respectively. The half-life time (T1/2) was 184.91 min at 50 °С, while being 179.53 min at 60 °С. The molecular weight of the subunits of L-asparaginase was found to be approximately 53 kDa by SDS–PAGE analysis. The purified L-asparaginase showed a final specific activity of 30.636 U/mg protein and was purified 3.338-fold. The present work for the first time reported more information in the production, purification and characterization of L-asparaginase produced by newly isolated actinomycetes Streptomyces fradiae NEAE-82.

No MeSH data available.


Effect of the temperature on L-asparaginase activity.
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f7: Effect of the temperature on L-asparaginase activity.

Mentions: The temperature optimum of L-asparaginase from Streptomyces fradiae NEAE-82 is shown in Fig. 7. It was active at wide range of temperature condition from 25–60 °C. The maximum L-asparaginase activity of 26.848 IU was obtained at 40 °C. At higher temperature the L-asparaginase activity declined. The enzyme retains 50.85% of its activity at 60 °C. Our results were in agreement with a previous study which reported that the maximum activity of L-asparaginase purified from Streptomyces gulbargensis was at 40 °C11. Manna et al.28 have found 37 °C to be the optimum temperature for the enzyme activity obtained from Pseudomonas stutzeri MB-405. However, optimum temperature for L-asparaginase activity obtained from Erwinia sp. showed maximum activity at 35 °C29.


Purification, characterization, cytotoxicity and anticancer activities of L-asparaginase, anti-colon cancer protein, from the newly isolated alkaliphilic Streptomyces fradiae NEAE-82
Effect of the temperature on L-asparaginase activity.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5015098&req=5

f7: Effect of the temperature on L-asparaginase activity.
Mentions: The temperature optimum of L-asparaginase from Streptomyces fradiae NEAE-82 is shown in Fig. 7. It was active at wide range of temperature condition from 25–60 °C. The maximum L-asparaginase activity of 26.848 IU was obtained at 40 °C. At higher temperature the L-asparaginase activity declined. The enzyme retains 50.85% of its activity at 60 °C. Our results were in agreement with a previous study which reported that the maximum activity of L-asparaginase purified from Streptomyces gulbargensis was at 40 °C11. Manna et al.28 have found 37 °C to be the optimum temperature for the enzyme activity obtained from Pseudomonas stutzeri MB-405. However, optimum temperature for L-asparaginase activity obtained from Erwinia sp. showed maximum activity at 35 °C29.

View Article: PubMed Central - PubMed

ABSTRACT

L-asparaginase is an important enzyme as therapeutic agents used in combination with other drugs in the treatment of acute lymphoblastic leukemia. A newly isolated actinomycetes strain, Streptomyces sp. NEAE-82, was potentially producing extracellular L-asparaginase, it was identified as Streptomyces fradiae NEAE-82, sequencing product was deposited in the GenBank database under accession number KJ467538. L-asparaginase was purified from the crude enzyme using ammonium sulfate precipitation, dialysis and ion exchange chromatography using DEAE Sepharose CL-6B. Further the kinetic studies of purified enzyme were carried out. The optimum pH, temperature and incubation time for maximum L-asparaginase activity were found to be 8.5, 40 °C and 30 min, respectively. The optimum substrate concentration was found to be 0.06 M. The Km and Vmax of the enzyme were 0.01007 M and 95.08 Uml−1min−1, respectively. The half-life time (T1/2) was 184.91 min at 50 °С, while being 179.53 min at 60 °С. The molecular weight of the subunits of L-asparaginase was found to be approximately 53 kDa by SDS–PAGE analysis. The purified L-asparaginase showed a final specific activity of 30.636 U/mg protein and was purified 3.338-fold. The present work for the first time reported more information in the production, purification and characterization of L-asparaginase produced by newly isolated actinomycetes Streptomyces fradiae NEAE-82.

No MeSH data available.