|
KMID : 0545120220320050551
|
|
Journal of Microbiology and Biotechnology
2022 Volume.32 No. 5 p.551 ~ p.563
|
|
|
Anticancer Activity of Extremely Effective Recombinant L-Asparaginase from Burkholderia pseudomallei
|
|
Darwesh Doaa B.
Al-Awthan Yahya S.
Elfaki Imadeldin
Habib Salem A.
Alnour Tarig M.
Darwish Ahmed B.
Youssef Magdy M.
|
|
|
Abstract
|
|
|
|
L-asparaginase (E.C. 3.5.1.1) purified from bacterial cells is widely used in the food industry, as well as in the treatment of childhood acute lymphoblastic leukemia. In the present study, the Burkholderia pseudomallei L-asparaginase gene was cloned into the pGEX-2T DNA plasmid, expressed in E. coli BL21 (DE3) pLysS, and purified to homogeneity using Glutathione Sepharose chromatography with 7.26 purification fold and 16.01% recovery. The purified enzyme exhibited a molecular weight of ~33.6 kDa with SDS-PAGE and showed maximal activity at 50°C and pH 8.0. It retained 95.1, 89.6%, and 70.2% initial activity after 60 min at 30°C, 40°C, and 50°C, respectively. The enzyme reserved its activity at 30°C and 37°C up to 24 h. The enzyme had optimum pH of 8 and reserved 50% activity up to 24 h. The recombinant enzyme showed the highest substrate specificity towards L-asparaginase substrate, while no detectable specificity was observed for L-glutamine, urea, and acrylamide at 10 mM concentration. THP-1, a human leukemia cell line, displayed significant morphological alterations after being treated with recombinant L-asparaginase and the IC50 of the purified enzyme was recorded as 0.8 IU. Furthermore, the purified recombinant L-asparaginase improved cytotoxicity in liver cancer HepG2 and breast cancer MCF-7 cell lines, with IC50 values of 1.53 and 18 IU, respectively.
|
|
|
KEYWORD
|
|
|
L-asparaginase, leukemia, cloning, DNA, purification, characterization
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
  
|
|