title: Characterization of a recombinant glutaminase-free L-asparaginase (ansA3) enzyme with high catalytic activity from Bacillus licheniformis.
abstract: L-Asparaginase (3.5.1.1) is an enzyme widely used to treat the acute lymphoblastic leukemia. Two genes coding for L-asparaginase (ansA1 and ansA3) from Bacillus licheniformis MTCC 429 were cloned and overexpressed in Escherichia coli BL21 (DE3) cells. The recombinant proteins were purified to homogeneity by one-step purification process and further characterized for various biochemical parameters. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that both the enzymes are monomers of ∼37 kDa. Recombinant ansA1 was found to be highly unstable, and recombinant ansA3 was catalytically active and stable, which showed an optimum activity of 407.65 IU/mg at 37 °C and pH 8. Recombinant ansA3 showed higher substrate specificity for L-asparagine with negligible glutaminase activity. Kinetic parameters like K m , V max, k cat, and k cat/K m were calculated for recombinant ansA3.
authors.affiliation: BRD School of Biosciences, Sardar Patel Maidan, Sardar Patel University, Vadtal Road, Satellite Campus, PO Box 39, Vallabh Vidyanagar, 388 120, Gujarat, India.
authors.forename: Ankit P
authors.initials: AP
authors.lastname: Sudhir
authors.forename: Bhaumik R
authors.initials: BR
authors.lastname: Dave
authors.forename: Anil S
authors.initials: AS
authors.lastname: Prajapati
authors.forename: Ketankumar
authors.initials: K
authors.lastname: Panchal
authors.forename: Darshan
authors.initials: D
authors.lastname: Patel
authors.forename: R B
authors.initials: RB
authors.lastname: Subramanian
mesh_terms: D001215:Asparaginase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D001216:Asparagine; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics; D002384:Catalysis; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D013379:Substrate Specificity
chemical_list: D011994:Recombinant Proteins; D001216:Asparagine; D001215:Asparaginase