Title

Characterization of a Recombinant Glutaminase-Free l-Asparaginase (ansA3) Enzyme with High Catalytic Activity from Bacillus licheniformis.

Authors

Sudhir, Ankit; Dave, Bhaumik; Prajapati, Anil; Panchal, Ketankumar; Patel, Darshan; Subramanian, R.

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, V, k, and k/ K were calculated for recombinant ansA3.

Subjects

BACILLUS licheniformis; ENZYME analysis; GLUTAMINASES; ASPARAGINASE; ESCHERICHIA coli; POLYACRYLAMIDE gel electrophoresis

Publication

Applied Biochemistry & Biotechnology, 2014, Vol 174, Issue 7, p2504

ISSN

0273-2289

Publication type

Academic Journal

DOI

10.1007/s12010-014-1200-z