[Isolation and properties of a homogeneous L-asparaginase preparation from Pseudomonas fluorescens AG].
Highly purified L-asparaginase having a specific activity of 500+/- +/-40 IU./mg protein is isolated from Pseudomonas fluorescens AG cells. The purification procedure includes isopropanol fractionation, gel filtration through Sephadex G-100, chromatography on hydroxylapatite and DEAE-cellulose columns. The asparaginase preparation is homogenous on the basis of polyacrylamide gel electrophoresis data. The pH optimum is found to be 8.0-9.0, isoelectric point and molecular weight are 4.5+/-0.05 and 70,000+/-5,000 respectively, Km for L-asparagine being-4.1-10(-4)M. The enzyme does not hydrolyse L-glutamine. The hydrolysis rate of D-glutamine is less than 1% of the deamydation rate of L-isomer. p-Chloro-mercurium benzoate at a concentration of 10(-4) M completely inhibits the asparaginase activity. Asparaginase from Ps. fluorescens AG possesses and antileucosic activity, inhibiting 3H-thymidine incorporation into DNA of Berkit lymphoma cells.
[Isolation and properties of a homogeneous L-asparaginase preparation from Pseudomonas fluorescens AG]. Highly purified L-asparaginase having a specific activity of 500+/- +/-40 IU./mg protein is isolated from Pseudomonas fluorescens AG cells. The purification procedure includes isopropanol fractionation, gel filtration through Sephadex G-100, chromatography on hydroxylapatite and DEAE-cellulose columns. The asparaginase preparation is homogenous on the basis of polyacrylamide gel electrophoresis data. The pH optimum is found to be 8.0-9.0, isoelectric point and molecular weight are 4.5+/-0.05 and 70,000+/-5,000 respectively, Km for L-asparagine being-4.1-10(-4)M. The enzyme does not hydrolyse L-glutamine. The hydrolysis rate of D-glutamine is less than 1% of the deamydation rate of L-isomer. p-Chloro-mercurium benzoate at a concentration of 10(-4) M completely inhibits the asparaginase activity. Asparaginase from Ps. fluorescens AG possesses and antileucosic activity, inhibiting 3H-thymidine incorporation into DNA of Berkit lymphoma cells.