Purification and properties of L-asparaginase from Mycobacterium phlei.
1. L-asparaginase from M. phlei was purified about 170-fold with an 11% yield. The purification procedure consisted of: fractionation with ammonium sulphate; adsorption of contaminating proteins on calcium phosphate gel; chromatography on Sephadex G-150 and DEAE-cellulose. The specific activity of the final preparation was 32.6 i.u./mg protein. 2. Molecular weight of the enzyme as determined by Sephadex G-100 filtration amounted to 126 000. Optimum pH was 8.8-9.2. The enzyme did not hydrolyse L-glutamine over the pH range 4-9, and was inhibited by D-asparagine. The apparent Michaelis constant for L-asparagine was 0.7 mM; energy of activation, 9800 cal/mole. 3. On polyacrylamide-gel electrophoresis the final preparation revealed two protein bands, one of which was coincident with the enzyme activity.
Purification and properties of L-asparaginase from Mycobacterium phlei. 1. L-asparaginase from M. phlei was purified about 170-fold with an 11% yield. The purification procedure consisted of: fractionation with ammonium sulphate; adsorption of contaminating proteins on calcium phosphate gel; chromatography on Sephadex G-150 and DEAE-cellulose. The specific activity of the final preparation was 32.6 i.u./mg protein. 2. Molecular weight of the enzyme as determined by Sephadex G-100 filtration amounted to 126 000. Optimum pH was 8.8-9.2. The enzyme did not hydrolyse L-glutamine over the pH range 4-9, and was inhibited by D-asparagine. The apparent Michaelis constant for L-asparagine was 0.7 mM; energy of activation, 9800 cal/mole. 3. On polyacrylamide-gel electrophoresis the final preparation revealed two protein bands, one of which was coincident with the enzyme activity.