The Properties and Large-scale Production of L-Asparaginase from Citrobacter

Summary: An intracellular L-asparaginase with antitumour activity was purified from a strain of Citrobacter. The optimum conditions for enzyme production by fermentation on scales up to 2700l were investigated. Highest enzyme yield was obtained in corn-steep liquor medium (9·2%, w/v) at 37°C. Oxygen limitation was not necessary for high enzyme yield. A total recovery of 4·3% from nucleic-acid-free extract and a 180-fold increase in specific activity were obtained after purification. The specific activity of the purified preparation was 45 i.u./mg protein. The enzyme hydrolysed D-asparagine and L-glutamine at 7 and 5%, respectively, of its activity toward L-asparagine, but L-glutaminase activity could be demonstrated only at substrate concentrations above 5 mM. The K m values for L-asparagine and D-asparagine were 2·6 x 10-5 and 1·4 x 10-4 respectively. The anti-lymphoma activity of the enzyme was demonstrated with Gardner lymphosarcoma and was found only slightly less potent that Crasnitin, the most active asparaginase so far tested in this system.