Characterization and Partial Purification of L-Asparaginase from Corynebacterium Glutamicum

• Published in: Journal of general microbiology Vol. 136; no. 3; pp. 515 - 519
• Main Authors: Mesas, Juan M, Gil, Jose A, Martin, Juan F
• Format: Journal Article
• Language: English
• Published: London Soc General Microbiol 01.03.1990; New York, NY Cambridge University Press
• Subjects: Asparaginase - isolation & purification; Aspartic Acid - biosynthesis; Aspartic Acid - pharmacology; Bacteriology; Biological and medical sciences; Corynebacterium - drug effects; Corynebacterium - enzymology; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Kinetics; Magnesium - pharmacology; Metabolism. Enzymes; Microbiology; Potassium - pharmacology; Substrate Specificity; Purification; Molecular weight determination; Enzyme; Enzyme inhibitor; Biosynthesis; Characterization; Enzymatic activity; Aminoacid; Corynebacterium glutamicum; Substrate specificity; Bacteria; Actinomycetes; Corynebacteriaceae; Aspartic acid; Asparaginase
• ISSN: 0022-1287
• DOI: 10.1099/00221287-136-3-515

Area de Microbiología, Facultad de Biología, Universidad de León, 24071 León, Spain ABSTRACT SUMMARY: A high L-asparaginase (L-asparagine amidohydrolase; EC 3.5.1.1 ) activity was found under conditions of lysine overproduction in cultures of Corynebacterium glutamicum . L-Asparaginase was purified 98-fold by protamine sulphate precipitation, DEAE-Sephacel anion exchange, ammonium sulphate precipitation and Sephacryl S-200 gel filtration. The asparaginase protein was subjected to PAGE under non-denaturing conditions, identified by an in situ reaction and eluted from the gel in an active form. The estimated M r from gel filtration and SDS-PAGE was 80000. The L-asparaginase activity was inhibited by the L-asparagine analogue 5-diazo-4-oxo-L-norvaline. Neither D-asparagine nor L-glutamine was a substrate for the enzyme. L-Asparaginase was produced constitutively; its role may be that of an overflow enzyme, converting excess asparagine into aspartic acid, the direct precursor of lysine and threonine.