CHARACTERIZATION AND PARTIAL-PURIFICATION OF L-ASPARAGINASE FROM CORYNEBACTERIUM-GLUTAMICUM

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.