ACTIVATION OF GLYCOSYLASPARAGINASE - FORMATION OF ACTIVE N-TERMINAL THREONINE BY INTRAMOLECULAR AUTOPROTEOLYSIS

The activation mechanism of glycosylasparaginase of Flavobacterium men ingosepticum has been analyzed by site directed mutagenesis and activa tion of purified precursors in vitro. Mutation of Thr-152 to Ser or Cy s leads to gene products that are not activated in vivo but are activa ted in vitro because processing of the mutant precursors is inhibited by certain amino acids in the cell, Kinetic studies reveal that activa tion is an intramolecular autoproteolytic process. The involvement of His150 and Thr/Ser/Cys-152 in activation suggests that autoproteolysis resembles proteolysis by serine/cysteine proteases. Multiple function s of the highly conserved active threonine residue are implicated.