Purification and properties of a membrane-bound L-asparaginase ofTetrahymena pyriformis

✍ Scribed by D. J. Triantafillou; J. G. Georgatsos; D. A. Kyriakidis

• Book ID: 104671732
• Publisher: Springer
• Year: 1988
• Tongue: English
• Weight: 569 KB
• Volume: 81
• Category: Article
• ISSN: 0300-8177
• DOI: 10.1007/bf00225652

✦ Synopsis

L-Asparaginase activity reaches maximal values at the stationary phase of growth of Tetrahymena pyriformis and fluctuates upon the growth conditions and the composition of the medium. Most of the L-asparaginase activity (80%) is associated with the endoplasmic reticulum, and the remaining with the pellicles. Detergents either alone or in combination with NaCl up to 0.5 M concentration failed to solubilize L-asparaginase. Solubilization can be accomplished by means of either the chaotropic agents KSCN and NaClO4, or 0.1 M sodium phosphate buffer pH 8.0, following pretreatment of the particulates with 2% w/v Triton X100. L-Asparaginase has been purified to near homogeneity by hydrophobic and gel filtration chromatography. The native enzyme has a relative molecular weight of 230,000. It is a multiple subunit enzyme, with subunit size of 39,000. Its isoelectric point is at pH 6.8. It acts optimally at pH 8.6 with a Km of 2.2 mM. It does not hydrolyse L-glutamine and its reaction is inhibited competitively by D-aspartic acid and D-asparagine as well as by L-asparagine analogues with substituents at the beta position.