Triantafillou D J, Georgatsos J G, Kyriakidis D A
Laboratory of Biochemistry, Faculty of Chemistry, Aristotelian University of Thessaloniki, Greece.
Mol Cell Biochem. 1988 May;81(1):43-51. doi: 10.1007/BF00225652.
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.
天冬酰胺酶活性在梨形四膜虫生长的稳定期达到最大值，并随生长条件和培养基成分而波动。大部分天冬酰胺酶活性（80%）与内质网相关，其余与表膜相关。单独使用洗涤剂或与浓度高达0.5M的氯化钠联合使用均无法使天冬酰胺酶溶解。在用2%（w/v） Triton X100预处理颗粒后，可通过离液剂硫氰酸钾和高氯酸钠或0.1M pH 8.0的磷酸钠缓冲液实现溶解。通过疏水色谱和凝胶过滤色谱，天冬酰胺酶已被纯化至接近均一。天然酶的相对分子量为230,000。它是一种多亚基酶，亚基大小为39,000。其等电点为pH 6.8。它在pH 8.6时作用最佳，Km为2.2mM。它不水解L-谷氨酰胺，其反应受到D-天冬氨酸、D-天冬酰胺以及β位带有取代基的L-天冬酰胺类似物的竞争性抑制。
