Horigome K, Hayakawa M, Inoue K, Nojima S
J Biochem. 1987 Mar;101(3):625-31. doi: 10.1093/jb/101.3.625.
It was found that phospholipase A2 and lysophospholipase, both of which were released from thrombin-stimulated rat platelets, had high affinity to insolubilized heparin. Phospholipase A2 released from rat platelets was purified by the sequential use of column chromatography on heparin-Sepharose and TSK gel G2000SW (high-performance liquid chromatography, HPLC). The enzyme was near homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and HPLC, and its Mr was estimated to be 13,500. The purified enzyme was labile and lost its activity within 1 h when incubated at 37 degrees C. Phospholipids or detergent in the solution protected the enzyme against inactivation. Phospholipase activity was inhibited by p-bromophenacylbromide, but not by diisopropylfluorophosphate or iodoacetamide. Lysophospholipase, which was also released from rat platelets, was separated from phospholipase A2 by chromatography on heparin-Sepharose.
研究发现，从凝血酶刺激的大鼠血小板中释放的磷脂酶A2和溶血磷脂酶对固定化肝素具有高亲和力。通过依次在肝素-琼脂糖柱和TSK凝胶G2000SW（高效液相色谱，HPLC）上进行柱色谱法，对从大鼠血小板中释放的磷脂酶A2进行了纯化。该酶在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和HPLC上接近均一，其Mr估计为13,500。纯化后的酶不稳定，在37℃孵育时1小时内失去活性。溶液中的磷脂或去污剂可保护该酶不被灭活。磷脂酶活性被对溴苯甲酰溴抑制，但不被二异丙基氟磷酸酯或碘乙酰胺抑制。同样从大鼠血小板中释放的溶血磷脂酶通过在肝素-琼脂糖上的色谱法与磷脂酶A2分离。
