Ishimaru K, Kihara H, Ohno M
J Biochem. 1980 Aug;88(2):443-51. doi: 10.1093/oxfordjournals.jbchem.a132991.
Phospholipase A was purified from venom of Trimeresurus flavoviridis (Habu snake) via three steps consisting of Sephadez G-100, CM-cellulose, and DEAE-cellulose column chromatographies. The apparent molecular weights determined by gel filtration on Sephadex G-75 and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were 28,000 and 14,000, respectively, suggesting that the enzyme is composed of two identical subunits. The isoelectric point was 7.9. The enzyme is characterized by high contents of aspartic acid, glycine, tyrosine, and lysine and contains one residue each of histidine and methionine, three or four tryptophans, and eight disulfide bonds per subunit. The enzyme was inactivated by reaction with p-bromophenacyl bromide following pseudo first order kinetics. The loss of activity was accompanied by loss of histidine, indicating that a single histidine residue is essential for activity. Oxidation with N-bromosuccinimide decreased the enzymic activity. Tryptophan residues appear to play some role in catalysis.
磷脂酶A是从竹叶青蛇（哈布蛇）毒液中通过三步纯化得到的，这三步包括葡聚糖凝胶G - 100、CM - 纤维素和DEAE - 纤维素柱层析。通过葡聚糖凝胶G - 75凝胶过滤法和十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳法测定的表观分子量分别为28,000和14,000，这表明该酶由两个相同的亚基组成。其等电点为7.9。该酶的特点是天冬氨酸、甘氨酸、酪氨酸和赖氨酸含量较高，每个亚基含有一个组氨酸和甲硫氨酸残基、三或四个色氨酸以及八个二硫键。该酶与对溴苯甲酰溴反应后按照准一级动力学失活。活性丧失伴随着组氨酸的丢失，表明单个组氨酸残基对活性至关重要。用N - 溴代琥珀酰亚胺氧化会降低酶活性。色氨酸残基似乎在催化中起一定作用。
