Guissani A
Biochimie. 1978;60(8):755-65. doi: 10.1016/s0300-9084(78)80020-0.
This report describes structural studies on purified polynucleotide phosphorylase from C. perfringens. A method is described for the purification of the enzyme which yields a product equivalent in activity to the native polynucleotide phosphorylase from E. coli. These studies revealed a molecular heterogeneity arising from successive stages of proteolysis, to which this enzyme is especially sensitive; unusally, the enzyme is obtained as a mixture of variable proportions of the native and proteolysed forms. We found in all cases a trimeric basic structure composed of the native (alpha) or proteolysed (lapha) or proteolysed (alpha', alpha") catalytic sub-units, However, the enzyme is rather easily dissociated into its sub-units, a phenomenon which seems to accompany proteolysis (Table). Under the action of either endogenous proteases or trypsin, two enzymatic forms are obtained: their quaternary structures seem analogous, but they differ in their catalytic properties from each other and from the initial enzyme. With some care at each step of purification, the polynucleotide phosphorylase of E. coli can be obtained exclusively in its native form. The greater susceptibility to proteolysis of the enzyme from C. perfrigens and the relationship between such degradation and quaternary structure seem to be at the origin of the peculiar behavior of this polynucleotide phosphorylase.
本报告描述了对产气荚膜梭菌纯化的多核苷酸磷酸化酶的结构研究。文中介绍了一种纯化该酶的方法，所得产物的活性与大肠杆菌天然多核苷酸磷酸化酶相当。这些研究揭示了由于蛋白水解的连续阶段而产生的分子异质性，该酶对此特别敏感；不同寻常的是，该酶是以天然形式和蛋白水解形式的可变比例混合物形式获得的。我们在所有情况下都发现了一种三聚体基本结构，由天然（α）或蛋白水解（α'、α''）催化亚基组成。然而，该酶相当容易解离成其亚基，这种现象似乎伴随着蛋白水解（表）。在内源蛋白酶或胰蛋白酶的作用下，可得到两种酶形式：它们的四级结构似乎相似，但它们的催化特性彼此不同，也与初始酶不同。在纯化的每一步都小心操作的情况下，大肠杆菌的多核苷酸磷酸化酶可以完全以其天然形式获得。产气荚膜梭菌的酶对蛋白水解的更高敏感性以及这种降解与四级结构之间的关系似乎是这种多核苷酸磷酸化酶特殊行为的根源。
