Panzeter P L, Althaus F R
Institute of Pharmacology and Toxicology, University of Zürich-Tierspital, Switzerland.
Biochemistry. 1994 Aug 16;33(32):9600-5. doi: 10.1021/bi00198a028.
The nuclear enzyme poly(ADP-ribose) polymerase participates in DNA excision repair. Following binding to DNA strand breaks through its amino-terminal Zn(2+)-finger domain, the enzyme is activated to form polymerase-associated ADP-ribose polymers of various sizes. Focusing on this "automodification" reaction, we observed that optimal enzyme activity and maximal polymer formation were attained only at a strict stoichiometry of two polymerase molecules per DNA fragment. Using various linearized DNAs and nicked circular DNA, we show that this stoichiometric dependence is dictated by the number of enzyme activating sites, i.e., DNA strand breaks. Deviations from the optimal ratio inevitably resulted in decreased polymer formation, ruling out a strict automodification mechanism of poly(ADP-ribosyl)ation. Our results suggest that the mechanism of poly(ADP-ribose) formation on polymerase molecules entails DNA strand break-mediated partitioning of the polymerase into two functional populations: one bound to the DNA breaks and catalytically active, the other, catalytically inactive, functioning as polymer acceptors.
核酶聚（ADP - 核糖）聚合酶参与DNA切除修复。该酶通过其氨基末端的锌（2+）指结构域与DNA链断裂结合后被激活，形成各种大小的与聚合酶相关的ADP - 核糖聚合物。针对这种“自身修饰”反应，我们观察到只有在每个DNA片段严格的两个聚合酶分子化学计量比下才能达到最佳酶活性和最大聚合物形成。使用各种线性化DNA和带切口的环状DNA，我们表明这种化学计量依赖性由酶激活位点的数量即DNA链断裂决定。偏离最佳比例不可避免地导致聚合物形成减少，排除了聚（ADP - 核糖基）化严格的自身修饰机制。我们的结果表明，聚合酶分子上聚（ADP - 核糖）形成的机制需要DNA链断裂介导的聚合酶分成两个功能群体：一个与DNA断裂结合并具有催化活性，另一个无催化活性，作为聚合物受体起作用。
