Turman M A, Douvas A
Department of Biochemistry, University of Colorado Health Sciences Center, Denver 80262.
Biochem Med Metab Biol. 1993 Oct;50(2):210-25. doi: 10.1006/bmmb.1993.1063.
Topoisomerase I (Topo I) is involved in many cellular functions that involve unwinding of supercoiled DNA, such as transcription and replication. Topo I is also the target of autoimmune antibodies in progressive systemic sclerosis (scleroderma), and abnormal regulation of Topo I may influence the excessive production of collagen found in scleroderma. Topo I is phosphorylated in vivo at serine residues and, in vitro, the activity of Topo I is increased by phosphorylation by casein kinase type II (CKII) and protein kinase C (PKC). In this study, a protein kinase activity from rat liver nuclei is shown to copurify with Topo I during Bio-Rex 70 cation exchange chromatography. The kinase can phosphorylate Topo I at serine residues, resulting in a threefold increase in topoisomerase activity. A relatively tight association between this kinase and Topo I is demonstrated by the ability to coprecipitate the kinase with scleroderma autoimmune anti-Topo I antibodies. The kinase activity is similar to CKII since it is Ca2+ and cyclic nucleotide independent, it can utilize either ATP or GTP as phosphate donor, and it can phosphorylate casein and phosvitin, but not histones. However, unlike typical CKII, the Topo I-associated kinase could utilize Mn2+ almost as well as Mg2+, it is not stimulated by polyamines, and it does not appear to undergo autophosphorylation. In conclusion, we demonstrate that rat liver Topo I is relatively tightly associated with a CKII-like protein kinase that can phosphorylate and activate Topo I. These findings provide corroborative evidence that CKII, or a CKII-like protein kinase, is a physiologic regulator of Topo I.
拓扑异构酶I（Topo I）参与许多涉及解开超螺旋DNA的细胞功能，如转录和复制。Topo I也是进行性系统性硬化症（硬皮病）中自身免疫抗体的靶标，Topo I的异常调节可能影响硬皮病中发现的胶原蛋白过度产生。Topo I在体内丝氨酸残基处被磷酸化，在体外，酪蛋白激酶II型（CKII）和蛋白激酶C（PKC）的磷酸化可增加Topo I的活性。在本研究中，大鼠肝细胞核中的一种蛋白激酶活性在Bio-Rex 70阳离子交换色谱过程中显示与Topo I共纯化。该激酶可在丝氨酸残基处磷酸化Topo I，导致拓扑异构酶活性增加三倍。通过用硬皮病自身免疫抗Topo I抗体共沉淀该激酶的能力，证明了该激酶与Topo I之间存在相对紧密的关联。该激酶活性与CKII相似，因为它不依赖Ca2+和环核苷酸，它可以利用ATP或GTP作为磷酸供体，并且它可以磷酸化酪蛋白和磷蛋白，但不能磷酸化组蛋白。然而，与典型的CKII不同，与Topo I相关的激酶几乎可以像Mg2+一样有效地利用Mn2+，它不受多胺刺激，并且似乎不会发生自身磷酸化。总之，我们证明大鼠肝脏Topo I与一种可磷酸化并激活Topo I的CKII样蛋白激酶相对紧密地相关。这些发现提供了确证性证据，表明CKII或一种CKII样蛋白激酶是Topo I的生理调节剂。
