Baranovskiy Andrey G, Babayeva Nigar D, Zhang Yinbo, Gu Jianyou, Suwa Yoshiaki, Pavlov Youri I, Tahirov Tahir H
From the Eppley Institute for Research in Cancer and Allied Diseases, Fred & Pamela Buffett Cancer Center and.
From the Eppley Institute for Research in Cancer and Allied Diseases, Fred & Pamela Buffett Cancer Center and the Departments of Biochemistry and Molecular Biology and.
J Biol Chem. 2016 May 6;291(19):10006-20. doi: 10.1074/jbc.M116.717405. Epub 2016 Mar 14.
The human primosome, a 340-kilodalton complex of primase and DNA polymerase α (Polα), synthesizes chimeric RNA-DNA primers to be extended by replicative DNA polymerases δ and ϵ. The intricate mechanism of concerted primer synthesis by two catalytic centers was an enigma for over three decades. Here we report the crystal structures of two key complexes, the human primosome and the C-terminal domain of the primase large subunit (p58C) with bound DNA/RNA duplex. These structures, along with analysis of primase/polymerase activities, provide a plausible mechanism for all transactions of the primosome including initiation, elongation, accurate counting of RNA primer length, primer transfer to Polα, and concerted autoregulation of alternate activation/inhibition of the catalytic centers. Our findings reveal a central role of p58C in the coordinated actions of two catalytic domains in the primosome and ultimately could impact the design of anticancer drugs.
人引发体是一种由引发酶和DNA聚合酶α（Polα）组成的340千道尔顿复合物，它合成嵌合的RNA-DNA引物，以供复制性DNA聚合酶δ和ε进行延伸。三十多年来，由两个催化中心协同进行引物合成的复杂机制一直是个谜。在此，我们报告了两种关键复合物的晶体结构，即人引发体以及与结合的DNA/RNA双链体结合的引发酶大亚基的C末端结构域（p58C）。这些结构，连同对引发酶/聚合酶活性的分析，为引发体的所有作用提供了一种合理的机制，包括起始、延伸、准确计算RNA引物长度、引物转移至Polα以及对催化中心交替激活/抑制的协同自动调节。我们的研究结果揭示了p58C在引发体中两个催化结构域的协同作用中的核心作用，最终可能会影响抗癌药物的设计。
