Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
Proc Natl Acad Sci U S A. 2011 Jun 7;108(23):9372-7. doi: 10.1073/pnas.1106678108. Epub 2011 May 23.
Interactions between gene 4 helicase and gene 5 DNA polymerase (gp5) are crucial for leading-strand DNA synthesis mediated by the replisome of bacteriophage T7. Interactions between the two proteins that assure high processivity are known but the interactions essential to initiate the leading-strand DNA synthesis remain unidentified. Replacement of solution-exposed basic residues (K587, K589, R590, and R591) located on the front surface of gp5 with neutral asparagines abolishes the ability of gp5 and the helicase to mediate strand-displacement synthesis. This front basic patch in gp5 contributes to physical interactions with the acidic C-terminal tail of the helicase. Nonetheless, the altered polymerase is able to replace gp5 and continue ongoing strand-displacement synthesis. The results suggest that the interaction between the C-terminal tail of the helicase and the basic patch of gp5 is critical for initiation of strand-displacement synthesis. Multiple interactions of T7 DNA polymerase and helicase coordinate replisome movement.
噬菌体 T7 复制体介导的领头链 DNA 合成，需要基因 4 解旋酶和基因 5 DNA 聚合酶（gp5）之间的相互作用。已知这两种蛋白之间存在确保高持续性的相互作用，但启动领头链 DNA 合成所必需的相互作用仍未确定。用中性天冬酰胺取代位于 gp5 前表面的暴露在溶液中的碱性残基（K587、K589、R590 和 R591），会使 gp5 和解旋酶丧失介导链置换合成的能力。gp5 中的这个前碱性斑与解旋酶的酸性 C 末端尾巴有物理相互作用。尽管如此，改变后的聚合酶仍能够取代 gp5 并继续进行正在进行的链置换合成。结果表明，解旋酶 C 末端尾巴与 gp5 的碱性斑之间的相互作用对链置换合成的起始至关重要。T7 DNA 聚合酶和解旋酶的多种相互作用协调复制体的运动。
