Mechanistic Studies of Replicative Polymerases

描述(申请人提供)：细胞复制酶的催化亚基与简单的聚合酶的区别在于复杂的模块化结构，允许与广泛的DNA复制蛋白相互作用。以细菌DNA聚合酶III全酶的α亚基为原型，我们将研究这些模块的功能以及它们之间的通信。我们将研究与β处理因子和DNAX复合体的tau“组织者”亚单位结合的位点，以及在Okazaki片段合成过程中实现循环的亲和力方面的相互作用。将确定α内epsilon校对亚基的结合部位，并进一步探索用于整合聚合酶和核酸外切酶活性部位的机制。我们还将确定复制聚合酶中发现的其他保守结构域和基序的功能。

DESCRIPTION (provided by applicant): The catalytic subunits of cellular replicases are distinguished from simpler polymerases by a complex modular structure that permits interaction with a wide array of DNA replication proteins. Using the alpha subunit of bacterial DNA polymerase III holoenzyme as a prototype, we will study the function of these modules and communication between them. We will investigate the sites involved in binding the beta processivity factor and the tau 'organizer' subunit of the DnaX complex, as well as the interplay in affinities that enables recycling during Okazaki fragment synthesis. The binding sites for the epsilon proofreading subunit within alpha will be determined, and the mechanism used to integrate the polymerase and exonuclease active sites will be further explored. We will also determine the function of other conserved domains and motifs found in replicative polymerases.