Mechanistic Studies of Replicative Polymerases

University Of Colorado, Boulder CO

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Abstract

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.