Date of Award

2010

Degree Type

Thesis

Degree Name

Master of Science

Program

Biochemistry

Supervisor

Dr. Megan J. Davey

Second Advisor

Dr. Christopher J. Brandi

Third Advisor

Dr. David B. Haniford

Abstract

Genome duplication occurs once and only once during each cell cycle. It is a highly ordered process and is separated into the formation of different multi-protein complexes. The pre-replicative complex (preRC) is formed during G1 -phase and is composed of ORC, Cdc6, Cdtl and Mcm2-7. Mcm2-7 is the replicative helicase in eukaryotic cells and is assembled on replication origins prior to S-phase. Cdtl is an essential component of the preRC. Cdtl has been shown to interact with Mcm2-7, however neither the requirements nor the effects of this interaction have explored. In this study, I show that Cdtl forms a complex with Mcm2-7 without the need for other factors. Furthermore, Cdtl modulates the helicase, ATPase and DNA binding activity of the Mcm2-7 complex. I propose a model where Cdtl modulates Mcm2-7 helicase activity by inhibiting ATP hydrolysis by Mcm2-7, thus preventing premature DNA unwinding. Furthermore, the increase in dsDNA binding affinity for the Mcm2-7/Cdtl complex (Mcm2-7*Cdtl) assists in its loading onto replication origins during preRC assembly. These results indicate a novel role for Cdtl in Mcm2-7 modulation and DNA replication control.

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