Title

A Dna Helicase Associated With Dna Polymerase Delta

Date of Award

1990

Availability

Article

Degree Name

Doctor of Philosophy (Ph.D.)

Department

Biochemistry and Molecular Biology

First Committee Member

Antero G. So, Committee Chair

Abstract

DNA polymerases alpha and delta are proposed to be the lagging strand and leading strand replicases in eukaryotes, based on the functional properties of these distinct DNA polymerases. One of the properties cited to support this hypothesis is the ability of partially purified DNA polymerase delta, but not DNA polymerase alpha to catalyze strand displacement synthesis. Characterization of strand displacement synthesis by DNA polymerase delta indicated that the proliferating cell nuclear antigen (PCNA) is absolutely required for strand displacement synthesis, and that strand displacement synthesis catalyzed by pol $\delta$/PCNA is very efficient, since the measured rate of replication fork movement approaches the in vivo rate and this rapid leading strand DNA synthesis is highly processive.Further dissection and reconstitution of this strand displacement synthesis system resulted in the separation of a strand displacement factor(s) (SDF) from pol delta core enzyme. SDF specifically endows pol $\delta$ core enzyme, but not pol $\alpha$, with the ability to catalyze strand displacement synthesis in the presence of PCNA, suggesting a specific protein-protein interaction. The stimulatory effect of SDF on strand displacement synthesis by pol $\delta$ core enzyme/PCNA is not due to stimulation of the DNA polymerase activity of pol $\delta$ core enzyme. SDF is associated with a DNA-dependent ATPase activity and a DNA helicase activity. We have partially purified and characterized the DNA helicase activity associated with pol $\delta$. The optimal conditions for $\delta$ helicase activity are identical to those for strand displacement synthesis. More importantly, delta helicase shares several properties with the prokaryotic DNA helicases known to be involved in DNA replication, i.e., it translocates in the 5$\sp\prime$ to 3$\sp\prime$ direction and requires a fork-like substrate for optimal activity. These findings, together with the observation that this DNA helicase is tightly associated with the leading strand replicase, suggest that the pol delta-associated DNA helicase is involved in chromosomal replication in eukaryotes.

Keywords

Biology, Molecular

Link to Full Text

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