Early dissociation of nuclear factor I from the origin during initiation of adenovirus DNA replication studied by origin immobilization.
AUTOR(ES)
Coenjaerts, F E
RESUMO
The DNA-binding domain of Nuclear Factor I (NFIBD) enhances initiation of adenovirus DNA replication up to 50-fold by binding to the auxiliary region of the origin and positioning the viral DNA polymerase. To study if and when NFIBD dissociates from the template, we immobilized origin DNA to glutathione-agarose beads by means of a GST-NFIBD fusion protein. This immobilized template is active in replication. By analyzing the release of prelabeled templates from the beads under different conditions, we show that NFIBD dissociates already early during initiation. During preinitiation NFIBD remains bound, but as soon as dCTP, dATP or dTTP are added, efficient dissociation occurs. A much lower dissociation level was induced by addition of dGTP. Since dCTP, dATP and dTTP are required for formation of a pTP-CAT initiation intermediate, we explain our results by conformational changes occurring in the polymerase during initiation leading to disruption of both the interaction between the polymerase and NFI as well as the interaction between NFI and the DNA.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=332066Documentos Relacionados
- Targeting of DNA polymerase to the adenovirus origin of DNA replication by interaction with nuclear factor I.
- Bending of Adenovirus Origin DNA by Nuclear Factor I as Shown by Scanning Force Microscopy Is Required for Optimal DNA Replication
- Adenovirus DNA replication in vitro: site-directed mutagenesis of the nuclear factor I binding site of the Ad2 origin.
- Nuclear factor I enhances adenovirus DNA replication by increasing the stability of a preinitiation complex.
- Interaction between the octamer-binding protein nuclear factor III and the adenovirus origin of DNA replication.