Abstract:
:The template requirements for efficient adenovirus DNA replication were studied in vitro in a reconstituted system with cloned DNA fragments, containing the Ad2 origin region, as templates. Replication is enhanced by nuclear factor I, a cellular protein that binds specifically to the Ad2 origin. This stimulation is shown to be strongly dependent on the concentration of the adenovirus DNA binding protein. Using synthetic oligonucleotides we have constructed plasmids with base substitutions in the nuclear factor I binding region. Footprint analysis and competition filter binding studies show that two of the three small blocks of conserved nucleotides in this region are involved in the binding of nuclear factor I. The binding affinity can be influenced by the base composition of the degenerate region just outside these two blocks. In vitro initiation and DNA chain elongation experiments with the mutants demonstrate that binding of nuclear factor I to the Ad2 origin is necessary for stimulation. However, binding alone is not always sufficient since a mutation which only slightly disturbs binding is strongly impaired in stimulation of DNA replication by nuclear factor I.
journal_name
Nucleic Acids Resjournal_title
Nucleic acids researchauthors
de Vries E,van Driel W,Tromp M,van Boom J,van der Vliet PCdoi
10.1093/nar/13.13.4935subject
Has Abstractpub_date
1985-07-11 00:00:00pages
4935-52issue
13eissn
0305-1048issn
1362-4962journal_volume
13pub_type
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