Complementation of the temperature-sensitive defect in H5ts125 adenovirus DNA replication in vitro.

Abstract:

:Soluble extracts of adenovirus-infected HeLa cell nuclei support DNA replication on exogenous adenovirus DNA templates. Conditions of synthesis using both wild-type and temperature-sensitive extracts have been defined. Nuclear extracts prepared from cells permissively infected with the adenovirus mutant H5ts125 expressed the temperature-sensitive phenotype and could be inactivated at 37 degrees C in vitro. These extracts were completely complemented by the addition of wild-type adenovirus DNA binding protein but not by H5ts125 DNA binding protein. Enhancement by binding protein in the mutant extracts represents replication, as demonstrated by the production of full-sized products and orderly chain elongation originating, as in vivo, at both ends of the linear DNA. Replicative synthesis required the 5'-terminal protein bound covalently to template DNA and could be inhibited by denaturation of this 55,000-dalton protein. Various inhibitors of eukaryotic DNA polymerases, such as aphidicolin and 2',3'-dideoxythymidine triphosphate, inhibited replication of exogenous adenovirus templates in this system as they do in previously reported systems that only elongate endogenous replicating intermediates.

authors

Kaplan LM,Ariga H,Hurwitz J,Horwitz MS

doi

10.1073/pnas.76.11.5534

subject

Has Abstract

pub_date

1979-11-01 00:00:00

pages

5534-8

issue

11

eissn

0027-8424

issn

1091-6490

journal_volume

76

pub_type

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