Isolation of Proteins Involved in the Replication of Adenoviral DNA In vitro

Abstract

The simple mechanism of replication of adenoviral DNA has made adenovirus an especially useful model system for studies of eukaryotic replication mechanisms. The adenoviral genome is a 35,000-bp linear, double-stranded DNA molecule containing inverted terminal repeats of approximately 100 bp (Wolfson and Dressier 1972; Shinagawa and Padmanabhan 1980). Adenoviruses type 2 (Ad2) and type 5 (Ad5), the serotypes commonly used for in vitro DNA replication studies, have repeats of 102–103 bp (Steenbergh et al. 1977; Arrand and Roberts 1979; Shinagawa and Padmanabhan 1979). The 5′ terminus of each viral DNA strand is covalently linked via a phosphodiester bond to a protein (the terminal protein) of 55,000 daltons (55K) (Robinson et al. 1973; Rekosh et al. 1977; Carusi 1977). The linkage joins the β-hydroxyl of a serine residue in the protein to the 5′-terminal phosphate of the DNA (Desiderio and Kelly 1981). The adenoviral DNA-protein complex (Ad DNA-pro) replicates by a...