Adenovirus DNA synthesis in vitro in an isolated complex

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Abstract
DNA-protein complexes isolated from adenovirus-infected [human oral carcinoma KB] cells by a modification of the M-band technique were used as an in vitro system for the study of adenovirus DNA replication. The synthesis in vitro was semiconservative, inhibited by N-ethylmaleimide and stimulated by ATP. Studies on DNA-negative mutants of adenovirus showed that the DNA synthesis in vitro represents a continuation of adenovirus DNA replication in vivo. DNA synthesis in vitro was inhibited 38% by 20 .mu.g of phosphonoacetic acid/ml, which is several-fold higher than the inhibition obtained with purified DNA polymerase .beta. or .gamma., but was similar to the degree of inhibition of DNA polymerase .alpha.. DNA synthesis in complexes from uninfected cells was much less sensitive to inhibition by phosphonoacetic acid. Complexes from infected cells contained a greater proportion of the .alpha.-polymerase than complexes from uninfected cells, suggesting that an association of .alpha.-polymerase with the replication complex may be occurring during adenovirus infection, with subsequent utilization of the .alpha.-polymerase for viral DNA synthesis.