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. 1989 Sep;86(17):6479–6483. doi: 10.1073/pnas.86.17.6479

Production of simian virus 40 large tumor antigen in bacteria: altered DNA-binding specificity and dna-replication activity of underphosphorylated large tumor antigen.

I J Mohr 1, Y Gluzman 1, M P Fairman 1, M Strauss 1, D McVey 1, B Stillman 1, R D Gerard 1
PMCID: PMC297867  PMID: 2549538

Abstract

A bacterial expression system was used to produce simian virus 40 large tumor antigen (T antigen) in the absence of the extensive posttranslational modifications that occur in mammalian cells. Wild-type T antigen produced in bacteria retained a specific subset of the biochemical activities displayed by its mammalian counterpart. Escherichia coli T antigen functioned as a helicase and bound to DNA fragments containing either site I or the wild-type origin of replication in a manner identical to mammalian T antigen. However, T antigen purified from E. coli did not efficiently bind to site II, an essential cis element within the simian virus 40 origin of replication. It therefore could not unwind origin-containing plasmids or efficiently replicate simian virus 40 DNA in vitro. The ability of protein phosphorylation to modulate the intrinsic preference of full-length T antigen for either site I or site II is discussed.

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