Protein-DNA cross-linking demonstrates stepwise ATP-dependent assembly of T4 DNA polymerase and its accessory proteins on the primer-template

Todd L. Capson, Stephen J. Benkovic, Nancy G. Nossal

Research output: Contribution to journalArticlepeer-review

68 Scopus citations

Abstract

T4 DNA polymerase, the 44 62 and 45 polymerase accessory proteins, and 32 single-stranded DNA-binding protein catalyze ATP-dependent DNA synthesis. Using DNA primers with cross-linkable residues at specific positions, we obtained structural data that reveal how these proteins assemble on the primer-template. With the nonhydrolyzable ATP analog ATPγS, assembly of the 44 62 and 45 proteins on the primer requires 32 protein but not polymerase. ATP hydrolysis changes the position and intensity of cross-linking to each of the accessory proteins and allows cross-linking of polymerase. Our data indicate that the initial binding of the three accessory proteins and ATP to a 32 protein-covered primer-template is followed by ATP hydrolysis, binding of polymerase, and movement of the accessory proteins to yield a complex capable of processive DNA synthesis.

Original languageEnglish (US)
Pages (from-to)249-258
Number of pages10
JournalCell
Volume65
Issue number2
DOIs
StatePublished - Apr 19 1991

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)

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