Analysis of the ATPase subassembly which initiates processive DNA synthesis by DNA polymerase III holoenzyme.
The gamma complex (gamma delta delta' chi psi) subassembly of DNA polymerase III holoenzyme transfers the beta subunit onto primed DNA in a reaction which requires ATP hydrolysis. Once on DNA, beta is a "sliding clamp" which tethers the polymerase to DNA for highly processive synthesis. We have examined beta and the gamma complex to identify which subunit(s) hydrolyzes ATP. We find the gamma complex is a DNA dependent ATPase. The beta subunit, which lacks ATPase activity, enhances the gamma complex ATPase when primed DNA is used as an effector. Hence, the gamma complex recognizes DNA and couples ATP hydrolysis to clamp beta onto primed DNA. Study of gamma complex subunits showed no single subunit contained significant ATPase activity. However, the heterodimers, gamma delta and gamma delta', were both DNA-dependent ATPases. Only the gamma delta ATPase was stimulated by beta and was functional in transferring the beta from solution to primed DNA. Similarity in ATPase activity of DNA polymerase III holoenzyme accessory proteins to accessory proteins of phage T4 DNA polymerase and mammalian DNA polymerase delta suggests the basic strategy of chromosome duplication has been conserved throughout evolution.
Onrust
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Howard Hughes Medical Institute, Microbiology Department, Cornell University Medical College, New York, New York 10021.
Stukenberg
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O'Donnell
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D000251:Adenosine Triphosphatases / Q000378:metabolism*; D010584:Bacteriophage phi X 174 / Q000235:genetics; D003090:Coliphages / Q000235:genetics; D004265:DNA Helicases / Q000378:metabolism; D004258:DNA Polymerase III / Q000378:metabolism*; D004261:DNA Replication; D004279:DNA, Viral / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D011061:Poly A / Q000378:metabolism; D011067:Poly dA-dT / Q000378:metabolism; D013379:Substrate Specificity; D013698:Templates, Genetic; D013816:Thermodynamics
D004279:DNA, Viral; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; C027903:oligo (dT); D011061:Poly A; C015465:poly(dA); D011067:Poly dA-dT; D004258:DNA Polymerase III; D000251:Adenosine Triphosphatases; D004265:DNA Helicases