Constitution of the twin polymerase of DNA polymerase III holoenzyme.
It is speculated that DNA polymerases which duplicate chromosomes are dimeric to provide concurrent replication of both leading and lagging strands. DNA polymerase III holoenzyme (holoenzyme), is the 10-subunit replicase of the Escherichia coli chromosome. A complex of the alpha (DNA polymerase) and epsilon (3'-5' exonuclease) subunits of the holoenzyme contains only one of each protein. Presumably, one of the eight other subunit(s) functions to dimerize the alpha epsilon polymerase within the holoenzyme. Based on dimeric subassemblies of the holoenzyme, two subunits have been elected as possible agents of polymerase dimerization, one of which is the tau subunit (McHenry, C. S. (1982) J. Biol. Chem. 257, 2657-2663). Here, we have used pure alpha, epsilon, and tau subunits in binding studies to determine whether tau can dimerize the polymerase. We find tau binds directly to alpha. Whereas alpha is monomeric, tau is a dimer in its native state and thereby serves as an efficient scaffold to dimerize the polymerase. The epsilon subunit does not associate directly with tau but becomes dimerized in the alpha epsilon tau complex by virtue of its interaction with alpha. We have analyzed the dimeric alpha epsilon tau complex by different physical methods to increase the confidence that this complex truly contains a dimeric polymerase. The tau subunit is comprised of the NH2-terminal two-thirds of tau but does not bind to alpha epsilon, identifying the COOH-terminal region of tau as essential to its polymerase dimerization function. The significance of these results with respect to the organization of subunits within the holoenzyme is discussed.
Studwell-Vaughan
P S
PS
Microbiology Department, Howard Hughes Medical Institute, Cornell University Medical College, New York, New York 10021.
O'Donnell
M
M
D002850:Chromatography, Gel; D002876:Chromosomes, Bacterial; D004258:DNA Polymerase III / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005798:Genes, Bacterial; D011487:Protein Conformation; D013379:Substrate Specificity
D004258:DNA Polymerase III