On the fidelity of DNA replication. Isolation of high fidelity DNA polymerase-primase complexes by immunoaffinity chromatography.


Error rates for conventionally purified DNA polymerase-alpha from calf thymus, chicken, and human sources have been reported to be one in 10,000 to one in 40,000 nucleotides incorporated. Isolation of polymerase-alpha by immunoaffinity chromatography yields a multiprotein high molecular weight replication complex that contains an associated DNA primase (Wong, S. W., Paborsky, L. R., Fisher, P. A., Wang, T. S-F., and Korn, D. (1986) J. Biol. Chem. 261, 7958-7968). We have isolated DNA polymerase-primase complexes from calf thymus, from a human lymphoblast cell line (TK-6), and from Chinese hamster lung cells (V-79) using two different methods of immunoaffinity chromatography. These enzyme complexes are 12- to 20-fold more accurate than conventionally purified calf thymus DNA polymerase-alpha when assayed using the phi X174am3 fidelity assay; estimated error rates are one in 460,000 to one in 830,000 nucleotides incorporated when the enzyme complex is freshly isolated. The polymerase-primase complex from calf thymus exhibited no detectable 3'----5' exonuclease activity using a heteroduplex substrate containing a single 3'-terminal mismatched nucleotide. Upon prolonged storage at -70 degrees C, the error rate of the immunoaffinity-purified calf thymus DNA polymerase-primase complex increases to about one in 50,000 nucleotides incorporated, an error rate similar to that exhibited by conventional isolates of DNA polymerase-alpha.




new topics/pols set partial results complete validated


No results available for this paper.

Entry validated by:

Log in to edit reference All References

Using Polbase tables:


Tables may be sorted by clicking on any of the column titles. A second click reverses the sort order. <Ctrl> + click on the column titles to sort by more than one column (e.g. family then name).


It is also possible to filter the table by typing into the search box above the table. This will instantly hide lines from the table that do not contain your search text.