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DNA helicases associated with DNA polymerases from human cells
T Sakurai1, M Suzuki, S Yoshida
1Laboratory of Cancer Cell Biology, Nagoya University School of Medicine, Japan.
Biochemistry and Molecular Biology International
|March 1, 1993
Summary
This study separated DNA polymerase alpha, delta, and epsilon. DNA helicase activities were found associated with DNA polymerase alpha and delta, suggesting large complex formation.
Area of Science:
- Molecular Biology
- Biochemistry
Background:
- DNA polymerases are crucial for DNA replication and repair.
- DNA helicases unwind DNA, a process essential for replication.
- The association between DNA polymerases and helicases is not fully understood.
Purpose of the Study:
- To separate and characterize DNA polymerase alpha, delta, and epsilon.
- To identify and characterize DNA helicase activities associated with these polymerases.
- To investigate the physical association between DNA polymerases and DNA helicases.
Main Methods:
- Fractionation of HeLa cell extracts using hydroxylapatite and heparin columns.
- Characterization of DNA helicase activity and substrate requirements.
- Size exclusion chromatography (Superose 6) to determine complex molecular weights.
Main Results:
- DNA polymerase alpha, delta, and epsilon were successfully separated.
- DNA helicase activities were detected in fractions containing DNA polymerase alpha and delta.
- These DNA helicases exhibited 3' to 5' translocation on single-stranded DNA and required ATP/dATP.
- Size exclusion chromatography indicated that DNA helicase activities were associated with DNA polymerase alpha (approx. 600 kDa) and delta (approx. 400 kDa) complexes.
Conclusions:
- DNA helicases are physically associated with DNA polymerase alpha and delta.
- These associations form large functional complexes, suggesting a coordinated role in DNA replication or repair.