Effect of calcium on the recovery and distribution of DNA polymerase alpha from cultured human cells.
It was recently reported (Lynch, W. E., Surrey, S., and Lieberman, I. (1975) J. Biol. Chem. 250, 8179-8183) that the extraction of regenerating rat liver in solutions of isotonic sucrose containing 4 mM CaCl2 leads to almost quantitative recovery of DNA polymerase alpha (Weissbach, A., Baltimore, D., Bollum, F., Gallo, R., and Korn, D. (1975) Science 190, 401--402) activity in the purified nuclear compartment. Our application of this method to the isolation of the DNA polymerase activities activities from cultured human epithelial and lymphoblastoid cells has led to substantially different results. We have observed that the inclusion of Ca2+ in either isotonic sucrose or hypotonic aqueous extraction media leads to the irreversible inactivation of the majority, cytoplasmic fraction of DNA polymerase alpha activity and is without quantitative effect on the recovery of the nuclear fraction of this activity.
D002118:Calcium / Q000494:pharmacology*; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000201:enzymology; D004257:DNA Polymerase II / Q000302:isolation & purification*; D004259:DNA-Directed DNA Polymerase / Q000302:isolation & purification*; D006801:Humans; D008274:Magnesium; D008722:Methods; D009994:Osmolar Concentration
D004257:DNA Polymerase II; D004259:DNA-Directed DNA Polymerase; D008274:Magnesium; D002118:Calcium