Increased ionic strength: effects on DNA fragmentation and poly(ADP-ribose) polymerase activity in HeLa nuclei.
Poly(ADP-ribose) polymerase activity was measured in a crude nuclear fraction isolated from HeLa cells. It was found that the addition of ammonium sulfate or other salts to the standard incubation medium inhibited the formation of poly(ADP-ribose). Through the use of alkaline sucrose density gradients it was also noted that this same increase in ionic strength inhibited the in vitro breakdown of the HeLa DNA. Additional experiments with alkaline sucrose density gradients and deoxyribonuclease I showed that the in vitro activity of poly(ADP-ribose) polymerase is largely dependent upon DNA fragmentation but that DNA fragmentation at least in vitro is not dependent upon the formation of poly(ADP-ribose). These observations imply that this nuclear enzyme is not extremely sensitive to changes in the ionic strength of the reaction media but is affected indirectly, supposedly through changes in the endonuclease activity of the HeLa nuclei. If this proves to be true, then the addition of salt to the incubation medium for poly(ADP-ribose) polymerase could prove to be a valuable tool for the study of ADP-ribosylation reactions.
D000645:Ammonium Sulfate / Q000494:pharmacology; D002467:Cell Nucleus / Q000201:enzymology*; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D003850:Deoxyribonuclease I / Q000378:metabolism; D006367:HeLa Cells / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D009994:Osmolar Concentration; D011064:Poly Adenosine Diphosphate Ribose / Q000096:biosynthesis; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D013696:Temperature
D011064:Poly Adenosine Diphosphate Ribose; D004247:DNA; D011065:Poly(ADP-ribose) Polymerases; D003850:Deoxyribonuclease I; D000645:Ammonium Sulfate