Cross-linking of proteins to DNA in newly synthesized chromatin by diisopropylfluorophosphate, a serine protease inhibitor.
When nascent DNA of SV40 pulse labeled with [alpha-32p]dCTP in a permeable cell system was treated in situ with diisopropylfluorophosphate (DFP), a significant fraction of radioactivity was found to be covalently complexed with proteins. The adduct formation was demonstrated by density separation in CsCl, selective precipitation of the complexed DNA with SDS-KC1, and visualization of cross-linked proteins after SDS-PAGE. No cross-linking occurred with mature SV40 chromatin labeled in vivo and extracted from nuclei of infected cells. The DFP-induced DNA-protein cross-linking reaction appears to involve the protein's sulfhydryl groups since pretreatment with some sulfhydryl reagents completely inhibited the reaction.
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D002843:Chromatin / Q000378:metabolism*; D003432:Cross-Linking Reagents; D004279:DNA, Viral / Q000378:metabolism*; D007531:Isoflurophate / Q000494:pharmacology*; D007668:Kidney; D007700:Kinetics; D009698:Nucleoproteins / Q000378:metabolism*; D011480:Protease Inhibitors; D011485:Protein Binding; D012697:Serine Endopeptidases; D013539:Simian virus 40 / Q000235:genetics
D002843:Chromatin; D003432:Cross-Linking Reagents; D004279:DNA, Viral; D009698:Nucleoproteins; D011480:Protease Inhibitors; D007531:Isoflurophate; D012697:Serine Endopeptidases