caa a22 4500 477119158 CHVBK 20180405111631.0 cr unu---uuuuu 170330e19960901xx s 000 0 eng 10.1007/BF00420918 doi (NATIONALLICENCE)springer-10.1007/BF00420918 Association of poly(ADP-ribose) polymerase with nuclear subfractions catalyzed with sodium tetrathionate and hydrogene peroxide crosslinks [Elektronische Daten] [Serge Desnoyers, James Kirkland, Guy Poirier] Poly(ADP-ribose) polymerase (PARP) is a nuclear enzyme which catalyzes the transfer of ADP-ribose units from NAD+ to a variety of nuclear proteins under the stimulation of DNA strand break. To examine its role in DNA repair, we have been studying the interaction of PARP with other nuclear proteins using disulfide cross-linking, initiated by sodium tetrathionate (NaTT). Chinese Hamster Ovary (CHO) cells were extracted sequentially with Nonidet P40 (detergent), nucleases (DNase + RNase), and high salt (1.6 M NaCl) with and without the addition of a sulfhydryl reducing agent. The residual structures are referred to as the nuclear matrix, and are implicated in the organization of DNA repair and replication. Treatment of the cells with NaTT causes the crosslinking of PARP to the nuclear matrix. Activating PARP by pretreating the cells with H2O2 did not increase the cross-linking of PARP with the nuclear matrix, suggesting a lack of additional interaction of the enzyme with the nuclear matrix during DNA repair. Both NaTT and H2O2 induced crosslinks of PARP that were extractable with high salt. To shorten the procedure, these crosslinks were extracted from cells without nucleases and high salt treatment, using phosphate buffer. Using western blotting, these crosslinks appeared as a smear of high molecular weight species including a possible dimer of PARP at 230 kDa, which return to 116 kDa following reduction with β-mercaptoethanol. Kluwer Academic Publishers, 1996 crosslinking nationallicence sodium tetrathionate nationallicence protein-protein interaction nationallicence poly(ADP-ribose) polymerase nationallicence oxidative stress nationallicence Desnoyers Serge Research Group on Poly(ADP-ribose) Metabolism, Health and Environment Unit, CHUL Research Center and Laval University, GIV 4G2, Sainte-Foy, Quebec, Canada aut Kirkland James Research Group on Poly(ADP-ribose) Metabolism, Health and Environment Unit, CHUL Research Center and Laval University, GIV 4G2, Sainte-Foy, Quebec, Canada aut Poirier Guy Research Group on Poly(ADP-ribose) Metabolism, Health and Environment Unit, CHUL Research Center and Laval University, GIV 4G2, Sainte-Foy, Quebec, Canada aut Molecular and Cellular Biochemistry Kluwer Academic Publishers 159/2(1996-09-01), 155-161 0300-8177 159:2<155 1996 159 11010 https://doi.org/10.1007/BF00420918 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/BF00420918 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Desnoyers Serge Research Group on Poly(ADP-ribose) Metabolism, Health and Environment Unit, CHUL Research Center and Laval University, GIV 4G2, Sainte-Foy, Quebec, Canada aut NATIONALLICENCE 700 1- Kirkland James Research Group on Poly(ADP-ribose) Metabolism, Health and Environment Unit, CHUL Research Center and Laval University, GIV 4G2, Sainte-Foy, Quebec, Canada aut NATIONALLICENCE 700 1- Poirier Guy Research Group on Poly(ADP-ribose) Metabolism, Health and Environment Unit, CHUL Research Center and Laval University, GIV 4G2, Sainte-Foy, Quebec, Canada aut NATIONALLICENCE 773 0- Molecular and Cellular Biochemistry Kluwer Academic Publishers 159/2(1996-09-01), 155-161 0300-8177 159:2<155 1996 159 11010 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer