Phosphodiesterase 2

All the salts and solvents, activated calf thymus DNA, nuclease P1 from Penicillium citrinum, phosphodiesterase II, phosphodiesterase I from Crotalus Adamanteus venom, DNAse I, DNAse II, alkaline phosphatase from bovine intestinal mucosa, erythro-9-(2-hydroxy-3-nonyl)adenine hydrochoride (EHNA), benzonase 99%, deferoxamine mesylate salt, BHT and pentostatin, were obtained from Sigma (Taufkirchen, Germany and Milan, Italy) while the 3 and 100 kDa cut-off filters were purchased from Millipore (Bedford, USA). Distilled and deionized water (ddH₂O) was purified by a Milli-Q system (Millipore, Bedford, USA). Synthesis of reference compounds and internal standards was based on previously reported protocols (Terzidis and Chatgilialoglu, in preparation).

Uvr proteins were purified as previously described [27 ]. Monoclonal antibodies to acrolein-dG DNA adducts were prepared as described [14 (link)]. UPIII antibodies were purchased from Abcam (Cambridge, MA), α-tubulin from Calbiochem (Billerica, MA), and antibodies against mouse/rabbit IgG from Amersham Biosciences (Pittsburgh, PA). Normal human bladder epithelial cells (HBEP cells) were obtained from Zen-Bio (Research Triangle Park, NC). UROtsa cells were obtained from the laboratory of JR Masters (University College London, UK) and normal human fibroblasts (CCL202 cells) were obtained from ATCC (American Type Culture Collection, Manassas, VA). Plasmid pSP189 was prepared as described by Canella and Seidman [28 (link)]. T4 kinase, protease K, and RNase A were purchased from New England Biolabs (Ipswich, MA); γ-³²P-ATP and α-³²P-dATP were purchased from Perkin Elmer (Waltham, MA), and nuclease P1 and phosphodiesterase II were purchased from Sigma (St. Louis, MO) and Worthington (Lakewood, NJ), respectively. Acrolein (purity 90%) was purchased from Sigma and N-hydroxy-4-ABP (N-OH-4-ABP) was synthesized as previously described (34).

8-Bromo-2′-deoxyadenosine, 2′-deoxyadenosine monohydrated, 7,8-dihydro-8-oxo-2′-deoxyadenosine (8-oxo-dA), 2′-deoxyguanosine monohydrated, 7,8-dihydro-8-oxo-2′-deoxyguanosine 8-oxo-dG), and 8-bromo-2′-deoxyguanosine were purchased from Berry & Associates Inc. (Dexter, USA). [¹⁵N₅]-2′-deoxyadenosine monohydrated and [¹⁵N₅]-2′-deoxyguanosine monohydrated (all >98% isotopic purity) were purchased from Cambridge Isotope Laboratories (Andover, USA). All the salts and solvents, thymidine, 2′-deoxycytidine, activated calf thymus DNA, nuclease P1 from Penicillium citrinum, phosphodiesterase II, phosphodiesterase I from Crotalus Adamanteus venom, DNAse I, DNAse II, alkaline phosphatase from bovine intestinal mucosa, erythro-9-(2-hydroxy-3-nonyl)adenine hydrochoride (EHNA), benzonase 99%, deferoxamine mesylate salt, BHT and pentostatin, were obtained from Sigma (Taufkirchen, Germany and Milan, Italy) while the 3 kDa filters were purchased from Millipore (Bedford, USA). Distilled and deionized water (ddH2O) was purified by a Milli-Q system (Millipore, Bedford, USA).