A human factor that recognizes DNA substituted with 2-chloroadenine, an antileukemic purine analog.
2-Chloro-2'-deoxyadenosine (cladribine), an analog of deoxyadenosine, is an important new drug for the treatment of hairy cell leukemia and other forms of adult and pediatric leukemia. By a gel-shift binding assay, we identified an activity in HeLa nuclear extracts that recognizes and binds to oligonucleotides substituted with 2-chloroadenine (ClAde). The activity was specific for ClAde residues because control oligomers did not readily compete out the complex. The binding factor was a monomeric protein that was resistant to inactivation by heating at 45 degrees C but sensitive to heating at 65 degrees C, proteinase K treatment, and 5 mM ZnCl2. This protein, designated ClAde recognition protein (CARP), appeared to be related to a protein that recognized other forms of DNA damage. Gel-shift binding reactions with ultraviolet (UV)-irradiated oligomers revealed a UV-specific protein/DNA complex that had an electrophoretic mobility similar to that of the CARP/DNA complex, and CARP binding to ClAde-containing oligomers was readily competed out by UV-irradiated DNA. Moreover, CARP activity was present in extracts prepared from UV-sensitive xeroderma pigmentosum group A cells but not in a subset of cells from group E, suggesting that CARP was similar to a previously described repair associated factor, xeroderma pigmentosum-E binding factor. Our findings support a possible repair process for ClAde residues incorporated into cellular DNA.
Hentosh
P
P
Department of Pharmacology and Molecular Biology, Chicago Medical School, Illinois 60064, USA.
Tibudan
M
M
Grippo
P
P
D000225:Adenine / Q000031:analogs & derivatives* / Q000378:metabolism; D001483:Base Sequence; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D014983:Xeroderma Pigmentosum / Q000378:metabolism
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; C069484:2-chloroadenine; D000225:Adenine