Presence of the anti-leukemic nucleotide analog, 2-chloro-2′-deoxyadenosine-5′-monophosphate, in a promoter sequence alters DNA binding of TATA-binding protein (TBP)

William R. Hartman, D. Eric Walters, Patricia Hentosh

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

2-Chlorodeoxyadenosine (CldAdo, Cladribine), a nucleoside analog used in the treatment of hairy cell leukemia, is phosphorylated and incorporated into DNA, but is not an absolute chain terminator. We hypothesized that the presence of a chlorine molecule projecting into the DNA minor groove would affect DNA:protein-binding interactions. Here, we investigated recognition of and binding to double-stranded CldAMP-substituted TATA promoter sequences by human TATA-binding protein (TBP) using mobility shift assays. Depending on the site, CldAMP in place of dAMP within a TATA sequence decreased in vitro TBP binding by ∼30% to 55% compared to control sites. When bound to a CldAMP-substituted TATA box, however, the TBP complex was more resistant to polyanions, suggesting enhanced stability. Limited exposure of the TBP:DNA complex to proteases indicated that TBP conformation was altered on CldAMP-substituted DNA compared to control. Further, binding of transcription factor IIB to TBP was diminished on analog-containing TATA sequences. These results suggest normal TBP-binding interactions-specifically recognition, stability, and conformation-are disrupted by CldAMP insertion into eukaryotic promoter sequences.

Original languageEnglish (US)
Pages (from-to)223-232
Number of pages10
JournalArchives of Biochemistry and Biophysics
Volume459
Issue number2
DOIs
StatePublished - Mar 15 2007

Keywords

  • Cancer drug
  • Cladribine
  • DNA minor groove
  • Hairy cell leukemia
  • Promoter sequence
  • Protein conformation
  • Protein:protein associations
  • RNA polymerase II
  • TATA-binding protein
  • Transcription factors

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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