Mean DNA bend angle and distribution of DNA bend angles in the CAP-DNA complex in solution

Achillefs N. Kapanidis, Yon W. Ebright, Richard D. Ludescher, Shirley Chan, Richard H. Ebright

Research output: Contribution to journalArticlepeer-review

47 Scopus citations

Abstract

In order to define the mean DNA bend angle and distribution of DNA bend angles in the catabolite activator protein (CAP)-DNA complex in solution under standard transcription initiation conditions, we have performed nanosecond time-resolved fluorescence measurements quantifying energy transfer between a probe incorporated at a specific site in CAP, and a complementary probe incorporated at each of five specific sites in DNA. The results indicate that the mean DNA bend angle is 77(±-3)° - consistent with the mean DNA bend angle observed in crystallographic structures (80(±12)°). Lifetime-distribution analysis indicates that the distribution of DNA bend angles is relatively narrow, with <10% of DNA bend angles exceeding 100°. Millisecond time-resolved luminescence measurements using lanthanide-chelate probes provide independent evidence that the upper limit of the distribution of DNA bend angles is ∼100°. The methods used here will permit mutational analysis of CAP-induced DNA bending and the role of CAP-induced DNA bending in transcriptional activation.

Original languageEnglish (US)
Pages (from-to)453-468
Number of pages16
JournalJournal of molecular biology
Volume312
Issue number3
DOIs
StatePublished - Sep 21 2001

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Molecular Biology

Keywords

  • Catabolite activator protein (CAP)
  • Fluorescence resonance energy transfer
  • Protein-induced DNA bending
  • Transcriptional activation
  • cAMP receptor protein (CRP)

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