Indirect readout of DNA sequence at the primary-kink site in the CAP-DNA complex: recognition of pyrimidine-purine and purine-purine steps

J Mol Biol. 2006 Mar 17;357(1):173-83. doi: 10.1016/j.jmb.2005.12.051. Epub 2006 Jan 3.

Abstract

The catabolite activator protein (CAP) bends DNA in the CAP-DNA complex, typically introducing a sharp DNA kink, with a roll angle of approximately 40 degrees and a twist angle of approximately 20 degrees, between positions 6 and 7 of the DNA half-site, 5'-A1A2A3T4G5T6G7A8T9C10T11 -3' ("primary kink"). In previous work, we showed that CAP recognizes the nucleotide immediately 5' to the primary-kink site, T6, through an "indirect-readout" mechanism involving sequence effects on energetics of primary-kink formation. Here, to understand further this example of indirect readout, we have determined crystal structures of CAP-DNA complexes containing each possible nucleotide at position 6. The structures show that CAP can introduce a DNA kink at the primary-kink site with any nucleotide at position 6. The DNA kink is sharp with the consensus pyrimidine-purine step T6G7 and the non-consensus pyrimidine-purine step C6G7 (roll angles of approximately 42 degrees, twist angles of approximately 16 degrees ), but is much less sharp with the non-consensus purine-purine steps A6G7 and G6G7 (roll angles of approximately 20 degrees, twist angles of approximately 17 degrees). We infer that CAP discriminates between consensus and non-consensus pyrimidine-purine steps at positions 6-7 solely based on differences in the energetics of DNA deformation, but that CAP discriminates between the consensus pyrimidine-purine step and non-consensus purine-purine steps at positions 6-7 both based on differences in the energetics of DNA deformation and based on qualitative differences in DNA deformation. The structures further show that CAP can achieve a similar, approximately 46 degrees per DNA half-site, overall DNA bend through a sharp DNA kink, a less sharp DNA kink, or a smooth DNA bend. Analysis of these and other crystal structures of CAP-DNA complexes indicates that there is a large, approximately 28 degrees per DNA half-site, out-of-plane component of CAP-induced DNA bending in structures not constrained by end-to-end DNA lattice interactions and that lattice contacts involving CAP tend to involve residues in or near biologically functional surfaces.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Cyclic AMP Receptor Protein / chemistry*
  • Cyclic AMP Receptor Protein / genetics
  • Cyclic AMP Receptor Protein / metabolism
  • DNA / chemistry*
  • DNA / metabolism
  • Escherichia coli / metabolism
  • Macromolecular Substances
  • Models, Molecular
  • Molecular Sequence Data
  • Nucleic Acid Conformation*
  • Protein Structure, Tertiary*
  • Purines / chemistry*
  • Pyrimidines / chemistry*
  • Sequence Analysis, DNA*

Substances

  • Cyclic AMP Receptor Protein
  • Macromolecular Substances
  • Purines
  • Pyrimidines
  • DNA

Associated data

  • PDB/1ZRC
  • PDB/1ZRD
  • PDB/1ZRE
  • PDB/1ZRF