Facilitated distortion of the DNA site enhances EcoRI endonuclease-DNA recognition.
We have measured the binding of EcoRI endonuclease to a complete set of purine-base analogue sites, each of which deletes one functional group that forms a hydrogen bond with the endonuclease in the canonical GAATTC complex. For five of six functional group deletions, the observed penalty in binding free energy is +1.3 to +1.7 kcal/mol. For two of these cases (replacement of adenine N7 with carbon) a single protein-base hydrogen bond is removed without deleting an interstrand Watson-Crick hydrogen bond or causing structural "adaptation" in the complex. This observation establishes that the incremental energetic contribution of one protein-base hydrogen bond is about -1.5 kcal/mol. By contrast, deletion of the N6-amino group of the inner adenine in the site improves binding by -1.0 kcal/mol because the penalty for deleting a protein-base hydrogen bond is outweighed by facilitation of the required DNA distortion ("kinking") in the complex. This result provides direct evidence that the energetic cost of distorting a DNA site can make an unfavorable contribution to protein-DNA binding.
Lesser
D R
DR
Department of Biological Sciences, University of Pittsburgh, PA 15260.
Kurpiewski
M R
MR
Waters
T
T
Connolly
B A
BA
Jen-Jacobson
L
L
D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D002151:Calorimetry; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D015246:Deoxyribonuclease EcoRI / Q000737:chemistry / Q000378:metabolism*; D006860:Hydrogen Bonding; D007700:Kinetics; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011487:Protein Conformation; D017384:Sequence Deletion
D004247:DNA; D015246:Deoxyribonuclease EcoRI