Mechanisms of coupling between DNA recognition specificity and catalysis in EcoRI endonuclease.
Proteins that bind to specific sites on DNA often do so in order to carry out catalysis or specific protein-protein interaction while bound to the recognition site. Functional specificity is enhanced if this second function is coupled to correct DNA site recognition. To analyze the structural and energetic basis of coupling between recognition and catalysis in EcoRI endonuclease, we have studied stereospecific phosphorothioate (PS) or methylphosphonate (PMe) substitutions at the scissile phosphate GpAATTC or at the adjacent phosphate GApATTC in combination with molecular-dynamics simulations of the catalytic center with bound Mg2+. The results show the roles in catalysis of individual phosphoryl oxygens and of DNA distortion and suggest that a "crosstalk ring" in the complex couples recognition to catalysis and couples the two catalytic sites to each other.
Kurpiewski
Michael R
MR
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.
Engler
Lisa E
LE
Wozniak
Lucyna A
LA
Kobylanska
Anna
A
Koziolkiewicz
Maria
M
Stec
Wojciech J
WJ
Jen-Jacobson
Linda
L
D000227:Adenine Nucleotides / Q000737:chemistry; D001483:Base Sequence; D001665:Binding Sites; D002384:Catalysis; D020134:Catalytic Domain; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D015246:Deoxyribonuclease EcoRI / Q000737:chemistry* / Q000378:metabolism; D008274:Magnesium / Q000737:chemistry; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D009943:Organophosphorus Compounds / Q000737:chemistry; D010100:Oxygen / Q000737:chemistry; D010710:Phosphates / Q000737:chemistry; D013379:Substrate Specificity
D000227:Adenine Nucleotides; D009943:Organophosphorus Compounds; D010710:Phosphates; C032627:methylphosphonic acid; C083120:phosphorodithioic acid; D004247:DNA; D015246:Deoxyribonuclease EcoRI; D008274:Magnesium; D010100:Oxygen