Bending and flexibility of methylated and unmethylated EcoRI DNA.
We used cyclization kinetics experiments and Monte Carlo simulations to determine a structural model for a DNA decamer containing the EcoRI restriction site. Our findings agree well with recent crystal and NMR structures of the EcoRI dodecamer, where an overall bend of seven degrees is distributed symmetrically over the molecule. Monte Carlo simulations indicate that the sequence has a higher flexibility, assumed to be isotropic, compared to that of a "generic" DNA sequence. This model was used as a starting point for the investigation of the effect of cytosine methylation on DNA bending and flexibility. While methylation did not affect bend magnitude or direction, it resulted in a reduction in bending flexibility and under-winding of the methylated nucleotides. We demonstrate that our approach can augment the understanding of DNA structure and dynamics by adding information about the global structure and flexibility of the sequence. We also show that cyclization kinetics can be used to study the properties of modified nucleotides.
Nathan
Dafna
D
Departments of Chemistry, Yale University, New Haven, CT 06520, USA.
Crothers
Donald M
DM
D001483:Base Sequence; D001665:Binding Sites; D003198:Computer Simulation; D003500:Cyclization; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation; D015246:Deoxyribonuclease EcoRI / Q000378:metabolism*; D007700:Kinetics; D009010:Monte Carlo Method; D009690:Nucleic Acid Conformation; D018583:Pliability; D013379:Substrate Specificity
D004247:DNA; D015246:Deoxyribonuclease EcoRI