Rate enhancements in the DNase I footprinting experiment.
Footprinting experiments for DNase I digests of a 139-base-pair segment of pBR-322 DNA in the presence of either netropsin or actinomycin D were carried out. Plots of oligonucleotide concentration as a function of drug concentration were analyzed to study the enhancement in cleavage rates at approximately 30 sites, accompanying drug binding at other sites. The pattern of enhancements is not consistent with drug-induced DNA structural changes, but agrees with a redistribution mechanism involving DNase I. Since the total number of enzyme molecules per fragment remains unchanged, drug binding at some sites increases the enzyme concentration at other sites, giving rise to increased cleavage. The consequences of the redistribution mechanism for analysis of footprinting experiments are indicated.
Ward
B
B
Department of Chemistry, Syracuse University, NY 13244-1200.
Rehfuss
R
R
Goodisman
J
J
Dabrowiak
J C
JC
D001482:Base Composition; D001483:Base Sequence; D004247:DNA; D004262:DNA Restriction Enzymes / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D003850:Deoxyribonuclease I / Q000378:metabolism*; D007700:Kinetics; D009429:Netropsin / Q000494:pharmacology; D009710:Nucleotide Mapping; D010957:Plasmids
D003609:Dactinomycin; D009429:Netropsin; D004247:DNA; D004262:DNA Restriction Enzymes; D003850:Deoxyribonuclease I