Analysis of a curved DNA constructed from alternating dAn:dTn-tracts in linear and supercoiled form by high resolution chemical probing.
Complex of osmium tetroxide and bipyridine (Os,bipy), KMnO4, and diethyl pyrocarbonate (DEPC) were used to probe curved DNA at single nucleotide resolution. The DNA was constructed from repeated dAn:dTn-blocks with dATATA and dAGAGA interblock sequences. The DNA was probed in the linear and supercoiled form at various salt concentrations. While all purines were available for DEPC attack, the thymines within the blocks were resistant to chemical probing by KMnO4 and Os,bipy. Only the 3'-flanking dTs were available for modification. The thymines within dTC and dTA sequences showed modification indicating that these thymines display an unstacked structure allowing both probes to attack. Under destabilizing conditions, at low ionic strength and superhelical stress, considerable unstacking was observed. We found experimental indications that under these destabilizing conditions unpaired regions might appear, probably within the dATATA sequence.
Institute of Biophysics, Academy of Sciences of the Czech Republic, Brno, Czech Republic. kane@ibp.cz
K
K
Nejedlý
E
E
Sýkorová
S
S
Diekmann
E
E
Palecek
D004278:DNA, Superhelical / Q000737:chemistry*; D003839:Deoxyadenosines / Q000737:chemistry*; D004047:Diethyl Pyrocarbonate / Q000737:chemistry; D009690:Nucleic Acid Conformation; D009993:Osmium Tetroxide / Q000737:chemistry; D010957:Plasmids / Q000737:chemistry; D011196:Potassium Permanganate / Q000737:chemistry; D011687:Purines / Q000737:chemistry; D013936:Thymidine / Q000737:chemistry*
D004278:DNA, Superhelical; D003839:Deoxyadenosines; D011687:Purines; D011196:Potassium Permanganate; D004047:Diethyl Pyrocarbonate; D009993:Osmium Tetroxide; D013936:Thymidine; C030985:purine