Synthetic repeating sequence DNAs containing phosphorothioates: nuclease sensitivity and triplex formation.
More than twenty repeating sequence DNAs containing phosphorothioates were prepared from the appropriate dXTPs with DNA polymerase I. The Tms of the modified DNAs were all lower than the parent polymers. A phosphorothioate group 5' to a pyrimidine gave rise to a large decrease than 5' to a purine, e.g., poly(dA).poly(dT) = 50 degrees; poly(dsA).poly(dT) = 44 degrees; poly(dA).poly(dsT) = 33 degrees; and poly(dsA).poly(dsT) = 26 degrees. The presence of phosphorothioate groups had a dramatic effect on triplex formation; poly[d(TC)].poly[d(sGsA)] spontaneously dismutases to a triplex at pH 8 whereas triplex formation in poly[d(sTsC)].poly[d(GA)] was inhibited. Surprisingly poly(dsG).poly(dC) had a Tm which initially decreased with increasing ionic strength. Resistance to digestion with pancreatic DNAse I did not correlate with phosphorothioate content. Poly[d(AsT)], poly[d(TsC)].poly[d(sGA)] and poly[d(sTG)].poly[d(sCA)] were resistant whereas poly[d(sAT)] and poly[d(sTsTG)].poly[d(CsAsA)] were rapidly degraded. Thus phosphorothioate groups cause small conformational changes and may reveal new families of conformational polymorphisms.
Latimer
L J
LJ
Department of Biochemistry, University of Saskatchewan, Saskatoon, Canada.
Hampel
K
K
Lee
J S
JS
D000911:Antibodies, Monoclonal; D004256:DNA Polymerase I / Q000378:metabolism; D007202:Indicators and Reagents; D007700:Kinetics; D009691:Nucleic Acid Denaturation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis*; D009994:Osmolar Concentration; D011089:Polydeoxyribonucleotides / Q000138:chemical synthesis*; D013816:Thermodynamics; D013873:Thionucleotides
D000911:Antibodies, Monoclonal; D007202:Indicators and Reagents; D009838:Oligodeoxyribonucleotides; D011089:Polydeoxyribonucleotides; D013873:Thionucleotides; D004256:DNA Polymerase I