1H and 31P NMR investigations of actinomycin D binding selectivity with oligodeoxyribonucleotides containing multiple adjacent d(GC) sites.
Imino proton and 31P NMR studies were conducted on the binding of actinomycin D (ActD) to self-complementary oligodeoxyribonucleotides with adjacent 5'-GC-3' sites. ActD showed very high specificity for binding to GC sites regardless of oligomer length and surrounding sequence. For a first class of duplexes with a central GCGC sequence, a mixture of 1:1 complexes was observed due to the two different orientations of the ActD phenoxazone ring system. Analysis of 1H chemical shifts suggested that the favored 1:1 complex had the benzenoid side of the phenoxazone ring over the G base in the central base pair of the GCGC sequence. This is the first case in which an unsymmetrical intercalator has been shown to bind to DNA in both possible orientations. A unique 2:1 complex, with significantly different 1H and 31P chemical shifts relative to those of the 1:1 complexes, was formed with these same oligomers, again with the benzenoid side of the ActD molecule over the G base of the central GC base pair. There is considerable anticooperativity to binding of the second ActD in a GCGC sequence. In titrations of oligomers with the GCGC sequence, only the two 1:1 complexes are found up to ratios of one ActD per oligomer. Increasing the ActD concentration, however, resulted in stoichiometric formation of the unique 2:1 adduct. Spectrophotometric binding studies indicated that the apparent binding equilibrium constant for a GC site adjacent to a bound site is reduced by approximately a factor of 20 relative to the ActD binding constant to an isolated GC site.
Scott
E V
EV
Department of Chemistry, Georgia State University, Atlanta 30303.
Jones
R L
RL
Banville
D L
DL
Zon
G
G
Marzilli
L G
LG
Wilson
W D
WD
D001483:Base Sequence; D003609:Dactinomycin; D003841:Deoxycytidine; D003849:Deoxyguanosine; D006859:Hydrogen; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009838:Oligodeoxyribonucleotides; D010758:Phosphorus
D009838:Oligodeoxyribonucleotides; D003841:Deoxycytidine; D003609:Dactinomycin; D010758:Phosphorus; D006859:Hydrogen; D003849:Deoxyguanosine