Molecular determinants for DNA minor groove recognition: design of a bis-guanidinium derivative of ethidium that is highly selective for AT-rich DNA sequences.
The phenanthridinium dye ethidium bromide is a prototypical DNA intercalating agent. For decades, this anti-trypanosomal agent has been known to intercalate into nucleic acids, with little preference for particular sequences. Only polydA-polydT tracts are relatively refractory to ethidium intercalation. In an effort to tune the sequence selectivity of known DNA binding agents, we report here the synthesis and detailed characterization of the mode of binding to DNA of a novel ethidium derivative possessing two guanidinium groups at positions 3 and 8. This compound, DB950, binds to DNA much more tightly than ethidium and exhibits distinct DNA-dependent absorption and fluorescence properties. The study of the mode of binding to DNA by means of circular and electric linear dichroism revealed that, unlike ethidium, DB950 forms minor groove complexes with AT sequences. Accurate quantification of binding affinities by surface plasmon resonance using A(n)T(n) hairpin oligomer indicated that the interaction of DB950 is over 10-50 times stronger than that of ethidium and comparable to that of the known minor groove binder furamidine. DB950 interacts weakly with GC sites by intercalation. DNase I footprinting experiments performed with different DNA fragments established that DB950 presents a pronounced selectivity for AT-rich sites, identical with that of furamidine. The replacement of the amino groups of ethidium with guanidinium groups has resulted in a marked gain of both affinity and sequence selectivity. DB950 provides protection against DNase I cleavage at AT-containing sites which frequently correspond to regions of enhanced cleavage in the presence of ethidium. Although DB950 maintains a planar phenanthridinium chromophore, the compound no longer intercalates at AT sites. The guanidinium groups of DB950, just like the amidinium group of furamidine (DB75), are the critical determinants for recognition of AT binding sites in DNA. The chemical modulation of the ethidium exocyclic amines is a profitable option to tune the nucleic acid recognition properties of phenylphenanthridinium dyes.
Bailly
Christian
C
INSERM U-524 et Laboratoire de Pharmacologie Antitumorale du Centre Oscar Lambret, IRCL, 59045 Lille, France. christian.bailly@panther.gsu.edu
Arafa
Reem K
RK
Tanious
Farial A
FA
Laine
William
W
Tardy
Christèle
C
Lansiaux
Amélie
A
Colson
Pierre
P
Boykin
David W
DW
Wilson
W David
WD
D001483:Base Sequence; D001665:Binding Sites; D002942:Circular Dichroism / Q000379:methods; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D018900:Dinucleotide Repeats; D004996:Ethidium / Q000138:chemical synthesis* / Q000378:metabolism; D019791:Guanidine / Q000031:analogs & derivatives / Q000138:chemical synthesis* / Q000378:metabolism; D007364:Intercalating Agents / Q000138:chemical synthesis* / Q000378:metabolism; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010617:Phenanthridines / Q000138:chemical synthesis* / Q000378:metabolism; D011061:Poly A / Q000378:metabolism; D011071:Poly T / Q000378:metabolism; D013053:Spectrophotometry; D020349:Surface Plasmon Resonance; D013816:Thermodynamics
D007364:Intercalating Agents; D010617:Phenanthridines; D011061:Poly A; D011071:Poly T; D004247:DNA; D004996:Ethidium; D019791:Guanidine