STRUCTURAL SPECIFICITY EFFECTS OF TRIVALENT POLYAMINE ANALOGUES ON THE STABILIZATION AND CONFORMATIONAL PLASTICITY OF TRIPLEX DNA

摘要

Natural polyamines, i.e. putrescine, spermidine and spermine, are excellent promoters of tripler DNA. Using melting temperature (T-m) measurements and CD spectroscopy, we found that structural alterations on spermidine backbone, including methylation, or acetylation at the N-1-, N-4- and/or N-8-positions had a profound influence on the stability and conformation of poly(dA).2poly(dT) tripler. The conformation of the polynucleotide complex underwent sequential changes from B-DNA to tripler DNA as the concentration of spermidine increased from 0 to 50 mu M in a buffer containing 10 mM sodium cacodylate and 1 mM EDTA (pH 7.2). At 60 mu M spermidine, the CD spectrum of tripler DNA was comparable with that of Psi-DNA, with a strong positive band centred around 260 nm. A negative band was also found at 295 nm. At higher concentrations of spermidine, however, the intensity of the positive band progressively decreased and the peak intensity was found at a 1 :0.3 molar ratio of DNA phosphate:spermidine. Temperature-dependent CD analysis showed that the Psi-DNA structure melted to single-stranded DNA at temperatures above the T-m determined from the absorbance versus temperature profile, Comparable effects were exerted on the conformation of triplex DNA by Co(NH3)(6)(3+), an inorganic trivalent cation. Substitution of the N-4-hydrogen of spermidine by a cyclohexyl ring or the fusion of the N-4-nitrogen in a cyclic ring system, as in piperidine, enhanced the ability of spermidine analogues to stabilize tripler and Psi-DNA forms over a wider concentration range compared with spermidine. These data demonstrate a differential effect of trivalent cations in stabilizing tripler DNA and provoking unusual conformations such as Psi-DNA, Synthetic homologues of spermidine that stabilize tripler DNA over a wider range of concentrations than that stabilized by spermidine itself might have potential therapeutic applications in the development of an anti-gene strategy against several diseases, including cancer and AIDS.