STABILIZATION OF PARALLEL (RECOMBINANT) TRIPLEX WITH PROPIDIUM IODIDE

Earlier we have shown that the oligonucleotide 5'-d(CATGCTAACT)-L-d(AG TTAGCATG)-L-d (CATGCTAACT)-3' [L = pO(CH2CH2O)(3)p] is able to fold ba ck forming intramolecular RecA-independent tripler with identical stra nds oriented parallel to each other (parallel tripler) [AK. Shchyolkin a, E.N. Timofeev, O.F. Borisova, I.A. Il'icheva, E.E. Minyat, E.V. Kho myakova, V.L. Florentiev, FEES Letters 339, 113-118 (1994) (1)]. In th is study the propidium iodide (PI) was found to intercalate into the p arallel tripler and increase its stability significantly (T-m increase d from 21.4up to 44.4 degrees C in 0.01 M Na phosphate buffer, pH 7, 0 .1 M NaCl when three PI molecules per tripler were bound). Fluorescenc e excitation and emission spectra, the quantum yield of fluorescence ( q = 0.16) and the fluorescence lifetime of PI(tau = 24.5 ns at 3 degre es C) for the parallel tripler studied were shown to be similar to tho se for DNA. Scatchard binding plots indicated an anticooperative mode of PI binding to the parallel tripler. The association constant is clo se to that of PI binding to DNA. The fluorescence experiments revealed the maximum number of binding sites to be five PI molecules per one t ripler molecule, Molecular mechanics calculation of possible structure s for the parallel tripler-PI complex were performed.