Phosphorescence and optically detected magnetic resonance studies of echinomycin-DNA complexes

Abstract

Echinomycin complexes with polymeric DNAs and model duplex oligonucleotides have been studied by low-temperature phosphorescence and optical detection of triplet-state magnetic resonance (ODMR) spectroscopy, with the quinoxaline chromophores of the drug used as intrinsic probes. Although not optically resolved, plots of ODMR transition frequencies versus monitored wavelength revealed heterogeneity in the phosphorescence emission of echinomycin, which was ascribed to the presence of two distinct quinoxaline triplet-state environments (referred to as the blue and red triplet states of echinomycin in this report). We think that a likely origin of the two triplet states of echinomycin is the occurrence of two or more distinct conformations of the drug in aqueous solutions. Spectroscopically observed perturbations of the triplet-state properties of echinomycin such as the phosphorescence emission spectrum, phosphorescence lifetime, ODMR spectrum, and zero-field splitting (zfs) energies were investigated upon drug binding to the double-stranded alternating copolymers poly(dG-dC).poly(dG-dC) [abbreviated as poly[d(G-C)2]] and poly(dA-dT).poly(dA-dT) [abbreviated as poly[d(A-T)2]], the homopolymer duplexes poly(dG).poly(dC) [abbreviated as poly(dG.dC)] and poly(dA).poly(dT) [abbreviated as poly(dA.dT)], and the natural DNAs from Escherichia coli, Micrococcus lysodeikticus, and calf thymus. Echinomycin bisintercalation complexes with the self-complementary oligonucleotides d(ACGT), d(CGTACG), and d(ACGTACGT), which are thought to model drug binding sites, were also investigated. Phosphorescence and ODMR spectroscopic results indicate that the quinoxaline chromophores of the drug are involved in aromatic stacking interactions in complexes with the natural DNAs as evidenced by red shifts in the phosphorescence 0,0 band of the drug, a small but significant reduction in the phosphorescence lifetime of the red triplet state, and reduction of the zfs D-value of both the blue and red triplet states upon drug complexation. These changes in the triplet-state properties of echinomycin are consistent with stacking interactions that increase the polarizability of the quinoxaline environment. The extent of the reduction of the D parameter for the red triplet state upon complexation with the polymeric DNAs was found to correlate with the binding affinities measured for these targets [Wakelin, L. P. G., & Waring, M. J. (1976) Biochem. J. 157, 721-740], with the single exception of the drug-poly[d(G-C)2] complex, for which an increase in the D-value was noted. In addition, upon drug binding to the natural DNAs, there is a reversal of signal polarity in the ODMR spectra of the red triplet state. Among the synthetic DNA polymers investigated, a reversal of ODMR signal polarity was found only with the echinomycin-poly[d(A-T)2] complex.(ABSTRACT TRUNCATED AT 400 WORDS)

Conformational heterogeneity of quinoxaline peptides in solution

“…21 The origin of the conformational heterogeneity of triostin A in nonpolar solutions has not been determined unequivocally. Based on nmr studies, Kuwano et al 33 attributed the existence of two conformations of triostin A in nonpolar solutions to cis-trans isomerization of the N-methyl peptide bonds.…”

“…Low-temperature phosphorescence measurements were performed using previously described instrumentation. 21 B & J Brand high-purity acetonitrile, methanol, chloroform, and 2-propanol and HPLC-grade water from J. T. Baker Co. were employed for spectroscopic and chromatographic studies. CDC13, methanol-d4, and acetonitrile-d3 for nmr measurements, and 2-quinoxalinoyl chloride, methylamine, and diethylamine for synthesis of the corresponding quinoxaline-2-carboxamides, were obtained from Aldrich Chemical Co.…”

“…16 Recent nmr solution studies of e c h i n~m y c i n '~-~~ and [ N-MeCys3,N-MeCys7] -TANDEM ( MCTANDEM) 2o have focused primarily on the structure of the DNA complexes and changes in the oligonucleotide targets upon binding in aqueous solutions. A recent report on the lowtemperature phosphorescence and optically detected triplet-state magnetic resonance ( ODMR) spectroscopic investigations of echinomycin and its DNA complexes 21 has suggested that the heterogeneity observed in the phosphorescence and ODMR of the unbound peptide in aqueous environments may be ascribed to the presence of two distinct triplet states of the peptide, possibly the result of two or more forms of the drug which are frozen out at low temperatures.…”

“…Nuclear magnetic resonance studies22-24 have shown that triostin A exists in deuteriochloroform solutions as two symmetrical conformers separated by an energy barrier of [20][21][22] kcal/mol. Based upon solvent effects on conformer equilibria, Blake et a1."…”

Binding of Echinomycin to d(GCGC)2and d(CCGG)2: Distinct Stacking Interactions Dictate the Sequence-Dependent Formation of Hoogsteen Base Pairs

“…In fact, the stacking interactions arising from these chromophores have been thoughtto enhance the stability of the complexes (39) and have also been suggested to contribute to the sequence requirements for Hoogsteen base pair formation (9,11,18). For these reasons, and in order to understand the differences among the four complexes studied and to facilitate the A) with the CO of the adjacent qxn-6 (see Figure 4).…”