Spin-Labeled Psoralen Probes for the Study of DNA Dynamics

Abstract

Six nitroxide spin-labeled psoralen derivatives have been synthesized and evaluated as probes for structural and dynamic studies. Sequence specific photoaddition of these derivatives to DNA oligonucleotides resulted in site-specifically cross-linked and spin-labeled oligomers. Comparison of the general line shape features of the observed electron paramagnetic resonance (EPR) spectra of several duplexes ranging in size from 8 to 46 base pairs with simulated EPR spectra indicate that the nitroxide spin-label probe reports the global tumbling motion of the oligomers. While there is no apparent large amplitude motion of the psoralen other than the overall tumbling of the DNA on the time scales investigated, there are indications of bending and other residual motions. The (A)BC excinuclease DNA repair system detects structural or dynamic features of the DNA that distinguish between damaged and undamaged DNA and are independent of the intrinsic structure of the lesion. NMR studies have shown that psoralencross-linked DNA has altered backbone dynamics and conformational populations in the immediate vicinity Psoralen-damaged DNA serves as an excellent substrate for the study of structural and dynamic motifs that DNA repair enzyme systems may recognize. Psoralens are photoreagents which form a well-characterized set of covalent nucleic acid adducts through photochemical addition ( Figure 1) (Straub et al., 1981; Kanne et al., 1982a,b). The primary reaction is cyclobutane ring formation between the 5,6 double bond of thymidine in DNA and either the 4',5' or 3,4 double bonds of the psoralen. Reaction at the 4'3' double bond creates a furanside monoadduct (MAf),' which can react further at a site with opposed and adjacent pyrimidines to create an interstrand cross-link (XL). The XL is a rigid unit formed by the psoralen and the opposite and adjacent pyrimidine bases, linking the two strands together. Reaction at the 3,4 double bond of the psoralen first creates a pyroneside monoadduct (MAP), which cannot form an interstrand cross-link.Psoralen-DNA monoadducts and cross-links are recognized and removed by the excision repair system in both

Interaction of cyanine dyes with nucleic acids. XXI. Arguments for half‐intercalation model of interaction

“…10 In particular, such a mode of interaction was clearly confirmed by NMR spectroscopy on the cyanine thiazole orange homodimeric dye TOTO. 11,12 The determined structure of the TOTO-DNA complex was similar to Lerman's classical full-intercalation model. 13 Intercalative and groove binding interactions were both observed at a high dye/DNA ratio for pseudoisocyanine 14 and for the monomethine dye YO-PRO-1.…”

Site‐Directed Spin Labelling of Nucleic Acids

“…Early on, alkylating or acylating agents, such as compounds 13 45 and 14 46 (Figure 6, A), were used for spin labelling,47 but for the aforementioned reasons had very limited sequence selectivity. In an attempt to increase the selectivity, bifunctional cross‐linking agents, such as the hydrazine mustard spin label 16 ,48 the psoralene derivative 17 49 and the spin‐labelled cisplatin 18 ,50 were developed. Although these cross‐linking agents have a sequence selectivity that spans a few nucleotides, such as intra‐strand cross‐linking of the sequence GG for 18 , they still lack enough specificity to be generally applicable for SDSL.…”

Molecular Beacon Probes of Oligonucleotides Photodamaged by Psoralen

“…They have been used for the treatment of skin diseases such as psoriasis, vitiligo and other skin pigmentation diseases (1–3). Psoralens can intercalate between pyrimidine bases of double‐stranded nucleic acids, and upon UVA (320–400 nm) irradiation, the intercalated psoralen can photoreact with adjacent pyrimidine bases (4,5). Monoadducts form first, with a cyclobutane ring formed between the 5,6 double bond of thymine on one strand and either the 4′,5′ (furan) or 3,4 (pyrone) double bond of the psoralen.…”

“…Monoadducts form first, with a cyclobutane ring formed between the 5,6 double bond of thymine on one strand and either the 4′,5′ (furan) or 3,4 (pyrone) double bond of the psoralen. Then, upon absorbing a second photon, it can form diadducts, linking the two strands of the double helix (4,6). Only the furanside monoadduct can react further with a pyrimidine on the opposite strand to create an interstrand crosslink (7).…”