Psoralen crosslinking

Psoralen, or derivatives of 9-methoxy-7H-furo[3,2-g]chromen-7-one tricyclic ring structures, are used as photoreactive groups in crosslinkers, biotinylation compounds, and nucleic acid probes. Psoralens have been used for many years as photochemotherapy agents for treatment of psoriasis and vitiligo (Smith and Barker, 2006). Psoralens react when exposed to UV light... 

Psoralens can react by two different routes upon photoactivation (Parsons, 1980 Pathak, 1984). The first route is through the well-known photoreaction mechanism that principally involves intercalation within double-stranded DNA or RNA with the formation of adducts with adjacent thymine bases. The furan-side and pyrone-side rings in psoralen both can form cycloaddition products with the 5,6-double bond of thymine to create a crosslink between two DNA strands (Reaction 57) or to a lesser extent, within double-strand regions of RNA. 

Oxygen-free reactions of psoralens, when in close proximity to the target, proceed via the first excited states in which the 3,4-and the 4, 5 7r-bonds of the pyrone and furan moieties, respectively, can undergo C4-cyclization reactions with, e.g., unsaturated bonds of lipids, or the C5=C6 double bonds of thymine in DNA. In reactions with DNA the psoralen is believed to intercalate with DNA in the dark. Subsequent irradiation at 400 nm usually leads to furan-side 4, 5 -monoadduct formation, whereas irradiation at 350 nm increases the formation of crosslinks in which the furan and pyrone rings form C4 cycloadducts to thymines on opposite strands [95], Subsequent irradiation of the 4, 5 -monoadducts at 350 nm leads to formation of crosslinks and conversion into pyrone-side 3,4-monoadducts. Shorter wave-... 

Photochemically reactive molecules have often been used as labels for specific sites in proteins and nucleic acids. Psoralen derivatives serve as relatively nonspecific photochemically activated crosslinking agents for DNA and double-stranded RNA.195 Aryl azides are converted by light to aryl nitrenes, which react in a variety of ways including insertion into C-H bonds (Eq. 23-27).200 201 In some cases UV irradiation can be used to join natural substrates to enzymes or hormones to receptors. For example, progesterone, testosterone, and other steroids have been used for direct photoaffinity labeling of their receptors.202 Synthetic benzophenones have also been used widely as photoactivated probes.203... 

The last major group of reagents are the psoralens which have earlier been exploited to form crosslinks in double-stranded DNA and RNA (Song and Tapley, 1979 Parsons, 1980 Hearst, 1981). Reaction occurs primarily by 2 + 2 addition to pyrimidines but also with purines (Fig. 2.9). 

The demonstration of stepwise adduct formation by psoralens was an important advance. At 390 nm monoadduct formation occurs a crosslink is completed by irradiation at 360 nm (Chatterjee and Cantor, 1978). DNA-psoralen monoadducts have been used to covalently label complementary sites on large RNA molecules. The sites may then be mapped by electron microscopy of the denatured nucleic acid complexes (Wollenzien and Cantor, 1982). Psoralen derivatives for attachment to proteins have been made (Cantor, 1980) and this will allow the exploration of protein binding sites on nucleic acids by photochemical crosslinking. 

Action mechanism of ABC excision nuclease on a DNA substrate containing a psoralen crosslink at a defined position. Proc. Natl. 43. 

Psoralens (furocoumarins used by some plants in defence from insects and fungi) photoreact with ds DNA to form interstrand crosslinks in the presence of near-UV light by two steps intercalation in the dark and formation of monoadducts or interstrand crosslinks (Song and Tapley, 1979). This reaction has the advantage that psoralen binds, in contrast to photobiotin, only to nucleic acid. 

The crosslinking process depends on the structure of the psoralen derivative. Linear furocoumarins form crosslinks efficiently with a yield up to 50% of the overall adducts formed. 

Pearlman et al. (196) built a model of a psoralen-DNA covalently crosslinked complex based on the crystal structure of a thymidine-psoralen-thymine complex, using computer graphics and molecular mechanics calcu-... 

Van Houten, B., Gamper, H., Holbrook, S. R., Hearst, J. E., and Sancar, A. (1986). Action mechanism of ABC excision nuclease on a DNA substrate containing a psoralen crosslink at a defined position. Proc. NatL Acad. Sci. USA 83, 8077-8081. 

Methoxsalen and trioxsalen are derived from a family of benzofuran-containing plant natural products known as psoralens. Both methoxsalen and trioxsalen act as photosensitizers that decompose in the presence of UV light into active metabolites that crosslink DNA. These compounds have found significant use in the treatment psoriasis, eczema, and vitiligo. 

Sogo JM, Ness PJ, Widmer RM, Parish RW, Koller T (1984) Psoralen crosslinking of DNA as a probe for the structure of the active nucleolar chromatin. J Mol Biol 178 897-928... 

Ben-Hur E, Prager A, Riklis E (1979) Measurement of DNA crosslinks by SI nuclease induction and repair in psoralen-plus-360 nm light treated E. coli. Photochem Photobiol 29 921-924... 

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