Phased psoralen cross-links do not bend the DNA double helix.
Although the chemical reaction of psoralens with nucleic acids is well understood, the structure of psoralen-DNA cross-linked products is still not clear. Model building studies base on the crystal structure of the psoralen-thymine monoadduct suggest that each cross-link bends the DNA double helix by 46.5 degrees [Pearlman, D. A., Holbrook, S. R., Pirkle, D. H., & Kim, S.-H. (1985) Science (Washington, D.C.) 227, 1304-1308]. On the other hand, Sinden and Hagerman [Sinden, R. R., & Hagerman, P. J. (1984) Biochemistry 23, 6299-6303] find that, in solution, psoralen cross-linked DNA is not bent. Here we use gel electrophoresis to test the validity of the current models. We have synthesized a series of DNA fragments (21-24 base pairs in length), each containing one unique T-A site for 4'-(hydroxymethyl)-4,5',8-trimethylpsoralen (HMT) cross-linking. Because of an estimated 28 degrees unwinding of the helix by HMT [Wiesehahn, G., & Hearst, J. E. (1978) Proc. Natl. Acad. Sci. U.S.A. 75, 2703-2707], one expects that the 22-bp cross-linked fragment will be repeated nearly in phase with the average helical screw when multimerized. In that sequence ligation will maximally amplify any deformation to the double helix. We find that the ligated multimers of cross-linked DNA migrate close to the multimers of non-cross-linked DNA on polyacrylamide gels. Our observations place an upper limit of 10 degrees on DNA bending induced by psoralen cross-linking and indicate unwinding by about 1 bp, as well as stiffening of the double helix. These properties are not unexpected for classical intercalators.
Haran
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Department of Chemistry, Yale University, New Haven, Connecticut 06511.
Crothers
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D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000187:drug effects*; D004270:DNA, Circular / Q000187:drug effects; D011564:Furocoumarins / Q000494:pharmacology*; D066298:In Vitro Techniques; D007364:Intercalating Agents / Q000494:pharmacology; D009690:Nucleic Acid Conformation / Q000187:drug effects*; D014307:Trioxsalen / Q000031:analogs & derivatives / Q000494:pharmacology*
D003432:Cross-Linking Reagents; D004270:DNA, Circular; D011564:Furocoumarins; D007364:Intercalating Agents; C016507:hydroxymethyltrioxsalen; D004247:DNA; D014307:Trioxsalen