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. 1984 Oct 15;223(2):519–526. doi: 10.1042/bj2230519

Photochemical cross-linking of protein and DNA in chromatin. Synthesis and application of a photosensitive cleavable derivative of 9-aminoacridine with two photoprobes connected through a disulphide-containing linker.

P E Nielsen, J B Hansen, O Buchardt
PMCID: PMC1144326  PMID: 6497860

Abstract

A novel cleavable photo-cross-linking reagent, N-(2-methoxy-6-azidoacridin-9-yl)-N'-(4-azidobenzoyl)cystamine, for analysis of protein-nucleic acid interactions, has been synthesized. The reagent contains two photosensitive groups that can be activated sequentially. The azidoacridinyl moiety is sensitive to u.v. and visible light (lambda less than or equal to 450 nm), whereas the azidobenzoyl part needs higher-energy light (lambda less than or equal to 350 nm). Furthermore, the disulphide bridge connecting the two photoactive groups can be cleaved by reduction with mercaptans. The reagent is shown to induce cleavable cross-links between all five major histones and DNA in chromatin from Ehrlich ascites cells on activation with long-wavelength u.v. light (lambda greater than 300 nm) at an efficiency of approximately 3% of the added reagent.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bloom K. S., Anderson J. N. Fractionation and characterization of chromosomal proteins by the hydroxyapatite dissociation method. J Biol Chem. 1978 Jun 25;253(12):4446–4450. [PubMed] [Google Scholar]
  2. Cao T. M., Sung M. T. Ultraviolet light induced preferential cross-linking of histone H3 to deoxyribonucleic acid in chromatin and nuclei of chicken erythrocytes. Biochemistry. 1982 Jul 6;21(14):3419–3427. doi: 10.1021/bi00257a027. [DOI] [PubMed] [Google Scholar]
  3. Fink G., Fasold H., Rommel W., Brimacombe R. Reagents suitable for the crosslinking of nucleic acids to proteins. Anal Biochem. 1980 Nov 1;108(2):394–401. doi: 10.1016/0003-2697(80)90604-1. [DOI] [PubMed] [Google Scholar]
  4. Georghiou S. Interaction of acridine drugs with DNA and nucleotides. Photochem Photobiol. 1977 Jul;26(1):59–68. doi: 10.1111/j.1751-1097.1977.tb07450.x. [DOI] [PubMed] [Google Scholar]
  5. Hansen J. B., Koch T., Buchardt O., Nielsen P. E., Wirth M., Nordén B. Acridine-psoralen amines and their interaction with deoxyribonucleic acid. Biochemistry. 1983 Oct 11;22(21):4878–4886. doi: 10.1021/bi00290a003. [DOI] [PubMed] [Google Scholar]
  6. Lerner M. R., Steitz J. A. Antibodies to small nuclear RNAs complexed with proteins are produced by patients with systemic lupus erythematosus. Proc Natl Acad Sci U S A. 1979 Nov;76(11):5495–5499. doi: 10.1073/pnas.76.11.5495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Maassen J. A. Cross-linking of ribosomal proteins by 4-(6-formyl-3-3-azidophenoxy)butyrimidate, a heterobifunctional, cleavable cross-linker. Biochemistry. 1979 Apr 3;18(7):1288–1292. doi: 10.1021/bi00574a026. [DOI] [PubMed] [Google Scholar]
  8. Mandel R., Kolomijtseva G., Brahms J. G. DNA-protein interactions in nucleosomes and in chromatin. Structural studies of chromatin stabilized by ultraviolet-light induced crosslinking. Eur J Biochem. 1979 May 15;96(2):257–265. doi: 10.1111/j.1432-1033.1979.tb13036.x. [DOI] [PubMed] [Google Scholar]
  9. Nielsen P. E. Photoaffinity labeling of chromatin. Synthesis and properties of arylazido derivatives of 9-aminoacridine: potential photolabels for chromatin studies. Eur J Biochem. 1982 Feb;122(2):283–289. doi: 10.1111/j.1432-1033.1982.tb05878.x. [DOI] [PubMed] [Google Scholar]
  10. Nielsen P. E. Photoaffinity labeling of chromatin: nuclease-sensitive chromatin shows preferential labeling of histone H1. FEBS Lett. 1981 Nov 30;135(1):173–176. doi: 10.1016/0014-5793(81)80970-2. [DOI] [PubMed] [Google Scholar]
  11. Sato K., Hosokawa K. The structure of adenovirion chromatin revealed by ultraviolet light-induced cross-linking. Biochem Biophys Res Commun. 1981 Aug 31;101(4):1318–1323. doi: 10.1016/0006-291x(81)91591-6. [DOI] [PubMed] [Google Scholar]
  12. Thomas J. O., Kornberg R. D. The study of histone--histone associations by chemical cross-linking. Methods Cell Biol. 1978;18:429–440. [PubMed] [Google Scholar]
  13. Vanin E. F., Ji T. H. Synthesis and application of cleavable photoactivable heterobifunctional reagents. Biochemistry. 1981 Nov 24;20(24):6754–6760. doi: 10.1021/bi00527a003. [DOI] [PubMed] [Google Scholar]

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