Visible light induced DNA-protein crosslinking in DNA-histone complex and sarcoma-180 chromatin in the presence of methylene blue

Formation of DNA-protein cross-links by the action of visible light in the presence of methylene blue was studied in calf thymus DNA-calf thymus histone complex and sarcoma-180 chromatin. The extent of cross-link formation decreases with a decrease in the histone to DNA ratio in the DNA-histone complex. In chromatin, it is at a maximum (93%) at a dye to DNA nucleotide ratio (D/P ratio) of 0.04 and is appreciable even at a very low dye concentration (75% at a D/P ratio of 0.0033). Sepharose 4B-CL column chromatography indicates that methylene blue acts as a mediator in the cross-linking process, but not as a linker in the DNA-protein cross-link. Dodecylsulphate-polyacrylamide gel electrophoresis patterns reveal that both histone and non-histone proteins are involved in cross-linking, but to a varied extent. Competition experiments with ethidium bromide demonstrated the necessity of intercalative binding of methylene blue in the formation of DNA--protein cross-links. Viscometric studies in 2 M NaCl indicate that the compact structure of chromatin is stabilized by cross-linking.