TY - JOUR
T1 - Accessibility of nucleic acid-complexed biomolecules to hydroxyl radicals correlates with their conformation
T2 - A fluorescence polarization spectroscopy study
AU - Makrigiorgos, G. M.
AU - Bump, E.
AU - Huang, C.
AU - Baranowska-Kortylewicz, J.
AU - Kassis, A. I.
N1 - Funding Information:
Support for this study was provided by USPHS CA 63334-01 (G.M.M.)and by USPHS CA 15523-20 (A .I .K .) .
PY - 1994
Y1 - 1994
N2 - A fluorescence methodology has been developed to examine the relationship between the conformational state of specific biomolecules in simple chromatin models and their accessibility to hydroxyl radicals (.OH). Polylysine and histone H1 were labelled with SECCA, the succinimidyl ester of coumarin-3-carboxylic acid, which generates the fluorescent derivative 7-OH-SECCA following its interaction with radiation-induced .OH in aqueous solution. The fluorescence induced per unit γray dose reflecting the accessibility of .OH to such SECCA-conjugated biomolecules was recorded. The biomolecules were also labelled with the fluorescent derivative 7-OH-SECCA in trace amounts to study their conformation under identical conditions via fluorescence polarization spectroscopy. When these biomolecules were complexed with a polynucleotide or DNA, a major increase in polarization anisotropy was recorded. Upon salt-induced dissociation of these biomolecules from the nucleic acids, the increase in anisotropy was reversed. The histone H1-DNA complexes also exhibited an initial increase in anisotropy with increasing NaCl concentration (maximum at 100 mmol dm-3) indicating the possible formation of a more compact conformation. The fluctuations in anisotropy were inversely proportional to the recorded fluorescence/Gy. The data indicate a direct correlation between the accessibility of .OH to polylysine or histone H1 complexed with nucleic acids and the conformation of these biomolecules.
AB - A fluorescence methodology has been developed to examine the relationship between the conformational state of specific biomolecules in simple chromatin models and their accessibility to hydroxyl radicals (.OH). Polylysine and histone H1 were labelled with SECCA, the succinimidyl ester of coumarin-3-carboxylic acid, which generates the fluorescent derivative 7-OH-SECCA following its interaction with radiation-induced .OH in aqueous solution. The fluorescence induced per unit γray dose reflecting the accessibility of .OH to such SECCA-conjugated biomolecules was recorded. The biomolecules were also labelled with the fluorescent derivative 7-OH-SECCA in trace amounts to study their conformation under identical conditions via fluorescence polarization spectroscopy. When these biomolecules were complexed with a polynucleotide or DNA, a major increase in polarization anisotropy was recorded. Upon salt-induced dissociation of these biomolecules from the nucleic acids, the increase in anisotropy was reversed. The histone H1-DNA complexes also exhibited an initial increase in anisotropy with increasing NaCl concentration (maximum at 100 mmol dm-3) indicating the possible formation of a more compact conformation. The fluctuations in anisotropy were inversely proportional to the recorded fluorescence/Gy. The data indicate a direct correlation between the accessibility of .OH to polylysine or histone H1 complexed with nucleic acids and the conformation of these biomolecules.
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U2 - 10.1080/09553009414551181
DO - 10.1080/09553009414551181
M3 - Article
C2 - 7930827
AN - SCOPUS:0027930039
SN - 0955-3002
VL - 66
SP - 247
EP - 257
JO - International Journal of Radiation Biology
JF - International Journal of Radiation Biology
IS - 3
ER -