title: Understanding nucleosomal histone and DNA interactions: a biophysical study.
abstract: Histones are associated with DNA to form nucleosome essential for chromatin structure and major nuclear processes like gene regulation and expression. Histones consist of H1, H2A, H2B and H3, H4 type proteins. In the present study, combined histones from calf thymus were complexed with ct DNA and their binding affinities were measured fluorimetrically. All the five histones were resolved on SDS page and their binding with DNA was visualized. The values of biding affinities varied with pH and salt concentration. Highest affinity (4.0 × 105M-1) was recorded at pH 6.5 in 50 mM phosphate buffer and 1.5 × 104M-1in 2 M NaCl at pH 7.0. The CD spectra support the highest binding affinity with maximum conformational changes at pH 7.0. The time-resolved fluorescence data recorded two life times for histone tyrosine residues at 300 nm emission in phosphate buffer pH 6.5. These life times did not show much change upon binding with DNA in buffer as well as in 2 M NaCl. The isothermal calorimetric studies yielded thermodynamic parameters ΔG, ΔH and ΔS as -1.6 × 105cal/mol, -1.13 × 103cal/mol and -3.80 cal/mol/deg, respectively, evidencing a spontaneous exothermic reaction. The dominant binding forces in building the nucleosome are electrostatic interactions.
authors.affiliation: a Department of Biotechnology , Maulana Abul Kalam Azad University of Technology , BF-142, Salt Lake, Sector-I, Kolkata 700 064 , West Bengal , India.
authors.forename: Chabita
authors.initials: C
authors.lastname: Saha
authors.affiliation: a Department of Biotechnology , Maulana Abul Kalam Azad University of Technology , BF-142, Salt Lake, Sector-I, Kolkata 700 064 , West Bengal , India.
authors.forename: Rahul
authors.initials: R
authors.lastname: Kumar
authors.affiliation: a Department of Biotechnology , Maulana Abul Kalam Azad University of Technology , BF-142, Salt Lake, Sector-I, Kolkata 700 064 , West Bengal , India.
authors.forename: Asmita
authors.initials: A
authors.lastname: Das
mesh_terms: D000818:Animals; D001703:Biophysics / Q000379:methods; D002417:Cattle; D002843:Chromatin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D009707:Nucleosomes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; D004247:DNA; C427203:calf thymus DNA
keywords: DNA; SDS page; circular dichroism; fluorescence; histone; pH; salt effect; tyrosine