Nucleic Acids Research, 2000, Vol. 28, No. 16 3092-3099
© 2000 Oxford University Press
DNase I digestion reveals alternating asymmetrical protection of the nucleosome by the higher order chromatin structure
Department of Respiratory Medicine and Allergy, GKT Medical School, King’s College London, Guy’s Hospital, London SE1 9RT, UK
DNase I was used to probe the higher order chromatin structure in whole nuclei. The digestion profiles obtained were the result of single-stranded cuts and were independent of pH, type of divalent ion and chromatin repeat length. Furthermore, the protection from digestion of the DNA at the entry/exit points on the nucleosome was found to be caused not by the H1/H5 histone tails, but by the compact structure that these proteins support. In order to resolve symmetry ambiguities, DNase I digestion fragments over several nucleosome repeat lengths were analysed quantitatively and compared with computer simulations using combinations of the experimentally obtained rate constants (some of which were converted to 0 to simulate steric protection from DNase I digestion). A clear picture of precisely defined, alternating, asymmetrically protected nucleosomes emerged. The linker DNA is inside the fibre, while the nucleosomes are positioned above and below a helical path and/or with alternating orientation towards the dyad axis. The dinucleosomal modulation of the digestion patterns comes from alternate protection of cutting sites inside the nucleosome and not from alternating exposure to the enzyme of the linker DNA.
* To whom correspondence should be addressed. Tel: +44 20 7955 4518; Fax: +44 20 7403 8640; Email: dontcho.staynov@kcl.ac.uk
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  M. Engelhardt Choreography for nucleosomes: the conformational freedom of the nucleosomal filament and its limitations Nucleic Acids Res., August 17, 2007; (2007) gkm560v1. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. J. J. Robinson, L. Fairall, V. A. T. Huynh, and D. Rhodes EM measurements define the dimensions of the "30-nm" chromatin fiber: Evidence for a compact, interdigitated structure PNAS, April 25, 2006; 103(17): 6506 - 6511. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. C. Bash, J. Yodh, Y. Lyubchenko, N. Woodbury, and D. Lohr Population Analysis of Subsaturated 172-12 Nucleosomal Arrays by Atomic Force Microscopy Detects Nonrandom Behavior That Is Favored by Histone Acetylation and Short Repeat Length J. Biol. Chem., December 14, 2001; 276(51): 48362 - 48370. [Abstract] [Full Text] [PDF]
|
|