Whereas the DNA path inside the nucleosome is well established, it is essentially unknown in the ''entry-exit'' region, a missing piece in our understanding of the chromatin fiber's folding. The three-dimensional structure of ''linker'' DNA was investigated here on single nucleosomes reconstituted without H1 on a 256-bp DNA fragment. Cryoelectron microscopy and three-dimensional reconstruction of the DNA axis reveal these nucleosomes as 1.61 +/- 0.15 left-handed superhelical turn particles whose DNA arms bend away from the core particle, thus preventing the occurrence of a crossing in the entry-exit region. (C) 1995 Academic Press, Inc.