Chemical mapping of H4S47C nucleosomes generally confirms the basic patterns of nucleosome positioning across the yeast genome inferred from nuclease-based assays and yields additional insights into rotational positioning of nucleosomes—the precise sequences for which the DNA major groove faces toward vs. away from the histone octamer. First, for nucleosomes which exhibit apparent “overlap”—where dyads are measured within ∼70 nt of one another—dyads are preferentially separated by multiples of 10 nt, indicating that the multiple nucleosome positions that contribute to “fuzzy” nucleosomes (see above) all exhibit a consistent shared rotational setting. Analysis of nucleosomal DNA sequences from chemical cleavage maps also reveals a much stronger signal for dinucleotide periodicity across the nucleosome than previously appreciated, emphasizing the role for DNA sequence flexibility/curvature in establishing the rotational positioning of nucleosomes. Finally, and most interestingly, analysis of dyad to dyad distances for adjacent (nonoverlapping) nucleosomes revealed a strong preference for linker DNA to be 10n + 5 nt long. This finding has implications for secondary structure of chromatin, as it suggests that adjacent nucleosomes will be positioned approximately on opposite sides of the DNA double helix, potentially consistent with two-start models for 30-nm fiber (Yao et al. 1993).
