ROLE OF INDIVIDUAL HISTONE TYROSINES IN THE FORMATION OF THE NUCLEOSOME COMPLEX

We have determined the accessibility of histone tyrosine residues to react with p-nitrobenzenesulfonyl fluoride (NBSF) in intact nuclei, salt-dissociated nucleosomes, isolated histone complexes, and individual core histones. Of the 15 core histone tyrosine residues, 13 are inaccessible in native nucleosomes; only Tyr121 near the C-terminus of H2B is fully accessible, and Tyr54 of H3 is partially accessible under near-physiological conditions. When H1 and the basic N-terminal tails of the core histones are dissociated from the DNA by treating nuclei with 0.4 and 0.8 M NaCl, the two tyrosines which are adjacent to the basic regions of H2B and H3 become accessible as well. This indicates that these tyrosine residues may be involved in histone-DNA interactions, either directly or indirectly. When the H2A-H2B dimers are dissociated from the chromatin by raising the NaCl concentration to 1.2 M, three to four tyrosines located in the structured regions of H2B and H4 are exposed, suggesting that these tyrosine residues may be located at the dimer-tetramer interface. Dissociating all the histones from the DNA at an even higher ionic strength as a mixture of dimers, tetramers, and octamers does not change the pattern of Tyr exposure, but reduces the reactivity of the tyrosines at the dimer-tetramer interface as would be expected from the reassociation of H2A-H2B dimers and H3-H4 tetramers. The fact that our results with native nucleosomes in intact nuclei are in every aspect consistent with the crystallographic structure proposed for the DNA-free histone octamer of Arents et al. [(1991) Proc. Natl. Acad. Sci. U.S.A. 88, 10148-10152] indicates that the structure of the protein core complex is not affected by removal of the DNA at high ionic strength. The exposure to NBSF modification of histone tyrosines at the histone-DNA and the dimer-tetramer interface may be a useful indicator for nucleosome unfolding which has been postulated to occur during chromatin transcription and replication.