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J. Biol. Chem., Vol. 267, Issue 21, 14622-14628, 07, 1992
GG Lindsey and P Thompson
The assembly of hybrid core particles onto long chicken DNA with histone
H2B in the chicken histone octamer replaced with either wheat histone
H2B(2) or sea urchin sperm histone H2B(1) or H2B(2) is described. All these
histone H2B variants have N-terminal extensions of between 18 and 20 amino
acids, although only those from sea urchin sperm have S(T)PXX motifs
present. Whereas chicken histone octamers protected 167 base pairs (bp)
(representing two full turns) of DNA against micrococcal nuclease digestion
(Lindsey, G. G., Orgeig, S., Thompson, P., Davies, N., and Maeder, D. L.
(1991) J. Mol. Biol. 218, 805-813), all the hybrid histone octamers
protected an additional 17-bp DNA against nuclease digestion. This
protection was more marked in the case of hybrid octamers containing sea
urchin sperm histone H2B variants and similar to that described previously
(Lindsey, G. G., Orgeig, S., Thompson, P., Davies, N., and Maeder, D. L.
(1991) J. Mol. Biol. 218, 805-813) for hybrid histone octamers containing
wheat histone H2A variants all of which also have S(T)PXX motifs present.
Continued micrococcal nuclease digestion reduced the length of DNA
associated with the core particle via 172-, 162-, and 152-bp intermediates
until the 146-bp core particle was obtained. These DNA lengths were
approximately 5 bp or half a helical turn longer than those reported
previously for stripped chicken chromatin and for core particles containing
histone octamers reconstituted using "normal" length histone H2B variants.
This protection pattern was also found in stripped sea urchin sperm
chromatin, demonstrating that the assembly/digestion methodology reflects
the in vivo situation. The interaction between the N-terminal histone H2B
extension and DNA of the "linker" region was confirmed by demonstrating
that stripped sea urchin sperm chromatin precipitated between 120 and 500
mM NaCl in a manner analogous to unstripped chromatin whereas stripped
chicken chromatin did not. Tryptic digestion to remove all the histone
tails abolished this precipitation as well as the protection of DNA outside
of the 167- bp core particle against nuclease digestion.
S(T)PXX motifs promote the interaction between the extended N-terminal tails of histone H2B with "linker" DNA
Department of Biochemistry, University of Cape Town, Rondebosch, Republic of South Africa.
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