Mononucleosomes isolated from micrococcal nuclease digests of stationary phase chromatin of the yeast Saccharomyces cerevisiae were compared both compositionally and physiochemically with those from chicken and bovine calf. It was found that while yeast mononucleosomes are similar in composition, their thermal denaturation profiles and circular dichroism spectra indicate a less constrained structure. Furthermore, yeast nucleosomes were discovered to be labile in solutions of low ionic strength and could not be reconstituted by methods applicable to calf and chicken nucleosomes. On the basis of the reconstitution of a hybrid nucleosome containing calf histones H2A, H2B, and H3 and yeast histone H4, it was concluded that variations in the yeast H4 sequence are unlikely to be responsible for the apparent decrease in the stability of yeast nucleosomes. Examinations of histone-histone interactions in free solution revealed a change in the H3-H4 interaction and together with the previously published results of other researchers it was inferred that changes in the H3 sequence might be responsible for this structural variation.