Figure 2

Z-DNA incorporation in a nucleosome. a. Body labeled PCR fragments described in Figure 1a, were assembled into nucleosomes by step salt dialysis using purified HeLa nucleosome cores. Nucleosome:DNA ratios were optimized to yield greater than 90% nucleosomal material as determined by native acrylamide gel separation of DNA from nucleosomes. All six constructs described assembled into nucleosomes at identical DNA:histone ratios. The positions of DNA and nucleosomes are indicated on the right side of the panel. b. Stability of the Z-DNA structure after assembly into a nucleosome was assayed by gel shift. Mononucleosomes assembled as in 2a, were incubated at room temperature for 30 minutes with purified Z-DNA specific binding protein Zaa. The binding reaction was run on a 4% native acrylamide gel and autoradiographed. The Z-DNA containing nucleosome assembly (Znuc) was bound by Zaa while the canonical nucleosome (Tnuc) and the GC-containing nucleosome (Bnuc) were not shifted when incubated with Zaa. The positions of shifted templates are indicated on the right side of the panel.