Divalent Metal- and High Mobility Group N Protein-Dependent Nucleosome Stability and Conformation
Figure 5
Off-centered NCP undergoes a transition in the presence of divalent metal and associates with HMGNs to form an S2-like state. (a) Crystal structure-based [19] models for off-centered and centered NCP. DNA strands are colored orange and cyan and histone proteins are grey. Off-centered NCP is missing an interaction between one DNA terminus and the N-terminal tail of H3 (MIA). (b) EMSAs of HMGN binding to centered (C; right) versus off-centered (O; left) NCP146b. Gel labels indicate HMGN identity (N1t, HMGN1t; N1, HMGN1; N2, HMGN2; n, NCP alone), and numbers designate HMGN:nucleosome molar stoichiometry. Off-centered samples (left 7 lanes) also contain a fraction of faster migrating centered NCP. Gels were run under three different buffer conditions: low ionic strength (top), near physiological ionic strength (middle), and near physiological ionic strength with 1 mM Mg2+ (bottom). HMGN : nucleosome molar stoichiometry (*) for centered samples (right 3 lanes) is 1 : 1 (top, middle) or 3 : 1 (bottom). Assignments of 1 : 1 and 2 : 1 HMGN : off-centered NCP species are inferred by comparison with those for centered NCP (see text for details). The additional band observed at low HMGN stoichiometry with Mg2+ (arrow) may coincide with a minor fraction of off-centered NCP in a compact state associated with one molecule of HMGN.
