Fig. 5

Functional characterization of Orc1-nucleosome interface. a–c Structural depiction of interactions between residues in Orc1 chosen for mutagenesis and nucleosomes (left panels), and the impact of these mutations on nucleosome binding measured by EMSA (right panels). a Orc1 E95 (orange) interacts with H18 of histone H4, and mutation of this residue to lysine (E95K) results in decreased affinity for the nucleosome as measured by EMSA. b Orc1 R202 makes a network of interactions that contribute to the stabilization of the LRS-interacting residues. The R202E mutation decreases the affinity of Orc1 for the nucleosome. c Acetylation of A2 at the N-terminus of Orc1 (yellow) helps to stabilize the conformation of Loop 3, and the L79I mutation (green) increases VdW interactions of the same Loop 3. A2P mutation, which prevents acetylation of the N-terminus of Orc1, results in decreased affinity for nucleosome. L79I has the opposite effect, resulting in an increased affinity of Orc1 for the nucleosome. Each data point and error bar represent the mean ± s.d. from three independent experiments. The standard errors of dissociation constants (K_D) are indicated