Schematic diagram of pKa calculations and electrostatic analyses of nucleosome structures. (a) Determination of pKa shifts in histone titratable residues induced by the protein environment in nucleosome or the nucleosome-partner protein interactions. (b) Calculation of the long-range electrostatic forces generated between nucleosomes and binding proteins at separating distances from 10 to 30 Å, with a step size of 2 Å, along the line connecting their centers of mass. (c) Calculation of the long-range electrostatic forces generated between nucleosomes and binding proteins at different rotational angles of the line connecting their centers of mass, from −30° to 30°, with a step size of 6°.

pKa shifts of histone titratable residues from histone dimer to octamer formation. (a) Comparison of our predicted pKa values with experimentally determined pKa values for nine titratable residues near the acidic patch. (b) Distribution of pKa shifts of histone titratable residues due to the protein environments in nucleosome. The pKa shift represents the difference in pKa values of histone titratable residues between the dimer and the octamer within the nucleosome structure. (c) and (d) Histone titratable residues exhibiting significant pKa shifts in nucleosome structures (|ΔpKa| ≥ 1.5). Histone residues belonging to different histone types were indicated by different colors.

Modulation of nucleosome surface electrostatic potentials by protonation states of histone ionizable residues. (a) Locations of identified histone titratable residues whose protonation states are sensitive to pH perturbations in cellular conditions. (b) Six histone titratable residues are located on the surface of the histone octamer, whose protonation states are sensitive to pH perturbations. (c) Nucleosome surface electrostatic potentials under different pH conditions: Left: under weakly acidic condition (pH 5-6.5); Middle: within the physiological pH range (pH 6.5-7.5); Right: under weakly basic condition (pH 7.5-9). The electrostatic potential is mapped onto the surface of histone octamer on a scale from +5 kT/e (blue) to −5 kT/e (red).

Protonation states of histone ionizable residues regulate the long-range electrostatic interactions between nucleosomes and regulatory proteins. (a) and (b) Comparison of the magnitudes of electrostatic forces between nucleosomes and binding proteins (Tumor suppressor p53-binding protein 1 on the left, and PCR1 on the right) under weakly acidic (pH 5 to 6.5), physiological (pH 6.5 to 7.5), and weakly basic (pH 7.5 to 9) conditions. The electrostatic forces were computed at various center-of-mass separation distances ranging from 10 to 30 Å, with 2 Å increments, using PDB structures 5KGF and 8GRM. (c) and (d) Comparison of the directions of electrostatic forces between nucleosomes and binding proteins (Tumor suppressor p53-binding protein 1 above, and PCR1 below) under weakly acidic (pH 5 to 6.5), physiological (pH 6.5 to 7.5), and weakly basic (pH 7.5 to 9) conditions. The directions of electrostatic interactions are represented by red arrows at separating distances of 10 Å, 20 Å, and 30 Å.

Effects of proton uptake or release of histone titratable residues on nucleosome-chromatin factor interactions. (a) Distribution of pKa shifts of histone titratable residues caused by the interactions between nucleosomes and binding proteins. The pKa shift represents the change in pKa value of the titratable group from the unbound state to the bound state. (b) and (c) Histone titratable residues that exhibited protonation state change upon binding of partner proteins to nucleosome. These residues were mapped onto the surface of nucleosome structure using the PDB 2PYO (shown as red). Protein H2A, H2B, H3 and H4 were shown as pink, yellow, green, and blue, respectively. (d) and (e) Comparison of the hydrogen bonds and salt bridges formed between nucleosomes and binding partner proteins under two different protonation states of histone titratable residues: in one set, histone residues were assigned protonation states corresponding to the unbound condition, while in the other set, histone residues were assigned protonation states reflecting the bound condition.

Impact of histone cancer mutations on nucleosome surface electrostatic potential and electrostatic interactions. (a) Classification of recurrent histone cancer mutations based on their impact on residue net charges. Among the charge-altering mutations, Type 1 represents a charged residue mutated to a neutral residue, Type 2 represents a neutral residue mutated to a charged residue, and Type 3 represents mutations that flip the net charge of residues. (b) Distribution of binding free energy changes (ΔΔGs) caused by recurrent histone cancer mutations in nucleosome-partner protein interactions for different mutation types. (c) Changes in the nucleosome surface electrostatic potential caused by histone cancer mutations: left, wild-type; right, mutant-type. The electrostatic potential is mapped onto the surface of the histone octamer on a scale from +5 kT/e (blue) to −5 kT/e (red). (d) and (e) Comparison of the magnitudes of electrostatic forces between the nucleosome and binding partners in wild-type and mutant states. The electrostatic forces were computed at various center-of-mass separating distances ranging from 10 to 30 Å, with 2 Å increments, using PDB structures 8GRM and 3TU4.

Protonation states of histone titratable residues regulate the long-range electrostatic interactions in higher-order nucleosome structures. (a) and (b) The strengths and directions of electrostatic forces between neighboring nucleosomes in two structures of nucleosome arrays: H1-bound 6-nucleosome array (top) and telomeric tetranucleosome structure (bottom). The electrostatic forces were computed at various center-of-mass separating distances using PDB structures 6HKT and 7V9K under weakly acidic (pH 5 to 6.5), physiological (pH 6.5 to 7.5), and weakly basic (pH 7.5 to 9) conditions.

Histone cancer mutations affect the electrostatic interactions between the H4 tail and acidic patch. (a) Analyses of the long-range electrostatic forces generated between nucleosomes and H4 tail at separating distances from 10 to 30 Å, along the line connecting their centers of mass. (b) Comparison of the magnitudes of electrostatic forces between nucleosome and histone H4 tail under weakly acidic (pH 5 to 6.5), physiological (pH 6.5 to 7.5), and weakly basic (pH 7.5 to 9) conditions. (c) and (d) Comparison of the magnitudes of electrostatic forces between the H4 tail and acidic patch in wild-type and mutant states. The electrostatic forces were computed at various center-of-mass separating distances ranging from 10 to 30 Å, with 2 Å increments. (e) and (f) Comparison of the directions of electrostatic forces between nucleosomes and H4 tails in wild-type and mutant states. The directions of electrostatic interactions are represented by red arrows at separating distances of 10 Å, 20 Å, and 30 Å.

A generalized model explaining pH perturbation and cancer mutation can affect nucleosomes’ interactions with chromatin factors and higher-order nucleosome structures by modulating the long-range electrostatic interactions. (a) Perturbations within the cellular pH ranges and charge-altering cancer mutations can affect the interactions between nucleosomes and chromatin factors. (b) Protonation states of histone ionizable residues can regulate nucleosome stacking in higher-order nucleosome structures. (c) Perturbations within the cellular pH ranges and charge-altering cancer mutations can affect the interactions of H4 tail with acidic patch in neighboring nucleosome.