Supporting Information

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1 Supporting Information Nucleosome histone tail conformation and dynamics: impacts of lysine acetylation and a nearby minor groove benzo[a]pyrene-derived lesion Iwen Fu 1, Yuqin Cai 1, Nicholas E. Geacintov 2, Yingkai Zhang 2,3, and Suse Broyde 1* Departments of Biology 1 and Chemistry 2, New York University, 100 Washington Square East, New York, NY 10003, United States 3 NYU-ECNU Center for Computational Chemistry at NYU Shanghai, Shanghai , China *Corresponding Author: Suse Broyde, broyde@nyu.edu Tel. (212) S1

2 Table of Contents Methods... 5 Initial nucleosome models containing full length H2B tail at SHL ~ 3 for MD simulations... 5 Lesion-free/unacetylated NCP... 5 Lesion-free/acetylated NCP... 5 Lesion-containing/unacetylated NCP... 5 Lesion-containing/acetylated NCP... 6 Force Field... 6 Protonation... 6 Molecular Dynamics Simulation Protocols... 7 Structural Analyses... 7 Root-Mean-Square Deviations (RMSDs) 1D- and 2D- RMSDs... 7 Root-Mean-Square Fluctuations (RMSFs)... 8 Contact Surface Area (CSA)... 8 Hydrogen Bonds Analyses... 8 Radius of Gyration... 8 Secondary Structures of the Tail... 8 Tables... 9 Figures S2

3 List of Supporting Tables Table S1. Box sizes, number of waters, and length of MD simulations of the investigated NCP models with a full length H2B tail Table S2. Hydrogen bonds between DNA gyres (gyre-1 and gyre-2) and the tail residues 1 to 34 in the lesion-free/unacetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given Table S3. Hydrogen bonds between DNA gyres (gyre-1 and gyre-2) and the tail residues 1 to 34 in the lesion-free/acetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given Table S4. Hydrogen bonds between tail residue lysine 17 (LYS17) and DNA (including B[a]P-dG) in the lesion-containing/unacetylated NCP. Hydrogen bond occupancies (> 10%), average distances, and average angles are given. B[a]P-dG lesion is designated as G* Table S5. Hydrogen bonds between DNA and tail residues 1 to 34 in the lesion-containing/unacetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. B[a]P-dG lesion is designated as G* Table S6. Intra-tail hydrogen bonds involving residues 1 to 25 in the lesion-containing/unacetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. B[a]P-dG lesion is designated as G* Table S7. Hydrogen bonds between DNA and tail residues 1 to 34 in the lesion-containing/acetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. B[a]P-dG lesion is designated as G* Table S8. Hydrogen bonds between B[a]P aromatic rings and acetylated lysine residues (ACK) in the lesion-containing/acetylated NCP. Hydrogen bond occupancies (> 1%), average distances, and average angles are given. B[a]P-dG lesion is designated as G* List of Supporting Figures Figure S1. Structures of the two DNA gyres and the local DNA sequence near the H2B tail and lesion modification site, with G* designating the lesion modified base are shown Figure S2. The acetylated tail is more dynamic than the unacetylated tail and even more so when the lesion is present Figure S3. Histone H2B unacetylated tail snapshots along the 3 μs MD simulation of the lesionfree/unacetylated NCP show that the tail remains between the two gyres up to ~ 1 μs, then becomes transiently solvent exposed, and subsequently, at ~1.5 μs, the tail stably collapses onto one gyre (gyre-1 in blue) Figure S4. Contact maps between tail residues 1 to 28 and nearby DNA base pairs show that both the lesion and acetylation reduce the total contacts and alter the DNA-tail contact pattern Figure S5. Acetylation decreases the DNA surface area which is contacted by the tail lysine/acetylated lysine residues, and shortens the distances between acetylated lysine residues. (A) The contact surface area between DNA and the lysine/acetylated lysine residues in the lesion-free NCPs. (B) Average distances between lysine pairs and acetylated lysine pairs are given in the left and right panel, respectively S3

4 Figure S6. Both lysine acetylation and the DNA lesion affect the secondary structures of the H2B tail residues in the NCPs Figure S7. Both lysine acetylation and the lesion increase the mobility of nearby DNA Figure S8. The lesion enlarges the minor groove and makes it more dynamic. Acetylation causes the groove to open and fluctuate more Figure S9. Entrapment of the tail by the B[a]P ring system through a network of stabilizing interactions in the lesion-containing/unacetylated NCP Figure S10. Van der Waals interaction energies between the B[a]P ring system and individual amino acid residues of the H2B tail show stages for the interactions of tail residues ACK13, ACK20, ACK21, ACK25, and ARG26 with the B[a]P ring system. These interactions are unstable because of the lysine acetylation Figure S11. Van der Waals interaction energies between the B[a]P ring system and amino acid residues of the H2B tail show that the unacetylated tail interacts stably with the B[a]P ring system while lysine acetylation causes the contacts of the tail with the lesion to be unstable Figure S12. The acetylated tail oscillates between two conformations, one (state-1, 52%) in which it is in the lesion-containing minor groove, and the other (state-2, 38%) where it is housed between the two gyres. The remaining ~ 10% of the population represents transitional conformation between state-1 and state-2. The interactions between the B[a]P ring system and the tail are always unstable (Figure S11), causing the acetylated tail in both states to be dynamic; this is shown in the greater ensemble average RMSF values compared to the lesion-containing/unacetylated NCP (Figure 3, main text). The tail in state-1 is extended, collapsed onto the lesion-containing minor groove and has more contact with the DNA and the lesion s ring system; the tail in state-2 is housed between the two gyres, and is more compact and has relatively less contact with the DNA and the lesion s rings List of Supporting Movies Movie S1: Best representative structure of the lesion-free/unacetylated NCP Movie S2: Best representative structure of the lesion-free/acetylated NCP Movie S3: Entrapment of the tail by the lesion in the lesion-containing/unacetylated NCP Movie S4: Entrapment of the tail by the lesion in the lesion-containing/acetylated NCP S4

5 Methods Initial nucleosome models containing full length H2B tail at SHL ~ 3 for MD simulations Lesion-free/unacetylated NCP We built an initial model for our simulations which contains only one full length tail, the histone H2B tail. Our initial model is a hybrid NCP model. We began with a NCP with PDB 1 entry 2NZD 2 in which all the tails are truncated. We investigated the H2B tail at SHL ~ 3 in which residues 1 27 from the N terminus were truncated. We modeled in these residues based on the NCP with PDB 1 entry 1KX5 3 in which full length tails were resolved, and the H2B tail at SHL ~ 3 is between the two DNA gyres. Note that 1KX5 3 contains the same histones as 2NZD 2. In the crystal structure 1KX5 3, the H2B tail, in which the numbers are counted from the N-terminus of Chain D, is consists of residues 1 34 AKSAP APKKG SKKAV TKTQK KDKK RRKTR KESY (Figure 1, main text). Note that in our model for the H2B tail, we did not include the first three amino acids, PEP, of the H2B tail in Xenopus laevis because they are missing in this crystal structure PDB coordinate data. Therefore, the number of the first residue (Ala) in crystal structure 1KX5 3, that we employed, corresponds to the 4 th residue in Xenopus laevis. All other models for our simulations were based on this hybrid model (lesion-free/unacetylated NCP). Lesion-free/acetylated NCP For the acetylated tail NCP cases, we acetylated all the lysine residues on the H2B tail in the lesion-free and lesion-containing NCPs. However, for the lesion-free/acetylated NCP, after running ~ 2 μs of MD simulation, we found that the tail remains between the two gyres as in the crystal structure of PDB entry 1KX5 3, instead of becoming solvent exposed at ~ 1 μs as it did in the lesion-free NCP (Figure S3 in SI). We concluded that the tail remained trapped between the two gyres in a local minimum, since the acetylated tail should have been released from the DNA due to the charge neutralization. Accordingly, we created a second initial model of the lesion-free/acetylated NCP for MD simulation. For this model, we selected a snapshot at ~ 1 μs from the lesion-free/unacetylated NCP simulation, in which the tail had the greatest solvent accessible surface area (SASA), shown in Figure S3C. We then acetylated all the lysine residues on the H2B tail from this snapshot for subsequent MD simulation. Lesion-containing/unacetylated NCP To create a nucleosome model with the lesion, we modeled a minor groove-positioned 10S (+)-trans-anti- B[a]P-N 2 -dg (B[a]P-dG) lesion 4, based on the NMR solution structure, into the lesion-free/unacetylated NCP at SHL ~ 3 (Figure 1, main text) where the H2B tail is nearby and the lesion in the minor groove faces the tail (lesion-containing/unacetylated NCP). The lesion modification site G* corresponds to sequence number 29 of Chain I in crystal structure 2NZD 2, and the local DNA sequence for each gyre near the lesion is shown in Figure S1. We note that Figure S6 of our previous study 5 shows a comparison of minor groove widths in the absence and the presence of the H2B tail, revealing how the minor groove enlarges and becomes more dynamic when the tail is present. In addition, a comparison of the lesion- DNA linkage site torsion angle β', shown below, displays the accommodation of the β' torsion angle to the presence of the tail, compared to the truncated tail in the prior work; the accommodation occurs before 300 ns in the current simulation. This β' torsion angle governs the orientation of the benzo[a]pyrenyl ring system in the groove. S5

6 Lesion-containing/acetylated NCP We acetylated the H2B tail for the lesion-containing case (lesion-containing/acetylated NCP). Force Field AMBER force field ff14sb 6, 7, which includes force field ff99sb 8 and corrections 6 for the backbone and side chain torsions of the amino acids were used in all systems. We note that methyl-π interactions are well-represented in the AMBER force field. Free energy changes stemming from methyl-π interactions calculated with the AMBER force field and the thermodynamic integration method agree with experimental results 9. The partial charges and parameters for 10S (+)-trans-anti-b[a]p-n 2 -dg lesion were obtained from Mocquet et al 10. Lysine acetylation removes positive charge of the ε-amino group by adding the acetyl group to the nitrogen, which also increases the volume of the lysine side chain. Since the force field used here does not provide parameters for acetylated lysine, we utilized the force field parameters published by Papageorgiou 11 on the website ( for the acetylated lysine. We used the Joung-Cheatham 12 model for the K+ ions. For DNA, the force fields ff99 13 and modifications 14-16, including Barcelona bsc0 corrections 15 for α/γ backbone torsions and OL1 modification 16 for backbone angles epsilon (ε) and zeta (ζ) torsion were utilized. Details on the force field for nucleic acids is given here 17. Protonation Protonation states for the histones were presented in our earlier publication 18 for the same nucleosome, with details of the methods provided. S6

7 Molecular Dynamics Simulation Protocols Protonation of the histones and construction of molecular topology and coordinate files for the initial models were performed using the tleap module of AmberTools14 6. The nucleosome structures were neutralized by adding potassium counterions (K + ) and were placed in a periodic box of explicit TIP3P water 19. Potassium counterions were used (instead of sodium counterions, Na + ) because KCl was used in the buffer for the X-ray structure analysis and corresponds to the monovalent ion that has the highest concentration in the cell nucleus 20. The distance from the surface of the box to the closest atom of the solute is set to 10 Å. The water box size and water molecule numbers are given in Table S1. All systems were subjected to energy minimization, equilibration, and production dynamics using the PMEMD module of AMBER14 6. All simulations were carried out according to the following simulation protocol. First, the counterions and water molecules were minimized for 2500 steps of steepest descent and 2500 steps of conjugate gradient energy minimization, with a force constant of 50 kcal/mol/å 2 restraint on the solute atoms. Then, 30 ps initial MD at 10 K with 25 kcal/mol/å 2 restraints on solute were performed to allow the solute to relax. Next, the system was heated from 10 K to 300 K at constant volume for 30 ps with 10 kcal/mol restraints on the solute. Restraints on the solute were then relaxed with 30 ps of 10 kcal/(molå 2 ), 40 ps of 1 kcal/(molå 2 ), and 50 ps of 0.1 kcal/(molå 2 ) restraints. Subsequently, unrestrained dynamics was propagated in the NPT ensemble with a 2 fs timestep. Production MD was conducted at 1 atmosphere, 300 K. Constant pressure was maintained with a weak-coupling (Berendsen 21 ) barostat with a time constant of 1 ps. The simulation temperature was regulated by a Berendsen thermostat with a coupled thermostat of 4 ps. In all MD simulations, the SHAKE 22 algorithm for constraining the length of bonds to hydrogen was used. The short-range cutoff for nonbonded interactions was 9.0 Å, and long-range electrostatic interactions were treated with the particle-mesh Ewald method 23. The simulations were run for 3 ~ 3.5 μs and the trajectories were saved every 100 ps for further analysis. The simulations were run initially for equilibration using the CPU version of the PMEMD.MPI implementation of SANDER from AMBER 14 6, followed by production runs using the GPU version of the PMEMD.CUDA implementation of SANDER in AMBER 14 6 on NVIDIA Tesla K80 cards 24. Structural Analyses Post-processing of all simulations was carried out using the CPPTRAJ module 25 of AMBER14 6. All structural analyses were obtained from the ~ 3 μs MD simulations with the first 1.5 μs discarded. This was based on the 2D RMSDs (Figure S4), which show that stable tail structures were achieved after ~ 1.5 μs in the lesion-free/unacetylated NCP. In other NCP cases, the structures were revealed to fluctuate throughout the simulations compared to the lesion-free/unacetylated NCP. The best representative structure was obtained using cluster analysis, which was performed using the average linkage hierarchical agglomerative method 26 and RMSD as the distance metric. In this study, we are interested in the impact of the lesion and the acetylation on the dynamics and structure of the full length tail. For this purpose, we used the heavy atoms of the nearby 11-mer DNA from both gyres (see Figure S2) and the histone H2B tail residues 1 to 26 for clustering. Root-Mean-Square Deviations (RMSDs) 1D- and 2D- RMSDs The conventional one-dimensional RMSDs were computed by superimposing individual trajectory frames onto their respective first frame following equilibration for the selected region. We included the DNA 11- mer (Figure S2) that contains the lesion modification site and the histone H2B tail residues 1 26 for the S7

8 calculation of 1D RMSDs. The 2D RMSDs were computed between any two frames for the tail residues 1 26 (only considering the positions of Cα atoms). Root-Mean-Square Fluctuations (RMSFs) The measurement of root-mean-square fluctuation is an index of the average atomic mobility 27. Root mean square fluctuations of the heavy atoms during the MD simulations were computed using the CPPTRAJ module. All frames were superposed to the initial structure following equilibration prior to this calculation. Contact Surface Area (CSA) The contact surface area (CSA) is the surface area of the DNA covered by the tail and was computed as the difference between the solvent accessible surface areas (SASA) of the DNA with and without the nearby histone tail. For the SASA calculation, we calculated the surface area in Å 2 of all atoms using the LCPO algorithm of Weiser et al 28. Lower CSA values indicate dissociation of the tail from the DNA. Hydrogen Bonds Analyses Hydrogen bonds (HB) were defined as bonds between donor (D) and acceptor (A) atoms with a hydrogen (H) between them (D H A), where the distance between D and A was less than 3.5 Å and the angle of D H A was larger than 120 degree. Radius of Gyration Radius of gyration is an indicator of compactness of protein structure. Mathematically the radius of gyration is the average distance of the sum of the root mean square distance of each atom from its center of mass. We determined the radius of gyration (R g ) along each frame for all the studied models. The predicted gyration value for a globular protein of the same length (residues 1 to 26 of H2B tail) is ~ 7.6. This value is computed from R g,gggggggg = 2.2N 0.38, a relation based on a power law best fit of R g as a function of sequence length for a set of globular proteins in the PDB 29 database. Similarly, the predicted R g value for a thermally denatured random coil of the same length is ~ 15.5 ( R g,ddddddddd = 2.2N 0.6, with N=26, residues 1 to 26 of H2B tail), a relation proposed by Flory s theory 30, which is confirmed by power fitting R g values determined by computation and experiment 31. Secondary Structures of the Tail The secondary structure of the tail presented for each modeled system was determined with the AMBER14 secstruct tool 6, which uses the DSSP program to identify hydrogen bond motifs through backbone amide (N H) and carbonyl (C=O) group positions 32. By definition, a 3 10 helix spans at least three consecutive residues requiring two hydrogen bonds between residues (i, i+3), and an α-helix spans at last four consecutive residues requiring two hydrogen bonds between residues (i, i+4). Each residue is assigned a secondary structure type, which includes no structure, parallel β, antiparallel β, 3 10 helix, α helix, π helix, and turn for each frame. In this analysis, for each residue, we determined the percentage of simulation snapshots in which each residue assumes a 3 10 or α-helix, which we refer to as the helical propensity per residue. S8

9 Tables Table S1. Box sizes, number of waters, and length of MD simulations of the investigated NCP models with a full length H2B tail. DNA lesion state H2B tail acetylated state Lesion-free cases Lesion-containing cases Unacetylated tail Acetylated tail Unacetylated tail Acetylated tail model name Lesionfree/unacetylated NCP Lesionfree/acetylated NCP Lesioncontaining/unacetylated NCP Lesioncontaining/acetylated NCP Box Size, Å Water No. 34,345 34,342 34,309 34,306 Simulation Time (μs) S9

10 Table S2. Hydrogen bonds between DNA gyres (gyre-1 and gyre-2) and the tail residues 1 to 34 in the lesion-free/unacetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. Hydrogen bonds between gyre-1 and tail S10

11 Hydrogen bonds between gyre-2 and tail S11

12 Table S3. Hydrogen bonds between DNA gyres (gyre-1 and gyre-2) and the tail residues 1 to 34 in the lesion-free/acetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. Hydrogen bonds between gyre-1 and tail Hydrogen bonds between gyre-2 and tail S12

13 Table S4. Hydrogen bonds between tail residue lysine 17 (LYS17) and DNA (including B[a]P-dG) in the lesion-containing/unacetylated NCP. Hydrogen bond occupancies (> 10%), average distances, and average angles are given. B[a]P-dG lesion is designated as G*. S13

14 Table S5. Hydrogen bonds between DNA and tail residues 1 to 34 in the lesion-containing/unacetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. B[a]P-dG lesion is designated as G*. Hydrogen bonds between gyre-1 and tail Hydrogen bonds between gyre-2 and tail S14

15 Table S6. Intra-tail hydrogen bonds involving residues 1 to 25 in the lesion-containing/unacetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. B[a]P-dG lesion is designated as G*. S15

16 Table S7. Hydrogen bonds between DNA and tail residues 1 to 34 in the lesion-containing/acetylated NCP. Hydrogen bond occupancies (> 30%), average distances, and average angles are given. B[a]P-dG lesion is designated as G*. Hydrogen bonds between gyre-1 and tail Hydrogen bonds between gyre-2 and tail S16

17 Table S8. Hydrogen bonds between B[a]P aromatic rings and acetylated lysine residues (ACK) in the lesion-containing/acetylated NCP. Hydrogen bond occupancies (> 1%), average distances, and average angles are given. B[a]P-dG lesion is designated as G*. Note that the individual hydrogen bonds between the B[a]P ring system and the acetylated lysine residues (ACK) are not strong (the occupancy is less than 20 %). However, the total occupancies are not low: 91% between B[a]P ring system and the acetyl lysine residue 25 (ACK25); 37% between B[a]P ring system and the acetyl lysine residue 13 (ACK13). S17

18 Figures Figure S1. Structures of the two DNA gyres and the local DNA sequence near the H2B tail and lesion modification site, with G* designating the lesion modified base are shown. (A) A view of the best representative structure shown in Figure 1A in the main text, rotated 90 degrees clockwise along the dyad axis, showing the two DNA gyres. Gyre-1 and gyre-2 correspond to SHL > 0 and SHL < 0, respectively. (B) The numbering, chain ID, and super-helical location (SHL) for the DNA sequence from the crystal structure with PDB 1 entry 2NZD 2. Note that the base pair, T: 25 A: 25, which is 25 base pairs away from the dyad axis and at the positively numbered SHL of 2.5 is assigned as bp25. Base pair C: 26 G: 26 as bp26 and so on for gyre-1. For gyre-2, the base pair, G: 50 C: 50, which is 50 base pairs away from the dyad axis and at the negatively numbered SHL of 5 is assigned as bp 50. Base pair A: 53 T: 53 is bp 53 and so on. S18

19 Figure S2. The acetylated tail is more dynamic than the unacetylated tail and even more so when the lesion is present. 2D-RMSD values of Cα atoms in tail residues 1 to 26 show that the acetylated tail explores more conformational space than the unacetylated tail. The 2D-RMSD of each structure is computed pairwise against every other structure. The lower pairwise RMSD values indicate greater similarity between a pair of frames. S19

20 Figure S3. Histone H2B unacetylated tail snapshots along the 3 μs MD simulation of the lesionfree/unacetylated NCP show that the tail remains between the two gyres up to ~ 1 μs, then becomes transiently solvent exposed, and subsequently, at ~ 1.5 μs, the tail stably collapses onto one gyre (gyre-1 in blue). (A) A view of the structure shows the tail protruding from the histone core between the two DNA gyres. The histone core is not shown for clarity. (B) Same structures as in (A), rotated 90 degrees counter- S20

21 clockwise along the dyad axis, emphasizing the exposure of the tail to solvent. Gyre-1 and gyre-2 correspond to SHL > 0 (blue) and SHL < 0 (grey), respectively. The lesion modification site is rendered as stick and colored by atom with carbons in green; the H2B tail residues 1 to 29 are in cartoon and colored in red; the Cα atom of the first residue from the N terminus is in black sphere. (C) Solvent accessible surface area (SASA) of the histone H2B tail during the course of the MD simulation, with high SASA values beyond about 1 μs. S21

22 Figure S4. Contact maps between tail residues 1 to 28 and nearby DNA base pairs show that both the lesion and acetylation reduce the total contacts and alter the DNA-tail contact pattern. Contact maps show how lysine acetylation and a lesion affect which DNA sites the tail residues bind. The total number of contacts between one residue from the tail and one DNA base pair is summed in the color bar. A contact is defined as a distance of less than 3.9 Å between individual amino acid residue heavy atoms and DNA base pair heavy atoms. See Figure S1 for identification of base pairs. S22

23 Figure S5. Acetylation decreases the DNA surface area which is contacted by the tail lysine/acetylated lysine residues, and shortens the distances between acetylated lysine residues. (A) The contact surface area between DNA and the lysine/acetylated lysine residues in the lesion-free NCPs. (B) Average distances between lysine pairs and acetylated lysine pairs are given in the left and right panel, respectively. (A) The DNA region contacting the lysine/acetylated lysine residues is highlighted as surface rendering in cyan; the tail backbone is shown in cartoon rendering in red; lysine/acetylated lysine residue side-chains are shown as sticks in blue; the Cα atom of the first residue is displayed as black sphere and the ε-amino atoms of lysine and acetyl lysine residues are shown as yellow spheres. (B) The average distances are measured between ε-amino atoms of all lysine and acetylated lysine residues. Acetylation shortens the average distance between all pair-wise acetylated lysine residue side-chains by reducing repulsive interactions upon acetylation. S23

24 Figure S6. Both lysine acetylation and the DNA lesion affect the secondary structures of the H2B tail residues in the NCPs. (A) The unacetylated tail contains helical conformations (Ala14, Val15, Thr16, Lys17) locally but they are flickering, fluctuating between helical and turn conformations. Note that we classify as helical structures those containing α, 3 10, and π helices. (B) When the tail is acetylated, it forms a stable β-hairpin loop conformation (residues 12 to 21) with flickering helical structures between these residues, and the tail is more compact. (C) When the lesion is nearby, the unacetylated tail forms a stable β-hairpin loop conformation encompassing residues 15 to 24; these are housed in the lesion-imposed enlarged minor groove (Figure 3A, in main text), and residues 16 to 19 and 26 are engulfed by the lesion ring system. The part of the tail not engaged with the lesion contains flickering helical conformations at residues 8 to 12. (D) For the acetylated tail, the lesion disrupts ordered secondary structures including helical and β conformations. Types of secondary structures are colored as indicated in the color bar. Note that the secondary structural analyses in the main text have the first 1.5 μs discarded. S24

25 Figure S7. Both lysine acetylation and the lesion increase the mobility of nearby DNA. Ensemble average values of RMSFs of the nucleosomal DNA in the investigated NCPs. The DNA numbering and corresponding super-helical locations (SHL) are given along the X-axis. The dyad is at SHL = 0. Gyre-1 and gyre-2 correspond to SHL > 0 and SHL < 0, respectively. See Figure S1 for DNA numbering, SHL and gyre identification. The lesion-modification site is colored grey. Note that the investigated NCP contains only one histone tail at SHL ~ 3, the H2B tail between the two DNA gyres. Therefore, we are only interested in the DNA dynamics near this tail as highlighted in pink. S25

26 Figure S8. The lesion enlarges the minor groove and makes it more dynamic. Acetylation causes the groove to open and fluctuate more. Minor groove widths are the distance between backbone phosphates P(29) in Chain I and P( 25) in Chain J, P(30) in Chain I and P( 26) in Chain J, etc., minus 5.8 Å to account for the van der Waals radii of the P atoms 33. The ensemble average values of minor groove widths with block average standard deviations are given 34, 35 for each NCP model. The maximum values of the groove widths with the lesion present are highlighted in green. S26

27 Figure S9. Entrapment of the tail by the B[a]P ring system through a network of stabilizing interactions in the lesion-containing/unacetylated NCP. (A) The van der Waals interaction energies between the B[a]P ring system and residues and 26 of the tail show that these interactions are stable after 0.3 μs during the 3 μs course of the MD simulation. (B) Lysine residue 17 (LYS17) interacts with the B[a]P ring system through its backbone atoms and interacts with the nearby DNA backbone through its side chain atoms (Table S3). Interactions include conventional hydrogen bonds and carbon-oxygen hydrogen bonds 36. (C) Methyl-π (Me-π) interactions 37 S27

28 between the threonine residue 16 (THR16) methyl group and the B[a]P aromatic rings. (D) Intra-tail hydrogen bonds, specifically between ALA14 and LYS24 and between THR16 and LYS24 (Table S6), contribute to the stable loop conformation, which is housed in the lesion-imposed enlarged minor groove (Figure S8A). Note that all the hydrogen bonds are shown with black dashed line; the B[a]P ring system is shown as sticks and colored by atom with carbons green; certain key residues of the tail are highlighted as sticks colored by atom with carbons in light-blue. (E) The stability of the tail loop conformation, including residues 14 to 26, is revealed in the 1-D RMSD (left) and the 2-D RMSD (right). In the 2-D RMSD, the RMSD of each structure in the simulation is computed pairwise against every other structure. Lower pairwise RMSD values (darker blue) indicate greater similarity between a pair of structures. S28

29 Figure S10. Van der Waals interaction energies between the B[a]P ring system and individual amino acid residues of the H2B tail show stages for the interactions of tail residues ACK13, ACK20, ACK21, ACK25, and ARG26 with the B[a]P ring system. These interactions are unstable because of the lysine acetylation. The B[a]P ring system is rendered as sphere and colored by atom with carbons in green; the H2B tail residues are rendered as tube in red; tail residues ACK13, ACK20, ACK21, ACK25, and ARG26 are rendered as sticks and colored by atom with carbons in blue. (A) The snapshot is at 990 ns for stage 1, (B) at 1550 ns for state 2, and (C) at 2700 ns for stage 3. S29

30 Figure S11. Van der Waals interaction energies between the B[a]P ring system and amino acid residues of the H2B tail show that the unacetylated tail interacts stably with the B[a]P ring system while lysine acetylation causes the contacts of the tail with the lesion to be unstable. (A) Total van der Waals interactions between the B[a]P ring system and amino acid residues of the H2B tail during the course of the MD simulation (left) and the van der Waals energy distributions (right). S30

31 Interactions for the acetylated case are much more unstable than for the unacetylated case. Mean and standard deviations of van der Waals interaction energies are: lesion-containing/unacetylated NCP, 17.6 (±1.3); lesion-containing/acetylated NCP, 17.2 (±3.7) kcal/mol. (B) and (C) show the contributions of the residue backbone atoms and side chain atoms to the van der Waals interactions with the B[a]P ring system. The majority of the unacetylated tail residues that contribute to the van der Waals interactions with the B[a]P aromatic rings are from the backbone atoms; however, when the tail is acetylated, the tail residue side chain atoms contact the B[a]P aromatic rings, with their backbone atoms directed away from the rings. Note that the van der Waals analyses in the main text have the first 1.5 μs discarded. S31

32 Figure S12. The acetylated tail oscillates between two conformations, one (state-1, 52%) in which it is in the lesion-containing minor groove, and the other (state-2, 38%) where it is housed between the two gyres. The remaining ~ 10% of the population represents transitional conformation between state-1 and state-2. The interactions between the B[a]P ring system and the tail are always unstable (Figure S11), causing the acetylated tail in both states to be dynamic; this is shown in the greater ensemble average RMSF values compared to the lesion-containing/unacetylated NCP (Figure 3, main text). The tail in state-1 is extended, collapsed onto the lesion-containing minor groove and has more contact with the DNA and the lesion s ring system; the tail in state-2 is housed between the two gyres, and is more compact and has relatively less contact with the DNA and the lesion s rings. (A) Average RMSF values of the tail in the two states; (B) the population of van der Waals interaction energies between the B[a]P ring system and the tail. Mean and standard deviations are: state-1, 19.4 (±3.0); state-2, 14.2 (±2.4) kcal/mol. (C) The population of the contact surface area (CSA) between DNA and the histone H2B tail. Mean and standard deviations are: state-1, 957 (±94) Å 2 ; state-2, 829 (±75) Å 2. Structures shown, designated by an asterisk(*) on the CSA, are representatives of the highest population cluster; (D) the population of radii of gyration (R g ) of the tail showing extended and compact states of the tail. Mean and standard deviations are: state-1, 12.2 (±0.9); state-2, 8.2 (±0.4) Å. S32

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