Analysis of variants of PRMT7 (Manuscript in preparation)
For specific parts of analysis see minimisation notes, M81del notes and scoring notes.
Briefly, a Swissmodel model based upon PDB:4C4A, the mouse PRMT7 (68% identity), was used for scoring of
variants in Pyrosetta.
It was energy minimised with the FastRelax mover,
a parameterised SAH ligand (rdkit_to_params module)
and with the zinc kept in place thanks to SetupMetalsMover.
Helper scripts from anothe repo of MF were used.
The score function used was ref2015, the beta_nov2016_cart was tried, but proved problematic with the coordinated zinc.
The ∆∆G (in kcal/mol) of the mutants are:
| model | mutation | complex_ddG | complex_native_dG | complex_mutant_dG | FA_RMSD | CA_RMSD | wt_rmsd | mut_rmsd | ratio_rmsd | wt_fa_atr | wt_fa_rep | wt_fa_sol | wt_fa_intra_rep | wt_fa_intra_sol_xover4 | wt_lk_ball_wtd | wt_fa_elec | wt_pro_close | wt_hbond_sr_bb | wt_hbond_lr_bb | wt_hbond_bb_sc | wt_hbond_sc | wt_dslf_fa13 | wt_metalbinding_constraint | wt_omega | wt_fa_dun | wt_p_aa_pp | wt_yhh_planarity | wt_ref | wt_rama_prepro | wt_total_score | mut_fa_atr | mut_fa_rep | mut_fa_sol | mut_fa_intra_rep | mut_fa_intra_sol_xover4 | mut_lk_ball_wtd | mut_fa_elec | mut_pro_close | mut_hbond_sr_bb | mut_hbond_lr_bb | mut_hbond_bb_sc | mut_hbond_sc | mut_dslf_fa13 | mut_metalbinding_constraint | mut_omega | mut_fa_dun | mut_p_aa_pp | mut_yhh_planarity | mut_ref | mut_rama_prepro | mut_total_score | delta_ref | delta_fa_dun | delta_fa_elec | delta_dslf_fa13 | delta_hbond_lr_bb | delta_omega | delta_pro_close | delta_fa_intra_rep | delta_fa_intra_sol_xover4 | delta_fa_sol | delta_hbond_sr_bb | delta_fa_rep | delta_fa_atr | delta_total_score | delta_rama_prepro | delta_lk_ball_wtd | delta_metalbinding_constraint | delta_p_aa_pp | delta_hbond_sc | delta_yhh_planarity | delta_hbond_bb_sc | highest_contributor | highest_contributor_value | highest_contributor_wordy | lowest_contributor | lowest_contributor_value | lowest_contributor_wordy | |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 0 | model | W494R | 11.5642 | -2636.87 | -2625.3 | 0.0751774 | 0.0534519 | 0.438244 | 0.261443 | 0.596571 | -4266.27 | 502.781 | 2365.99 | 8.08926 | 135.348 | -92.4808 | -1364.08 | 6.22781 | -206.578 | -219.666 | -222.104 | -151.417 | 0 | 9.86076e-29 | 30.9287 | 769.475 | -157.417 | 0 | 215.226 | 9.07527 | -0 | -4259.44 | 502.128 | 2370.75 | 8.04969 | 135.072 | -93.0502 | -1361.64 | 6.17517 | -206.224 | -219.896 | -221.646 | -151.579 | 0 | 9.86076e-29 | 31.1934 | 769.536 | -156.867 | 0.00722052 | 212.87 | 9.25613 | -0 | -2.35573 | 0.0604955 | 2.44202 | 0 | -0.22989 | 0.264638 | -0.0526317 | -0.0395649 | -0.275995 | 4.75991 | 0.353592 | -0.653089 | 6.82532 | 0 | 0.180856 | -0.569347 | 0 | 0.549845 | -0.162218 | 0.00722052 | 0.458815 | fa_atr | 6.82532 | Lennard-Jones attractive between atoms in different residues (r^6 term, London dispersion forces). | ref | -2.35573 | Reference energy for each amino acid. Balances internal energy of amino acid terms. Plays role in design. |
| 1 | model | R32T | 1.8513 | -2636.87 | -2635.02 | 0.0430545 | 0.0311775 | 0.507276 | 0.4322 | 0.852001 | -4266.27 | 502.781 | 2365.99 | 8.08926 | 135.348 | -92.4808 | -1364.08 | 6.22781 | -206.578 | -219.666 | -222.104 | -151.417 | 0 | 9.86076e-29 | 30.9287 | 769.475 | -157.417 | 0 | 215.226 | 9.07527 | -0 | -4259.85 | 502.292 | 2358.48 | 8.08464 | 135.206 | -90.6887 | -1362.4 | 6.24715 | -206.539 | -219.642 | -221.33 | -151.481 | 0 | 0.0497096 | 30.9976 | 767.77 | -157.563 | 0.000361175 | 216.473 | 8.87581 | -0 | 1.24649 | -1.70496 | 1.67898 | 0 | 0.0233144 | 0.0688236 | 0.0193443 | -0.0046154 | -0.141243 | -7.51055 | 0.0385872 | -0.488713 | 6.41866 | 0 | -0.199459 | 1.79213 | 0.0497096 | -0.145879 | -0.06434 | 0.000361175 | 0.77466 | fa_atr | 6.41866 | Lennard-Jones attractive between atoms in different residues (r^6 term, London dispersion forces). | fa_sol | -7.51055 | Lazaridis-Karplus solvation energy. |
| 2 | model | R387G | 5.48082 | -2636.87 | -2631.39 | 0.0366166 | 0.0331456 | 0.370686 | 0.64359 | 1.73621 | -4266.27 | 502.781 | 2365.99 | 8.08926 | 135.348 | -92.4808 | -1364.08 | 6.22781 | -206.578 | -219.666 | -222.104 | -151.417 | 0 | 9.86076e-29 | 30.9287 | 769.475 | -157.417 | 0 | 215.226 | 9.07527 | -0 | -4257.61 | 502.503 | 2357.96 | 8.07043 | 134.731 | -92.7794 | -1360.05 | 6.2686 | -206.65 | -219.614 | -222.139 | -148.301 | 0 | 0.0129584 | 30.9599 | 766.054 | -156.817 | 0.000562709 | 216.119 | 9.88497 | -0 | 0.8929 | -3.42088 | 4.03492 | 0 | 0.0515675 | 0.0311272 | 0.0407945 | -0.0188271 | -0.616943 | -8.02839 | -0.071805 | -0.277596 | 8.65802 | 0 | 0.8097 | -0.298601 | 0.0129584 | 0.599977 | 3.11643 | 0.000562709 | -0.0350902 | fa_atr | 8.65802 | Lennard-Jones attractive between atoms in different residues (r^6 term, London dispersion forces). | fa_sol | -8.02839 | Lazaridis-Karplus solvation energy. |
| 3 | model | R497Q | 7.64264 | -2636.87 | -2629.23 | 0.0825427 | 0.0423215 | 0.250685 | 0.153106 | 0.610752 | -4266.27 | 502.781 | 2365.99 | 8.08926 | 135.348 | -92.4808 | -1364.08 | 6.22781 | -206.578 | -219.666 | -222.104 | -151.417 | 0 | 9.86076e-29 | 30.9287 | 769.475 | -157.417 | 0 | 215.226 | 9.07527 | -0 | -4261.09 | 503.187 | 2361.16 | 8.05313 | 134.755 | -93.4404 | -1356.88 | 6.13576 | -206.611 | -219.752 | -220.678 | -150.807 | 0 | 9.86076e-29 | 31.1039 | 769.463 | -157.187 | 0.0117609 | 213.87 | 9.48035 | -0 | -1.35621 | -0.0120372 | 7.20412 | 0 | -0.0858482 | 0.17516 | -0.0920469 | -0.0361254 | -0.593117 | -4.8295 | -0.0330634 | 0.406041 | 5.1725 | 0 | 0.405074 | -0.959527 | 0 | 0.229551 | 0.609898 | 0.0117609 | 1.42602 | fa_elec | 7.20412 | Coulombic electrostatic potential with a distance-dependent dielectric. | fa_sol | -4.8295 | Lazaridis-Karplus solvation energy. |
| 4 | model | E94K | 28.6743 | -2636.87 | -2608.19 | 0.293049 | 0.142442 | 0.218453 | 0.201877 | 0.924123 | -4266.27 | 502.781 | 2365.99 | 8.08926 | 135.348 | -92.4808 | -1364.08 | 6.22781 | -206.578 | -219.666 | -222.104 | -151.417 | 0 | 9.86076e-29 | 30.9287 | 769.475 | -157.417 | 0 | 215.226 | 9.07527 | -0 | -4265.41 | 508.027 | 2366.33 | 8.07766 | 134.235 | -93.0443 | -1350.93 | 6.20731 | -206.728 | -219.793 | -217.286 | -149.004 | 0 | 0.0451577 | 31.841 | 770.745 | -157.53 | 0.00362233 | 217.236 | 8.78323 | -0 | 2.00995 | 1.26976 | 13.149 | 0 | -0.127533 | 0.912204 | -0.0204994 | -0.0115955 | -1.11288 | 0.339037 | -0.150496 | 5.24619 | 0.860409 | 0 | -0.292038 | -0.563469 | 0.0451577 | -0.113428 | 2.4128 | 0.00362233 | 4.8181 | fa_elec | 13.149 | Coulombic electrostatic potential with a distance-dependent dielectric. | fa_intra_sol_xover4 | -1.11288 | Intra-residue LK solvation, counted for the atom-pairs beyond torsion-relationship. |
| 5 | model | E125G | 7.72434 | -2636.87 | -2629.14 | 0.0518754 | 0.0563029 | 0.34083 | 0.397814 | 1.16719 | -4266.27 | 502.781 | 2365.99 | 8.08926 | 135.348 | -92.4808 | -1364.08 | 6.22781 | -206.578 | -219.666 | -222.104 | -151.417 | 0 | 9.86076e-29 | 30.9287 | 769.475 | -157.417 | 0 | 215.226 | 9.07527 | -0 | -4262.2 | 504.784 | 2361.65 | 8.07048 | 135.067 | -92.9581 | -1360.49 | 6.22807 | -206.641 | -219.607 | -220.221 | -151.457 | 0 | 0.0108582 | 30.955 | 766.442 | -157.06 | 0 | 218.749 | 9.54048 | -0 | 3.52269 | -3.03302 | 3.59083 | 0 | 0.0585111 | 0.0262586 | 0.000264657 | -0.0187804 | -0.280606 | -4.34434 | -0.0627722 | 2.0034 | 4.06332 | 0 | 0.465211 | -0.477267 | 0.0108582 | 0.357265 | -0.0404141 | 0 | 1.88292 | fa_atr | 4.06332 | Lennard-Jones attractive between atoms in different residues (r^6 term, London dispersion forces). | fa_sol | -4.34434 | Lazaridis-Karplus solvation energy. |
| 6 | model | C571R | 24.7828 | -2636.87 | -2612.09 | 0.426129 | 0.157989 | 0.132207 | 0.230358 | 1.74241 | -4266.27 | 502.781 | 2365.99 | 8.08926 | 135.348 | -92.4808 | -1364.08 | 6.22781 | -206.578 | -219.666 | -222.104 | -151.417 | 0 | 9.86076e-29 | 30.9287 | 769.475 | -157.417 | 0 | 215.226 | 9.07527 | -0 | -4275.56 | 523.255 | 2374.81 | 8.07035 | 135.764 | -91.6686 | -1362.92 | 6.18973 | -206.584 | -221.566 | -222.657 | -151.081 | 0 | 0.0126243 | 31.9591 | 775.514 | -157.272 | 0.00237123 | 211.877 | 9.76565 | -0 | -3.34953 | 6.03872 | 1.15993 | 0 | -1.89984 | 1.03032 | -0.0380723 | -0.0189122 | 0.416559 | 8.81787 | -0.00615661 | 20.4745 | -9.28845 | 0 | 0.690382 | 0.812273 | 0.0126243 | 0.145503 | 0.33565 | 0.00237123 | -0.552972 | fa_rep | 20.4745 | Lennard-Jones repulsive between atoms in different residues (r^12 term, Pauli repulsion forces). | fa_atr | -9.28845 | Lennard-Jones attractive between atoms in different residues (r^6 term, London dispersion forces). |
R32 is a special case, as it is frequently monomethylated in Phosphosite Plus. So is likely to have a regulatory role, which is lost with R32T. Model with resn NMM not made as it is not a structural issue.
The M81∆ variant was made with the Remodel mover under the null hypothesis that the active site is unaffected and the helix away from the active site accomodates the different interactions resulting from the 100° offset. The lowest energy attainable was +27 kcal/mol. None of the neighbours had a positive potential, but were mostly smaller than the residue pose —indicating that it is not a single residue erroneously placed, but the whole neighbourhood has a smaller difference in folding energy. Therefore, this variant is likely to be unstable.
| Residue | dG_mut | dG_wt | ddG | |
|---|---|---|---|---|
| 0 | V56 A | -4.4 | -5.9 | 1.4 |
| 1 | K60 A | -4.3 | -2.3 | -2 |
| 2 | A66 A | -2.9 | -4.7 | 1.7 |
| 3 | L77 A | -4.2 | -5.7 | 1.5 |
| 4 | A85 A | -0.9 | -2.4 | 1.5 |
| 5 | G86 A | -0.8 | -2 | 1.2 |
| 6 | A87 A | -0.1 | -5.4 | 5.3 |
| 7 | D88 A | -1.6 | -6.2 | 4.6 |
| 8 | F89 A | -0.2 | -2.4 | 2.1 |
| 9 | F146 A | -2.2 | -3.3 | 1.1 |
| 10 | K195 A | -3.6 | -6 | 2.4 |
| 11 | L196 A | -4.2 | -5.3 | 1 |
| 12 | P212 A | -3 | -4 | 1.1 |
| 13 | D215 A | -4.4 | -3.1 | -1.3 |
| 14 | F299 A | 0.2 | 2.6 | -2.4 |
| 16 | F540 A | -3.5 | -4.9 | 1.4 |
| 17 | I544 A | -2.7 | -0.7 | -2 |