Starting phenix.real_space_refine on Tue Aug 4 14:14:59 2026 by dcliebschner =============================================================================== Processing files: ------------------------------------------------------------------------------- Found model, /net/cci-nas-00/data/ceres_data/8ws8_37801/08_2026/8ws8_37801.cif Found real_map, /net/cci-nas-00/data/ceres_data/8ws8_37801/08_2026/8ws8_37801.map Processing PHIL parameters: ------------------------------------------------------------------------------- Adding command-line PHIL: ------------------------- refinement.macro_cycles=10 scattering_table=electron resolution=2.96 write_initial_geo_file=False Final processed PHIL parameters: ------------------------------------------------------------------------------- data_manager { model { file = "/net/cci-nas-00/data/ceres_data/8ws8_37801/08_2026/8ws8_37801.cif" } default_model = "/net/cci-nas-00/data/ceres_data/8ws8_37801/08_2026/8ws8_37801.cif" real_map_files = "/net/cci-nas-00/data/ceres_data/8ws8_37801/08_2026/8ws8_37801.map" default_real_map = "/net/cci-nas-00/data/ceres_data/8ws8_37801/08_2026/8ws8_37801.map" } resolution = 2.96 write_initial_geo_file = False refinement { macro_cycles = 10 } Starting job =============================================================================== ------------------------------------------------------------------------------- Citation: ********* Afonine PV, Poon BK, Read RJ, Sobolev OV, Terwilliger TC, Urzhumtsev A, Adams PD. (2018) Real-space refinement in PHENIX for cryo-EM and crystallography. Acta Cryst. D74:531-544. Validating inputs Origin is already at (0, 0, 0), no shifts will be applied ------------------------------------------------------------------------------- Processing inputs ***************** Set random seed Set to: 0 Set model cs if undefined Decide on map wrapping Map wrapping is set to: False Normalize map: mean=0, sd=1 Input map: mean= 0.000 sd= 0.014 Set stop_for_unknowns flag Set to: True Assert model is a single copy model Assert all atoms have isotropic ADPs Construct map_model_manager Extract box with map and model Check model and map are aligned Set scattering table Set to: electron Number of scattering types: 5 Type Number sf(0) Gaussians P 74 5.49 5 S 3 5.16 5 C 2211 2.51 5 N 706 2.21 5 O 898 1.98 5 sf(0) = scattering factor at diffraction angle 0. Process input model Symmetric amino acids flipped. Time to flip 3 residue(s): 0.00s Monomer Library directory: "/net/cci-filer3/home/dcliebschner/04_cryoem/Phenix-2.2rc3-6140/lib/python3.11/site-packages/chem_data/mon_lib" Total number of atoms: 3892 Number of models: 1 Model: "" Number of chains: 4 Chain: "A" Number of atoms: 2348 Number of conformers: 1 Conformer: "" Number of residues, atoms: 298, 2348 Classifications: {'peptide': 298} Incomplete info: {'backbone_only': 1, 'truncation_to_alanine': 1} Link IDs: {'PTRANS': 29, 'TRANS': 268} Unresolved chain link angles: 1 Unresolved non-hydrogen bonds: 8 Unresolved non-hydrogen angles: 11 Unresolved non-hydrogen dihedrals: 6 Unresolved non-hydrogen chiralities: 1 Planarities with less than four sites: {'GLU:plan': 1} Unresolved non-hydrogen planarities: 4 Chain: "C" Number of atoms: 562 Number of conformers: 1 Conformer: "" Number of residues, atoms: 27, 562 Classifications: {'DNA': 27} Link IDs: {'rna3p': 26} Chain: "B" Number of atoms: 843 Number of conformers: 1 Conformer: "" Number of residues, atoms: 40, 843 Classifications: {'RNA': 40} Modifications used: {'rna2p_pyr': 6, 'rna3p_pur': 16, 'rna3p_pyr': 18} Link IDs: {'rna2p': 6, 'rna3p': 33} Chain: "D" Number of atoms: 139 Number of conformers: 1 Conformer: "" Number of residues, atoms: 7, 139 Classifications: {'DNA': 7} Link IDs: {'rna3p': 6} Time building chain proxies: 1.11, per 1000 atoms: 0.29 Number of scatterers: 3892 At special positions: 0 Unit cell: (68.64, 89.76, 106.26, 90, 90, 90) Space group: P 1 (No. 1) Number of sites at special positions: 0 Number of scattering types: 5 Type Number sf(0) S 3 16.00 P 74 15.00 O 898 8.00 N 706 7.00 C 2211 6.00 sf(0) = scattering factor at diffraction angle 0. Number of disulfides: simple=0, symmetry=0 Automatic linking Parameters for automatic linking Linking & cutoffs Metal : Auto - 3.00 Amino acid : True - 1.90 Carbohydrate : True - 1.99 Ligands : True - 1.99 Small molecules : False - 1.98 Amino acid - RNA/DNA : False Number of custom bonds: simple=0, symmetry=0 Time building additional restraints: 0.44 Conformation dependent library (CDL) restraints added in 110.7 milliseconds 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. Adding C-beta torsion restraints... Number of C-beta restraints generated: 542 Finding SS restraints... Secondary structure from input PDB file: 9 helices and 1 sheets defined 35.6% alpha, 6.4% beta 13 base pairs and 37 stacking pairs defined. Time for finding SS restraints: 0.30 Creating SS restraints... Processing helix chain 'A' and resid 75 through 90 removed outlier: 3.885A pdb=" N GLY A 90 " --> pdb=" O GLU A 86 " (cutoff:3.500A) Processing helix chain 'A' and resid 94 through 99 Processing helix chain 'A' and resid 105 through 117 removed outlier: 3.780A pdb=" N PHE A 113 " --> pdb=" O LEU A 109 " (cutoff:3.500A) removed outlier: 3.528A pdb=" N ALA A 114 " --> pdb=" O ASP A 110 " (cutoff:3.500A) Processing helix chain 'A' and resid 125 through 131 removed outlier: 3.701A pdb=" N LEU A 131 " --> pdb=" O GLN A 127 " (cutoff:3.500A) Processing helix chain 'A' and resid 133 through 169 removed outlier: 3.689A pdb=" N ILE A 144 " --> pdb=" O TYR A 140 " (cutoff:3.500A) removed outlier: 3.933A pdb=" N ARG A 150 " --> pdb=" O LYS A 146 " (cutoff:3.500A) removed outlier: 4.448A pdb=" N ASN A 151 " --> pdb=" O VAL A 147 " (cutoff:3.500A) removed outlier: 3.548A pdb=" N GLU A 152 " --> pdb=" O GLU A 148 " (cutoff:3.500A) removed outlier: 3.863A pdb=" N LYS A 153 " --> pdb=" O THR A 149 " (cutoff:3.500A) Processing helix chain 'A' and resid 247 through 251 Processing helix chain 'A' and resid 281 through 288 Processing helix chain 'A' and resid 310 through 317 removed outlier: 3.569A pdb=" N TYR A 317 " --> pdb=" O LEU A 313 " (cutoff:3.500A) Processing helix chain 'A' and resid 327 through 332 Processing sheet with id=AA1, first strand: chain 'A' and resid 198 through 199 removed outlier: 3.759A pdb=" N ILE A 347 " --> pdb=" O ASP A 342 " (cutoff:3.500A) removed outlier: 4.456A pdb=" N ASN A 64 " --> pdb=" O ASP A 308 " (cutoff:3.500A) removed outlier: 7.057A pdb=" N VAL A 306 " --> pdb=" O LEU A 66 " (cutoff:3.500A) removed outlier: 3.574A pdb=" N ILE A 296 " --> pdb=" O MET A 307 " (cutoff:3.500A) 75 hydrogen bonds defined for protein. 210 hydrogen bond angles defined for protein. Restraints generated for nucleic acids: 34 hydrogen bonds 64 hydrogen bond angles 0 basepair planarities 13 basepair parallelities 37 stacking parallelities Total time for adding SS restraints: 0.67 Time building geometry restraints manager: 0.56 seconds NOTE: a complete listing of the restraints can be obtained by requesting output of .geo file. Histogram of bond lengths: 1.20 - 1.32: 516 1.32 - 1.44: 1486 1.44 - 1.57: 1985 1.57 - 1.69: 145 1.69 - 1.81: 4 Bond restraints: 4136 Sorted by residual: bond pdb=" CA LYS A 273 " pdb=" C LYS A 273 " ideal model delta sigma weight residual 1.519 1.560 -0.041 1.16e-02 7.43e+03 1.23e+01 bond pdb=" C3' DA C -10 " pdb=" C2' DA C -10 " ideal model delta sigma weight residual 1.516 1.544 -0.028 8.00e-03 1.56e+04 1.21e+01 bond pdb=" C1' DT C -18 " pdb=" N1 DT C -18 " ideal model delta sigma weight residual 1.468 1.512 -0.044 1.40e-02 5.10e+03 9.83e+00 bond pdb=" C3' DG C -5 " pdb=" C2' DG C -5 " ideal model delta sigma weight residual 1.516 1.540 -0.024 8.00e-03 1.56e+04 9.05e+00 bond pdb=" C3' DA C -6 " pdb=" C2' DA C -6 " ideal model delta sigma weight residual 1.516 1.539 -0.023 8.00e-03 1.56e+04 8.36e+00 ... (remaining 4131 not shown) Histogram of bond angle deviations from ideal: 0.00 - 1.89: 5615 1.89 - 3.79: 304 3.79 - 5.68: 17 5.68 - 7.57: 3 7.57 - 9.46: 1 Bond angle restraints: 5940 Sorted by residual: angle pdb=" O4' DT D -2 " pdb=" C4' DT D -2 " pdb=" C3' DT D -2 " ideal model delta sigma weight residual 106.00 102.78 3.22 6.00e-01 2.78e+00 2.88e+01 angle pdb=" N3 DT C -7 " pdb=" C4 DT C -7 " pdb=" O4 DT C -7 " ideal model delta sigma weight residual 119.90 122.71 -2.81 6.00e-01 2.78e+00 2.19e+01 angle pdb=" O4 DT C -7 " pdb=" C4 DT C -7 " pdb=" C5 DT C -7 " ideal model delta sigma weight residual 124.90 121.87 3.03 7.00e-01 2.04e+00 1.87e+01 angle pdb=" N3 DT D -1 " pdb=" C4 DT D -1 " pdb=" O4 DT D -1 " ideal model delta sigma weight residual 119.90 122.42 -2.52 6.00e-01 2.78e+00 1.76e+01 angle pdb=" N3 DT C -19 " pdb=" C4 DT C -19 " pdb=" O4 DT C -19 " ideal model delta sigma weight residual 119.90 122.37 -2.47 6.00e-01 2.78e+00 1.69e+01 ... (remaining 5935 not shown) Histogram of dihedral angle deviations from ideal: 0.00 - 34.92: 2208 34.92 - 69.84: 251 69.84 - 104.76: 16 104.76 - 139.68: 0 139.68 - 174.60: 2 Dihedral angle restraints: 2477 sinusoidal: 1628 harmonic: 849 Sorted by residual: dihedral pdb=" O4' U B 10 " pdb=" C1' U B 10 " pdb=" N1 U B 10 " pdb=" C2 U B 10 " ideal model delta sinusoidal sigma weight residual 200.00 38.61 161.39 1 1.50e+01 4.44e-03 8.31e+01 dihedral pdb=" O4' U B -9 " pdb=" C1' U B -9 " pdb=" N1 U B -9 " pdb=" C2 U B -9 " ideal model delta sinusoidal sigma weight residual 232.00 57.40 174.60 1 1.70e+01 3.46e-03 6.63e+01 dihedral pdb=" O4' A B 18 " pdb=" C1' A B 18 " pdb=" N9 A B 18 " pdb=" C4 A B 18 " ideal model delta sinusoidal sigma weight residual 70.00 3.04 66.96 1 2.00e+01 2.50e-03 1.46e+01 ... (remaining 2474 not shown) Histogram of chiral volume deviations from ideal: 0.000 - 0.043: 467 0.043 - 0.086: 115 0.086 - 0.129: 67 0.129 - 0.172: 31 0.172 - 0.215: 3 Chirality restraints: 683 Sorted by residual: chirality pdb=" CA ILE A 294 " pdb=" N ILE A 294 " pdb=" C ILE A 294 " pdb=" CB ILE A 294 " both_signs ideal model delta sigma weight residual False 2.43 2.65 -0.22 2.00e-01 2.50e+01 1.16e+00 chirality pdb=" CA ASP A 303 " pdb=" N ASP A 303 " pdb=" C ASP A 303 " pdb=" CB ASP A 303 " both_signs ideal model delta sigma weight residual False 2.51 2.69 -0.17 2.00e-01 2.50e+01 7.62e-01 chirality pdb=" CA ILE A 144 " pdb=" N ILE A 144 " pdb=" C ILE A 144 " pdb=" CB ILE A 144 " both_signs ideal model delta sigma weight residual False 2.43 2.61 -0.17 2.00e-01 2.50e+01 7.62e-01 ... (remaining 680 not shown) Planarity restraints: 500 Sorted by residual: delta sigma weight rms_deltas residual plane pdb=" C VAL A 118 " 0.050 5.00e-02 4.00e+02 7.62e-02 9.29e+00 pdb=" N PRO A 119 " -0.132 5.00e-02 4.00e+02 pdb=" CA PRO A 119 " 0.040 5.00e-02 4.00e+02 pdb=" CD PRO A 119 " 0.042 5.00e-02 4.00e+02 delta sigma weight rms_deltas residual plane pdb=" C ILE A 294 " 0.030 5.00e-02 4.00e+02 4.50e-02 3.23e+00 pdb=" N PRO A 295 " -0.078 5.00e-02 4.00e+02 pdb=" CA PRO A 295 " 0.022 5.00e-02 4.00e+02 pdb=" CD PRO A 295 " 0.026 5.00e-02 4.00e+02 delta sigma weight rms_deltas residual plane pdb=" C TRP A 75 " 0.029 5.00e-02 4.00e+02 4.46e-02 3.18e+00 pdb=" N PRO A 76 " -0.077 5.00e-02 4.00e+02 pdb=" CA PRO A 76 " 0.023 5.00e-02 4.00e+02 pdb=" CD PRO A 76 " 0.025 5.00e-02 4.00e+02 ... (remaining 497 not shown) Histogram of nonbonded interaction distances: 2.25 - 2.78: 884 2.78 - 3.31: 3342 3.31 - 3.84: 7116 3.84 - 4.37: 8363 4.37 - 4.90: 12372 Nonbonded interactions: 32077 Sorted by model distance: nonbonded pdb=" N2 DG C 5 " pdb=" O2 DC D -5 " model vdw 2.253 3.120 nonbonded pdb=" NH1 ARG A 260 " pdb=" O6 G B -16 " model vdw 2.297 3.120 nonbonded pdb=" O2' U B -10 " pdb=" O2 U B -9 " model vdw 2.342 3.040 nonbonded pdb=" NH2 ARG A 260 " pdb=" OP2 U B -17 " model vdw 2.352 3.120 nonbonded pdb=" N1 DG C -12 " pdb=" O6 G B 11 " model vdw 2.357 3.120 ... (remaining 32072 not shown) NOTE: a complete listing of the restraints can be obtained by requesting output of .geo file. Find NCS groups from input model Time spend for trying shortcut: 0.00 ========== WARNING! ============ No NCS relation were found !!! ================================ Found NCS groups: found none. Set up NCS constraints No NCS constraints will be used in refinement. Set refine NCS operators Adjust number of macro_cycles Number of macro_cycles: 10 Reset NCS operators Extract rigid body selections Check and reset occupancies Occupancies: min=1.00 max=1.00 mean=1.00 Load rotamer database and sin/cos tables Set ADP refinement strategy ADPs will be refined as individual isotropic Make a string to write initial .geo file Internal consistency checks Time: Set random seed: 0.000 Set model cs if undefined: 0.000 Decide on map wrapping: 0.000 Normalize map: mean=0, sd=1: 4.420 Set stop_for_unknowns flag: 0.000 Assert model is a single copy model: 0.000 Assert all atoms have isotropic ADPs: 0.000 Construct map_model_manager: 0.010 Extract box with map and model: 0.110 Check model and map are aligned: 0.010 Set scattering table: 0.010 Process input model: 5.710 Find NCS groups from input model: 0.010 Set up NCS constraints: 0.010 Set refine NCS operators: 0.000 Adjust number of macro_cycles: 0.000 Reset NCS operators: 0.000 Extract rigid body selections: 0.000 Check and reset occupancies: 0.000 Load rotamer database and sin/cos tables:6.440 Set ADP refinement strategy: 0.000 Make a string to write initial .geo file:0.000 Internal consistency checks: 0.000 Total: 16.730 ------------------------------------------------------------------------------- Set refinement monitor ********************** ------------------------------------------------------------------------------- Setup refinement engine *********************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.6644 moved from start: 0.0000 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.005 0.044 4136 Z= 0.412 Angle : 0.821 9.464 5940 Z= 0.634 Chirality : 0.056 0.215 683 Planarity : 0.007 0.076 500 Dihedral : 22.749 174.597 1935 Min Nonbonded Distance : 2.253 Molprobity Statistics. All-atom Clashscore : 7.46 Ramachandran Plot: Outliers : 0.00 % Allowed : 7.09 % Favored : 92.91 % Rotamer: Outliers : 0.81 % Allowed : 2.43 % Favored : 96.76 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.94 (0.51), residues: 296 helix: -0.09 (0.53), residues: 104 sheet: -1.65 (1.04), residues: 26 loop : -0.74 (0.51), residues: 166 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.007 0.001 ARG A 319 TYR 0.013 0.002 TYR A 317 PHE 0.013 0.001 PHE A 137 TRP 0.017 0.002 TRP A 318 HIS 0.001 0.000 HIS A 214 Details of bonding type rmsd/Z covalent geometry : bond 0.00498 / 0.41 ( 4136) covalent geometry : angle 0.82109 / 0.63 ( 5940) hydrogen bonds : bond 0.12271 / 7.67 ( 109) hydrogen bonds : angle 6.22272 / 4.25 ( 274) *********************** REFINEMENT MACRO_CYCLE 1 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 47 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 2 poor density : 45 time to evaluate : 0.109 Fit side-chains REVERT: A 138 ASN cc_start: 0.7705 (m110) cc_final: 0.7463 (m110) REVERT: A 153 LYS cc_start: 0.6839 (mppt) cc_final: 0.6580 (mptt) REVERT: A 314 ARG cc_start: 0.6120 (ttt-90) cc_final: 0.5300 (ttt-90) outliers start: 2 outliers final: 0 residues processed: 47 average time/residue: 0.7899 time to fit residues: 38.0280 Evaluate side-chains 40 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 0 poor density : 40 time to evaluate : 0.100 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Rotamers are restrained with sigma=5.00 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 24 optimal weight: 0.7980 chunk 26 optimal weight: 0.4980 chunk 2 optimal weight: 0.0980 chunk 16 optimal weight: 0.0470 chunk 32 optimal weight: 2.9990 chunk 31 optimal weight: 2.9990 chunk 25 optimal weight: 0.5980 chunk 19 optimal weight: 0.9990 chunk 30 optimal weight: 4.9990 chunk 22 optimal weight: 0.5980 chunk 13 optimal weight: 0.0980 overall best weight: 0.2678 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 135 HIS ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 1 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4274 r_free = 0.4274 target = 0.184895 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 55)----------------| | r_work = 0.3903 r_free = 0.3903 target = 0.153978 restraints weight = 5318.940| |-----------------------------------------------------------------------------| r_work (start): 0.3891 rms_B_bonded: 1.31 r_work: 0.3789 rms_B_bonded: 2.21 restraints_weight: 0.5000 r_work: 0.3665 rms_B_bonded: 3.79 restraints_weight: 0.2500 r_work (final): 0.3665 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.6965 moved from start: 0.0886 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.003 0.027 4136 Z= 0.151 Angle : 0.597 11.485 5940 Z= 0.341 Chirality : 0.042 0.183 683 Planarity : 0.006 0.041 500 Dihedral : 25.072 177.561 1341 Min Nonbonded Distance : 2.433 Molprobity Statistics. All-atom Clashscore : 8.02 Ramachandran Plot: Outliers : 0.00 % Allowed : 6.76 % Favored : 93.24 % Rotamer: Outliers : 1.62 % Allowed : 12.55 % Favored : 85.83 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.66 (0.50), residues: 296 helix: 0.17 (0.52), residues: 105 sheet: -1.01 (1.07), residues: 26 loop : -0.68 (0.51), residues: 165 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.004 0.000 ARG A 319 TYR 0.009 0.001 TYR A 89 PHE 0.009 0.001 PHE A 137 TRP 0.015 0.002 TRP A 318 HIS 0.001 0.000 HIS A 214 Details of bonding type rmsd/Z covalent geometry : bond 0.00272 / 0.15 ( 4136) covalent geometry : angle 0.59737 / 0.34 ( 5940) hydrogen bonds : bond 0.05817 / 4.26 ( 109) hydrogen bonds : angle 4.73494 / 3.21 ( 274) *********************** REFINEMENT MACRO_CYCLE 2 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 37 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 4 poor density : 33 time to evaluate : 0.126 Fit side-chains REVERT: A 97 GLU cc_start: 0.7660 (mp0) cc_final: 0.7340 (mp0) REVERT: A 138 ASN cc_start: 0.7936 (m110) cc_final: 0.7515 (m110) REVERT: A 153 LYS cc_start: 0.7097 (mppt) cc_final: 0.6693 (mptt) REVERT: A 314 ARG cc_start: 0.6193 (ttt-90) cc_final: 0.5232 (ttt-90) outliers start: 4 outliers final: 3 residues processed: 35 average time/residue: 0.8124 time to fit residues: 29.1337 Evaluate side-chains 36 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 3 poor density : 33 time to evaluate : 0.111 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=4.50 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 21 optimal weight: 0.6980 chunk 0 optimal weight: 1.9990 chunk 9 optimal weight: 0.2980 chunk 5 optimal weight: 0.2980 chunk 28 optimal weight: 0.7980 chunk 12 optimal weight: 0.0070 chunk 15 optimal weight: 0.7980 chunk 23 optimal weight: 0.0970 chunk 29 optimal weight: 10.0000 chunk 16 optimal weight: 0.9990 chunk 34 optimal weight: 9.9990 overall best weight: 0.2796 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 1 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4277 r_free = 0.4277 target = 0.185061 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 55)----------------| | r_work = 0.3909 r_free = 0.3909 target = 0.154432 restraints weight = 5189.126| |-----------------------------------------------------------------------------| r_work (start): 0.3904 rms_B_bonded: 1.28 r_work: 0.3804 rms_B_bonded: 2.16 restraints_weight: 0.5000 r_work: 0.3682 rms_B_bonded: 3.74 restraints_weight: 0.2500 r_work (final): 0.3682 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.6956 moved from start: 0.1102 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.003 0.029 4136 Z= 0.146 Angle : 0.554 10.017 5940 Z= 0.322 Chirality : 0.040 0.168 683 Planarity : 0.005 0.036 500 Dihedral : 24.977 177.367 1341 Min Nonbonded Distance : 2.431 Molprobity Statistics. All-atom Clashscore : 6.76 Ramachandran Plot: Outliers : 0.00 % Allowed : 5.41 % Favored : 94.59 % Rotamer: Outliers : 2.02 % Allowed : 14.17 % Favored : 83.81 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.52 (0.51), residues: 296 helix: 0.31 (0.52), residues: 107 sheet: -0.95 (1.15), residues: 26 loop : -0.63 (0.52), residues: 163 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.002 0.000 ARG A 262 TYR 0.007 0.001 TYR A 201 PHE 0.008 0.001 PHE A 137 TRP 0.012 0.001 TRP A 318 HIS 0.002 0.001 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00266 / 0.15 ( 4136) covalent geometry : angle 0.55371 / 0.32 ( 5940) hydrogen bonds : bond 0.05338 / 3.81 ( 109) hydrogen bonds : angle 4.28108 / 2.88 ( 274) *********************** REFINEMENT MACRO_CYCLE 3 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 40 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 5 poor density : 35 time to evaluate : 0.093 Fit side-chains REVERT: A 97 GLU cc_start: 0.7640 (mp0) cc_final: 0.7390 (mp0) REVERT: A 138 ASN cc_start: 0.7962 (m110) cc_final: 0.7480 (m110) REVERT: A 153 LYS cc_start: 0.7123 (mppt) cc_final: 0.6760 (mptt) REVERT: A 314 ARG cc_start: 0.6340 (ttt-90) cc_final: 0.5048 (ttt-90) outliers start: 5 outliers final: 4 residues processed: 36 average time/residue: 0.7632 time to fit residues: 28.1748 Evaluate side-chains 37 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 4 poor density : 33 time to evaluate : 0.070 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=4.00 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 23 optimal weight: 0.9990 chunk 13 optimal weight: 1.9990 chunk 15 optimal weight: 0.6980 chunk 12 optimal weight: 0.5980 chunk 20 optimal weight: 0.0020 chunk 1 optimal weight: 0.0470 chunk 6 optimal weight: 0.7980 chunk 3 optimal weight: 0.5980 chunk 5 optimal weight: 0.5980 chunk 14 optimal weight: 0.2980 chunk 22 optimal weight: 2.9990 overall best weight: 0.3086 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 1 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4274 r_free = 0.4274 target = 0.184776 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 53)----------------| | r_work = 0.3900 r_free = 0.3900 target = 0.153737 restraints weight = 5165.789| |-----------------------------------------------------------------------------| r_work (start): 0.3894 rms_B_bonded: 1.26 r_work: 0.3790 rms_B_bonded: 2.19 restraints_weight: 0.5000 r_work: 0.3665 rms_B_bonded: 3.80 restraints_weight: 0.2500 r_work (final): 0.3665 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.6979 moved from start: 0.1253 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.003 0.031 4136 Z= 0.144 Angle : 0.548 10.164 5940 Z= 0.316 Chirality : 0.040 0.171 683 Planarity : 0.005 0.036 500 Dihedral : 24.924 177.826 1341 Min Nonbonded Distance : 2.433 Molprobity Statistics. All-atom Clashscore : 7.04 Ramachandran Plot: Outliers : 0.00 % Allowed : 6.08 % Favored : 93.92 % Rotamer: Outliers : 2.83 % Allowed : 16.60 % Favored : 80.57 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.44 (0.51), residues: 296 helix: 0.48 (0.53), residues: 107 sheet: -1.03 (1.12), residues: 26 loop : -0.63 (0.52), residues: 163 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.003 0.000 ARG A 262 TYR 0.017 0.002 TYR A 140 PHE 0.009 0.001 PHE A 137 TRP 0.011 0.001 TRP A 318 HIS 0.001 0.000 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00263 / 0.14 ( 4136) covalent geometry : angle 0.54809 / 0.32 ( 5940) hydrogen bonds : bond 0.05208 / 3.72 ( 109) hydrogen bonds : angle 4.12095 / 2.75 ( 274) *********************** REFINEMENT MACRO_CYCLE 4 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 39 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 7 poor density : 32 time to evaluate : 0.114 Fit side-chains REVERT: A 97 GLU cc_start: 0.7629 (OUTLIER) cc_final: 0.7410 (mp0) REVERT: A 138 ASN cc_start: 0.7977 (m110) cc_final: 0.7483 (m110) REVERT: A 153 LYS cc_start: 0.7161 (mppt) cc_final: 0.6761 (mptt) REVERT: A 314 ARG cc_start: 0.6339 (ttt-90) cc_final: 0.4734 (ttt-90) outliers start: 7 outliers final: 4 residues processed: 36 average time/residue: 0.7013 time to fit residues: 25.9302 Evaluate side-chains 38 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 5 poor density : 33 time to evaluate : 0.062 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 63 VAL Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 97 GLU Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=3.50 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 6 optimal weight: 0.6980 chunk 34 optimal weight: 9.9990 chunk 3 optimal weight: 0.0980 chunk 2 optimal weight: 0.5980 chunk 27 optimal weight: 0.6980 chunk 26 optimal weight: 0.7980 chunk 30 optimal weight: 3.9990 chunk 13 optimal weight: 0.7980 chunk 1 optimal weight: 0.2980 chunk 20 optimal weight: 0.0970 chunk 0 optimal weight: 1.9990 overall best weight: 0.3578 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 1 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4265 r_free = 0.4265 target = 0.184082 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 66)----------------| | r_work = 0.3889 r_free = 0.3889 target = 0.152923 restraints weight = 5221.810| |-----------------------------------------------------------------------------| r_work (start): 0.3887 rms_B_bonded: 1.29 r_work: 0.3785 rms_B_bonded: 2.19 restraints_weight: 0.5000 r_work: 0.3661 rms_B_bonded: 3.77 restraints_weight: 0.2500 r_work (final): 0.3661 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.6982 moved from start: 0.1355 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.003 0.030 4136 Z= 0.152 Angle : 0.557 10.357 5940 Z= 0.318 Chirality : 0.040 0.172 683 Planarity : 0.005 0.038 500 Dihedral : 24.897 178.491 1341 Min Nonbonded Distance : 2.436 Molprobity Statistics. All-atom Clashscore : 7.60 Ramachandran Plot: Outliers : 0.00 % Allowed : 5.41 % Favored : 94.59 % Rotamer: Outliers : 4.86 % Allowed : 15.38 % Favored : 79.76 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.39 (0.52), residues: 296 helix: 0.53 (0.53), residues: 107 sheet: -0.95 (1.13), residues: 26 loop : -0.62 (0.52), residues: 163 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.004 0.000 ARG A 262 TYR 0.011 0.001 TYR A 140 PHE 0.010 0.001 PHE A 137 TRP 0.011 0.002 TRP A 318 HIS 0.001 0.000 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00282 / 0.15 ( 4136) covalent geometry : angle 0.55708 / 0.32 ( 5940) hydrogen bonds : bond 0.05268 / 3.73 ( 109) hydrogen bonds : angle 4.08450 / 2.73 ( 274) *********************** REFINEMENT MACRO_CYCLE 5 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 45 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 12 poor density : 33 time to evaluate : 0.125 Fit side-chains REVERT: A 97 GLU cc_start: 0.7660 (OUTLIER) cc_final: 0.7459 (mp0) REVERT: A 138 ASN cc_start: 0.7996 (m110) cc_final: 0.7483 (m-40) REVERT: A 153 LYS cc_start: 0.7164 (mppt) cc_final: 0.6738 (mptt) REVERT: A 314 ARG cc_start: 0.6415 (ttt-90) cc_final: 0.4413 (ttt-90) outliers start: 12 outliers final: 5 residues processed: 41 average time/residue: 0.7513 time to fit residues: 31.6094 Evaluate side-chains 39 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 6 poor density : 33 time to evaluate : 0.105 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 63 VAL Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 97 GLU Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=3.00 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 32 optimal weight: 2.9990 chunk 24 optimal weight: 0.3980 chunk 12 optimal weight: 0.0040 chunk 2 optimal weight: 0.9980 chunk 15 optimal weight: 0.5980 chunk 3 optimal weight: 0.7980 chunk 18 optimal weight: 0.2980 chunk 33 optimal weight: 2.9990 chunk 29 optimal weight: 9.9990 chunk 21 optimal weight: 0.0770 chunk 26 optimal weight: 0.9980 overall best weight: 0.2750 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 1 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4266 r_free = 0.4266 target = 0.183623 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 56)----------------| | r_work = 0.3886 r_free = 0.3886 target = 0.152217 restraints weight = 5328.798| |-----------------------------------------------------------------------------| r_work (start): 0.3883 rms_B_bonded: 1.23 r_work: 0.3781 rms_B_bonded: 2.17 restraints_weight: 0.5000 r_work: 0.3658 rms_B_bonded: 3.75 restraints_weight: 0.2500 r_work (final): 0.3658 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.6983 moved from start: 0.1492 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.002 0.029 4136 Z= 0.137 Angle : 0.535 9.065 5940 Z= 0.309 Chirality : 0.040 0.169 683 Planarity : 0.005 0.035 500 Dihedral : 24.824 178.315 1341 Min Nonbonded Distance : 2.436 Molprobity Statistics. All-atom Clashscore : 7.60 Ramachandran Plot: Outliers : 0.00 % Allowed : 5.07 % Favored : 94.93 % Rotamer: Outliers : 2.43 % Allowed : 17.41 % Favored : 80.16 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.35 (0.52), residues: 296 helix: 0.56 (0.54), residues: 107 sheet: -0.98 (1.11), residues: 26 loop : -0.58 (0.52), residues: 163 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.004 0.000 ARG A 262 TYR 0.012 0.001 TYR A 140 PHE 0.008 0.001 PHE A 137 TRP 0.010 0.001 TRP A 318 HIS 0.001 0.000 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00247 / 0.14 ( 4136) covalent geometry : angle 0.53471 / 0.31 ( 5940) hydrogen bonds : bond 0.04872 / 3.44 ( 109) hydrogen bonds : angle 4.02386 / 2.69 ( 274) *********************** REFINEMENT MACRO_CYCLE 6 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 38 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 6 poor density : 32 time to evaluate : 0.064 Fit side-chains REVERT: A 153 LYS cc_start: 0.7152 (mppt) cc_final: 0.6757 (mptt) REVERT: A 314 ARG cc_start: 0.6366 (ttt-90) cc_final: 0.4423 (ttt-90) outliers start: 6 outliers final: 5 residues processed: 34 average time/residue: 0.6900 time to fit residues: 24.0498 Evaluate side-chains 37 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 5 poor density : 32 time to evaluate : 0.109 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 63 VAL Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=2.50 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 30 optimal weight: 3.9990 chunk 31 optimal weight: 3.9990 chunk 32 optimal weight: 2.9990 chunk 18 optimal weight: 0.0980 chunk 33 optimal weight: 2.9990 chunk 13 optimal weight: 0.0970 chunk 27 optimal weight: 0.9990 chunk 4 optimal weight: 0.3980 chunk 29 optimal weight: 9.9990 chunk 2 optimal weight: 0.7980 chunk 14 optimal weight: 0.4980 overall best weight: 0.3778 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN A 138 ASN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 2 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4253 r_free = 0.4253 target = 0.182465 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 53)----------------| | r_work = 0.3870 r_free = 0.3870 target = 0.151016 restraints weight = 5337.814| |-----------------------------------------------------------------------------| r_work (start): 0.3867 rms_B_bonded: 1.25 r_work: 0.3765 rms_B_bonded: 2.19 restraints_weight: 0.5000 r_work: 0.3641 rms_B_bonded: 3.74 restraints_weight: 0.2500 r_work (final): 0.3641 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.7010 moved from start: 0.1584 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.003 0.030 4136 Z= 0.155 Angle : 0.554 10.197 5940 Z= 0.314 Chirality : 0.040 0.174 683 Planarity : 0.005 0.041 500 Dihedral : 24.784 179.411 1341 Min Nonbonded Distance : 2.441 Molprobity Statistics. All-atom Clashscore : 7.88 Ramachandran Plot: Outliers : 0.00 % Allowed : 5.41 % Favored : 94.59 % Rotamer: Outliers : 3.64 % Allowed : 17.00 % Favored : 79.35 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.40 (0.52), residues: 296 helix: 0.51 (0.54), residues: 107 sheet: -1.01 (1.12), residues: 26 loop : -0.61 (0.52), residues: 163 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.003 0.000 ARG A 96 TYR 0.010 0.001 TYR A 140 PHE 0.009 0.001 PHE A 137 TRP 0.007 0.002 TRP A 318 HIS 0.001 0.000 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00290 / 0.15 ( 4136) covalent geometry : angle 0.55410 / 0.31 ( 5940) hydrogen bonds : bond 0.05192 / 3.66 ( 109) hydrogen bonds : angle 4.09951 / 2.74 ( 274) *********************** REFINEMENT MACRO_CYCLE 7 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 41 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 9 poor density : 32 time to evaluate : 0.114 Fit side-chains REVERT: A 153 LYS cc_start: 0.7139 (mppt) cc_final: 0.6761 (mptt) REVERT: A 264 TRP cc_start: 0.6326 (OUTLIER) cc_final: 0.6090 (p-90) REVERT: A 314 ARG cc_start: 0.6389 (ttt-90) cc_final: 0.4396 (ttt-90) outliers start: 9 outliers final: 5 residues processed: 37 average time/residue: 0.6550 time to fit residues: 24.9668 Evaluate side-chains 38 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 6 poor density : 32 time to evaluate : 0.064 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 63 VAL Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 264 TRP Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=2.00 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 29 optimal weight: 9.9990 chunk 28 optimal weight: 0.6980 chunk 27 optimal weight: 0.5980 chunk 22 optimal weight: 3.9990 chunk 8 optimal weight: 0.5980 chunk 10 optimal weight: 10.0000 chunk 23 optimal weight: 1.9990 chunk 2 optimal weight: 0.8980 chunk 15 optimal weight: 0.8980 chunk 12 optimal weight: 0.9990 chunk 0 optimal weight: 1.9990 overall best weight: 0.7380 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN A 138 ASN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 2 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4220 r_free = 0.4220 target = 0.179525 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 63)----------------| | r_work = 0.3825 r_free = 0.3825 target = 0.147800 restraints weight = 5215.412| |-----------------------------------------------------------------------------| r_work (start): 0.3822 rms_B_bonded: 1.26 r_work: 0.3720 rms_B_bonded: 2.16 restraints_weight: 0.5000 r_work: 0.3596 rms_B_bonded: 3.67 restraints_weight: 0.2500 r_work (final): 0.3596 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.7068 moved from start: 0.1679 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.005 0.038 4136 Z= 0.228 Angle : 0.660 13.215 5940 Z= 0.358 Chirality : 0.045 0.181 683 Planarity : 0.006 0.056 500 Dihedral : 24.949 179.441 1341 Min Nonbonded Distance : 2.445 Molprobity Statistics. All-atom Clashscore : 9.15 Ramachandran Plot: Outliers : 0.00 % Allowed : 6.76 % Favored : 93.24 % Rotamer: Outliers : 4.05 % Allowed : 17.81 % Favored : 78.14 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.85 (0.51), residues: 296 helix: 0.10 (0.51), residues: 112 sheet: -1.33 (1.12), residues: 26 loop : -0.86 (0.52), residues: 158 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.005 0.001 ARG A 276 TYR 0.011 0.002 TYR A 265 PHE 0.013 0.002 PHE A 137 TRP 0.012 0.002 TRP A 249 HIS 0.002 0.001 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00463 / 0.23 ( 4136) covalent geometry : angle 0.66027 / 0.36 ( 5940) hydrogen bonds : bond 0.06375 / 4.48 ( 109) hydrogen bonds : angle 4.50959 / 3.02 ( 274) *********************** REFINEMENT MACRO_CYCLE 8 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 45 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 10 poor density : 35 time to evaluate : 0.067 Fit side-chains REVERT: A 153 LYS cc_start: 0.7139 (mppt) cc_final: 0.6750 (mptt) REVERT: A 314 ARG cc_start: 0.6493 (ttt-90) cc_final: 0.4512 (ttt-90) outliers start: 10 outliers final: 6 residues processed: 41 average time/residue: 0.6520 time to fit residues: 27.4106 Evaluate side-chains 41 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 6 poor density : 35 time to evaluate : 0.123 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 63 VAL Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 264 TRP Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=1.50 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 3 optimal weight: 0.5980 chunk 1 optimal weight: 0.1980 chunk 27 optimal weight: 0.5980 chunk 6 optimal weight: 0.0980 chunk 24 optimal weight: 0.6980 chunk 22 optimal weight: 0.7980 chunk 10 optimal weight: 10.0000 chunk 15 optimal weight: 0.5980 chunk 9 optimal weight: 0.8980 chunk 4 optimal weight: 0.9990 chunk 23 optimal weight: 0.5980 overall best weight: 0.4180 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN A 138 ASN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 2 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4239 r_free = 0.4239 target = 0.181012 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 54)----------------| | r_work = 0.3848 r_free = 0.3848 target = 0.149409 restraints weight = 5262.832| |-----------------------------------------------------------------------------| r_work (start): 0.3848 rms_B_bonded: 1.23 r_work: 0.3746 rms_B_bonded: 2.18 restraints_weight: 0.5000 r_work: 0.3620 rms_B_bonded: 3.76 restraints_weight: 0.2500 r_work (final): 0.3620 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.7043 moved from start: 0.1725 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.003 0.029 4136 Z= 0.165 Angle : 0.585 10.624 5940 Z= 0.326 Chirality : 0.041 0.162 683 Planarity : 0.006 0.040 500 Dihedral : 24.865 179.507 1341 Min Nonbonded Distance : 2.442 Molprobity Statistics. All-atom Clashscore : 8.45 Ramachandran Plot: Outliers : 0.00 % Allowed : 5.07 % Favored : 94.93 % Rotamer: Outliers : 3.64 % Allowed : 19.03 % Favored : 77.33 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.73 (0.51), residues: 296 helix: 0.19 (0.51), residues: 112 sheet: -1.35 (1.10), residues: 26 loop : -0.77 (0.53), residues: 158 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.004 0.000 ARG A 115 TYR 0.009 0.002 TYR A 140 PHE 0.010 0.001 PHE A 137 TRP 0.011 0.002 TRP A 318 HIS 0.004 0.001 HIS A 253 Details of bonding type rmsd/Z covalent geometry : bond 0.00319 / 0.17 ( 4136) covalent geometry : angle 0.58485 / 0.33 ( 5940) hydrogen bonds : bond 0.05126 / 3.75 ( 109) hydrogen bonds : angle 4.31721 / 2.88 ( 274) *********************** REFINEMENT MACRO_CYCLE 9 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 41 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 9 poor density : 32 time to evaluate : 0.087 Fit side-chains REVERT: A 153 LYS cc_start: 0.7164 (mppt) cc_final: 0.6766 (mptt) REVERT: A 264 TRP cc_start: 0.6413 (OUTLIER) cc_final: 0.6112 (p-90) REVERT: A 314 ARG cc_start: 0.6434 (ttt-90) cc_final: 0.4589 (ttt-90) outliers start: 9 outliers final: 6 residues processed: 37 average time/residue: 0.6607 time to fit residues: 25.1696 Evaluate side-chains 39 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 7 poor density : 32 time to evaluate : 0.104 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 63 VAL Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 264 TRP Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 305 ILE Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=1.00 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 23 optimal weight: 0.9980 chunk 30 optimal weight: 4.9990 chunk 16 optimal weight: 1.9990 chunk 25 optimal weight: 0.5980 chunk 6 optimal weight: 0.5980 chunk 3 optimal weight: 0.9980 chunk 17 optimal weight: 0.9990 chunk 31 optimal weight: 2.9990 chunk 11 optimal weight: 30.0000 chunk 18 optimal weight: 0.5980 chunk 13 optimal weight: 0.9980 overall best weight: 0.7580 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN A 138 ASN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 2 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4209 r_free = 0.4209 target = 0.178582 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 60)----------------| | r_work = 0.3816 r_free = 0.3816 target = 0.147087 restraints weight = 5200.738| |-----------------------------------------------------------------------------| r_work (start): 0.3813 rms_B_bonded: 1.25 r_work: 0.3710 rms_B_bonded: 2.16 restraints_weight: 0.5000 r_work: 0.3586 rms_B_bonded: 3.67 restraints_weight: 0.2500 r_work (final): 0.3586 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.7082 moved from start: 0.1821 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.005 0.043 4136 Z= 0.234 Angle : 0.680 13.486 5940 Z= 0.366 Chirality : 0.045 0.187 683 Planarity : 0.006 0.067 500 Dihedral : 25.011 179.106 1341 Min Nonbonded Distance : 2.448 Molprobity Statistics. All-atom Clashscore : 9.99 Ramachandran Plot: Outliers : 0.00 % Allowed : 6.76 % Favored : 93.24 % Rotamer: Outliers : 3.64 % Allowed : 19.03 % Favored : 77.33 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -1.08 (0.50), residues: 296 helix: -0.05 (0.51), residues: 112 sheet: -1.68 (1.11), residues: 26 loop : -0.98 (0.51), residues: 158 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.006 0.001 ARG A 276 TYR 0.009 0.002 TYR A 201 PHE 0.014 0.002 PHE A 137 TRP 0.013 0.002 TRP A 249 HIS 0.002 0.001 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00477 / 0.23 ( 4136) covalent geometry : angle 0.67952 / 0.37 ( 5940) hydrogen bonds : bond 0.06632 / 4.71 ( 109) hydrogen bonds : angle 4.62963 / 3.11 ( 274) ********************** REFINEMENT MACRO_CYCLE 10 OF 10 ************************ ------------------------------------------------------------------------------- Update Rama plot phi/psi targets (oldfield only) ************************************************ 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Re-set Ramachandran plot restraints *********************************** favored: oldfield allowed: oldfield outlier: oldfield 592 Ramachandran restraints generated. 296 Oldfield, 0 Emsley, 296 emsley8k and 0 Phi/Psi/2. ------------------------------------------------------------------------------- Optimize residue side-chains **************************** Evaluate side-chains 46 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 9 poor density : 37 time to evaluate : 0.104 Fit side-chains REVERT: A 153 LYS cc_start: 0.7136 (mppt) cc_final: 0.6750 (mptt) REVERT: A 314 ARG cc_start: 0.6502 (ttt-90) cc_final: 0.4406 (ttt-90) outliers start: 9 outliers final: 6 residues processed: 42 average time/residue: 0.6475 time to fit residues: 27.9420 Evaluate side-chains 42 residues out of total 249 (non-[ALA,GLY,PRO]) need to be fit. rotamer outliers: 6 poor density : 36 time to evaluate : 0.109 ------------------------------------------------------------------------------- Set rotamer restraints ********************** Chi-restraints excluded: chain A residue 63 VAL Chi-restraints excluded: chain A residue 77 ILE Chi-restraints excluded: chain A residue 92 THR Chi-restraints excluded: chain A residue 264 TRP Chi-restraints excluded: chain A residue 302 LYS Chi-restraints excluded: chain A residue 336 SER Rotamers are restrained with sigma=1.00 ------------------------------------------------------------------------------- XYZ refinement ************** Weight determination summary: number of chunks: 35 random chunks: chunk 33 optimal weight: 2.9990 chunk 8 optimal weight: 0.6980 chunk 30 optimal weight: 2.9990 chunk 16 optimal weight: 0.6980 chunk 25 optimal weight: 0.2980 chunk 29 optimal weight: 9.9990 chunk 10 optimal weight: 10.0000 chunk 15 optimal weight: 0.8980 chunk 20 optimal weight: 0.0050 chunk 22 optimal weight: 0.5980 chunk 5 optimal weight: 0.0980 overall best weight: 0.3394 ------------------------------------------------------------------------------- NQH flips ********* Analyzing N/Q/H residues for possible flip corrections... A 127 GLN A 138 ASN ** A 203 GLN ** both conformations clash, **PLEASE CHECK MANUALLY** Total number of N/Q/H flips: 2 ------------------------------------------------------------------------------- ADP refinement ************** |-group b-factor refinement (macro cycle = 0; iterations = 0)-----------------| | r_work = 0.4240 r_free = 0.4240 target = 0.181054 restraints weight = None | |-----------------------------------------------------------------------------| |-group b-factor refinement (macro cycle = 1; iterations = 62)----------------| | r_work = 0.3852 r_free = 0.3852 target = 0.149630 restraints weight = 5224.916| |-----------------------------------------------------------------------------| r_work (start): 0.3851 rms_B_bonded: 1.22 r_work: 0.3748 rms_B_bonded: 2.16 restraints_weight: 0.5000 r_work: 0.3626 rms_B_bonded: 3.69 restraints_weight: 0.2500 r_work (final): 0.3626 ------------------------------------------------------------------------------- Occupancy refinement ******************** ------------------------------------------------------------------------------- Overall statistics ****************** model-to-map fit, CC_mask: 0.7032 moved from start: 0.1836 Geometry Restraints Library: GeoStd + Monomer Library + CDL v1.2 Deviations from Ideal Values - rmsd, rmsZ for bonds and angles. Bond : 0.003 0.029 4136 Z= 0.156 Angle : 0.570 9.797 5940 Z= 0.322 Chirality : 0.040 0.149 683 Planarity : 0.005 0.039 500 Dihedral : 24.831 179.933 1341 Min Nonbonded Distance : 2.443 Molprobity Statistics. All-atom Clashscore : 9.43 Ramachandran Plot: Outliers : 0.00 % Allowed : 4.39 % Favored : 95.61 % Rotamer: Outliers : 2.83 % Allowed : 19.84 % Favored : 77.33 % Cbeta Deviations : 0.00 % Peptide Plane: Cis-proline : 0.00 % Cis-general : 0.00 % Twisted Proline : 0.00 % Twisted General : 0.00 % Rama-Z values with (uncertainties): Interpretation: poor |Rama-Z| > 3; suspicious 2 < |Rama-Z| < 3; good |Rama-Z| < 2. Scores below are scaled independently, so they are not related in a simple way. whole: -0.82 (0.51), residues: 296 helix: 0.17 (0.52), residues: 112 sheet: -1.66 (1.09), residues: 26 loop : -0.81 (0.53), residues: 158 Max deviation from planes: Type MaxDev MeanDev LineInFile ARG 0.004 0.000 ARG A 262 TYR 0.009 0.001 TYR A 140 PHE 0.010 0.002 PHE A 137 TRP 0.012 0.002 TRP A 318 HIS 0.001 0.000 HIS A 135 Details of bonding type rmsd/Z covalent geometry : bond 0.00292 / 0.16 ( 4136) covalent geometry : angle 0.57026 / 0.32 ( 5940) hydrogen bonds : bond 0.04873 / 3.62 ( 109) hydrogen bonds : angle 4.31966 / 2.89 ( 274) Origin is already at (0, 0, 0), no shifts will be applied =============================================================================== Job complete usr+sys time: 1676.76 seconds wall clock time: 29 minutes 12.73 seconds (1752.73 seconds total)