++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE - PHASING AND DENSITY MODIFICATION - Version 2019/1 + + Copyright (c) George M. Sheldrick and Isabel Uson 2001-19 + + Started at 12:23:26 on 15 Dec 2020 + ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ Please cite: I. Uson & G.M. Sheldrick (2018), "An introduction to experimental phasing of macromolecules illustrated by SHELX; new autotracing features" Acta Cryst. D74, 106-116 (Open Access) if SHELXE proves useful. Command line parameters: P4_visc-standard P4_visc-standard_fa -a5 -h -s0.5 -m5 Cell, symmetry and heavy atoms from: P4_visc-standard_fa.res FA and alpha from P4_visc-standard_fa.hkl Native data from P4_visc-standard.hkl Listing output to P4_visc-standard.lst Phases output to P4_visc-standard.phs Revised heavy atom sites output to P4_visc-standard.hat Revised heavy atom phases output to P4_visc-standard.pha Poly-Ala trace output to P4_visc-standard.pdb Summary of parameters to be employed: -a 5 global autotracing cycles -b 5.0 extra B for revised heavy atom sites -B unset just build single beta-strands -c 0.400 fraction of pixels in crossover region -d 0.000 high resolution limit to be applied to input data -D unset do not fuse disulfides for NCS -e unset fill in missing data up to maximum resolution + 0.2 Ang. -f unset read intensity not F from native .hkl file -F 0.800 fractional weight for phases from previous global cycle -g 1.100 solvent gamma flipping factor -G 0.700 FOM threshold for initial tripeptides and chain building -h heavy atoms present in native - use all with occ > 0.2 -i unset no structure inversion -k 4.5 minimum height/sigma for revised heavy atom sites -l 2 space for 2000000 reflections -L 6 minimum number of residues/chain (if more than 3 chains) -m 5 cycles of density modification -n unset do not apply NCS -p unset no phosphate search -q unset no alpha-helix search -r 3.00 map resolution (multiplies maximum indices) -s 0.500 solvent fraction -S 2.42 radius of sphere of influence -t 1.00 time for initial searches (-t6 or more if difficult) -u 500 MB allocatable memory for fragment optimization -v unset density sharpening factor dependent on resolution -w 0.200 weight for experimental phases after cycle 1 -x unset no phase and trace diagnostics -z unset do not optimize heavy atoms -z 0 read heavy atoms from .res Space group: P 4 Allowed origin shift code: 7 21 atoms read from file P4_visc-standard_fa.res Trimmed to 19 atoms with occupancy > 0.2 14490 Reflections read from file P4_visc-standard_fa.hkl 22220 Reflections read from file P4_visc-standard.hkl 22220 Unique data, highest resolution = 1.702 Angstroms Anisotropic scaling: intensities multiplied by 0.000220h^2 +0.000220k^2 -0.000674l^2 +0.000000kl +0.000000hl +0.000000hk 9 Reflections with d > 1.902 and 0 in range 1.902 > d > 1.702 added Density sharpening factor set to 1.20 Fourier grid = 128 x 128 x 82 0.000 <= z <= 1.000 92 Point spherical net set up with radius 2.42A 32 Extra Fourier layers will be generated <|E^2-1|> = 0.760 Overall CC between Eobs (from delF) and Ecalc (from heavy atoms) = 9.72% = 0.088 for phases from phiT = phiA + alpha = 0.142 after including heavy atoms = 0.126, Contrast = 0.020, Connect. = 0.514 for dens.mod. cycle 1 = 0.148, Contrast = 0.085, Connect. = 0.591 for dens.mod. cycle 2 = 0.169, Contrast = 0.115, Connect. = 0.617 for dens.mod. cycle 3 = 0.186, Contrast = 0.117, Connect. = 0.608 for dens.mod. cycle 4 = 0.198, Contrast = 0.118, Connect. = 0.598 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 17 heavy atoms with Occ*Z > 4.80 added to NOGO map 1683 peaks > 0.5 sigma used to seed fragment search Space for about 203 unique residues taking solvent into account 119 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 7 2.290 1.437 0.198 0.246 0.667 0.667 CB 1.093 B: 6 2.768 1.325 0.267 0.393 0.800 0.600 N 1.033 C: 10 3.598 1.388 0.402 0.339 0.889 0.667 CB 0.826 D: 7 2.941 1.276 0.529 0.403 0.667 0.333 N 1.022 6 1.945 1.340 0.051 0.447 0.800 0.600 CB 0.815 ? E: 8 2.696 1.249 0.305 0.325 0.857 0.571 CB 0.884 F: 6 4.635 1.181 0.682 0.606 1.000 0.400 CB 0.944 38 residues left after pruning, divided into chains as follows: A: 7 B: 10 C: 7 D: 8 E: 6 CC for partial structure against native data = 2.46 % ------------------------------------------------------------------------------ Global autotracing cycle 2 = 0.286, Contrast = 0.174, Connect. = 0.532 for dens.mod. cycle 1 = 0.287, Contrast = 0.183, Connect. = 0.475 for dens.mod. cycle 2 = 0.287, Contrast = 0.189, Connect. = 0.489 for dens.mod. cycle 3 = 0.287, Contrast = 0.191, Connect. = 0.502 for dens.mod. cycle 4 = 0.287, Contrast = 0.193, Connect. = 0.512 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 17 heavy atoms with Occ*Z > 4.80 added to NOGO map 2225 peaks > 0.5 sigma used to seed fragment search Space for about 203 unique residues taking solvent into account 134 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 20 11.076 2.225 0.283 0.264 0.684 0.421 CB 1.703 B: 9 8.229 2.139 0.095 0.551 0.750 0.375 CB 1.576 C: 11 7.198 2.247 0.091 0.240 0.700 0.400 CA 1.793 D: 9 5.143 2.069 -0.018 0.435 0.625 0.375 CB 1.532 6 0.662 1.851 0.151 0.332 0.400 0.400 CB 0.418 ? E: 13 6.552 2.169 0.021 0.220 0.750 0.583 CB 1.581 13 residues pruned to eliminate duplicates F: 10 9.860 2.145 0.192 0.355 0.778 0.667 CB 1.933 10 residues pruned to eliminate duplicates Using tripeptides from previous cycle as seeds G: 14 13.609 2.472 0.051 0.284 0.846 0.462 CB 2.191 20 residues pruned to eliminate duplicates H: 6 3.643 1.840 0.352 0.358 0.800 0.800 CB 0.955 49 residues left after pruning, divided into chains as follows: A: 9 B: 11 C: 9 D: 14 E: 6 CC for partial structure against native data = 3.87 % ------------------------------------------------------------------------------ Global autotracing cycle 3 = 0.287, Contrast = 0.220, Connect. = 0.510 for dens.mod. cycle 1 = 0.287, Contrast = 0.228, Connect. = 0.496 for dens.mod. cycle 2 = 0.287, Contrast = 0.232, Connect. = 0.508 for dens.mod. cycle 3 = 0.287, Contrast = 0.232, Connect. = 0.519 for dens.mod. cycle 4 = 0.287, Contrast = 0.232, Connect. = 0.528 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 17 heavy atoms with Occ*Z > 4.80 added to NOGO map 2161 peaks > 0.5 sigma used to seed fragment search Space for about 203 unique residues taking solvent into account 141 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 16 13.880 2.562 0.058 0.249 0.800 0.467 CB 2.207 B: 24 12.267 2.297 0.209 0.261 0.739 0.348 CB 1.638 C: 12 11.467 2.379 0.090 0.379 0.818 0.273 CB 1.853 D: 6 4.172 2.225 0.363 0.373 0.600 0.600 CB 1.174 E: 8 2.530 1.275 0.184 0.455 0.857 0.429 CB 0.773 Using tripeptides from previous cycle as seeds 56 residues left after pruning, divided into chains as follows: A: 16 B: 8 C: 14 D: 12 E: 6 CC for partial structure against native data = 5.00 % ------------------------------------------------------------------------------ Global autotracing cycle 4 = 0.287, Contrast = 0.229, Connect. = 0.513 for dens.mod. cycle 1 = 0.287, Contrast = 0.235, Connect. = 0.505 for dens.mod. cycle 2 = 0.287, Contrast = 0.239, Connect. = 0.516 for dens.mod. cycle 3 = 0.287, Contrast = 0.239, Connect. = 0.525 for dens.mod. cycle 4 = 0.287, Contrast = 0.238, Connect. = 0.533 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 17 heavy atoms with Occ*Z > 4.80 added to NOGO map 2121 peaks > 0.5 sigma used to seed fragment search Space for about 203 unique residues taking solvent into account 149 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 18 18.250 2.627 0.167 0.271 0.824 0.471 CA 2.275 B: 8 10.416 2.780 -0.024 0.372 0.714 0.429 CB 2.330 C: 11 11.612 2.464 0.141 0.352 0.800 0.600 CB 1.915 D: 8 9.687 2.387 0.230 0.470 0.714 0.571 CB 1.801 E: 15 10.973 2.280 0.098 0.383 0.714 0.214 CB 1.858 Using tripeptides from previous cycle as seeds F: 18 9.143 2.359 0.090 0.225 0.706 0.294 CB 1.684 9 residues pruned to eliminate duplicates 64 residues left after pruning, divided into chains as follows: A: 18 B: 11 C: 8 D: 15 E: 12 CC for partial structure against native data = 4.82 % ------------------------------------------------------------------------------ Global autotracing cycle 5 = 0.287, Contrast = 0.229, Connect. = 0.518 for dens.mod. cycle 1 = 0.287, Contrast = 0.233, Connect. = 0.507 for dens.mod. cycle 2 = 0.287, Contrast = 0.238, Connect. = 0.518 for dens.mod. cycle 3 = 0.287, Contrast = 0.239, Connect. = 0.527 for dens.mod. cycle 4 = 0.287, Contrast = 0.238, Connect. = 0.535 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 17 heavy atoms with Occ*Z > 4.80 added to NOGO map 2138 peaks > 0.5 sigma used to seed fragment search Space for about 203 unique residues taking solvent into account 166 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 15 16.999 2.767 0.091 0.230 0.857 0.500 CA 2.407 B: 13 7.182 2.401 -0.068 0.292 0.667 0.583 CA 1.753 C: 17 11.202 2.100 0.305 0.397 0.688 0.250 CB 1.656 D: 18 13.104 2.373 0.102 0.303 0.765 0.118 CB 1.979 14 residues pruned to eliminate duplicates E: 10 13.305 2.252 0.354 0.483 0.778 0.556 CB 1.903 F: 15 10.856 2.549 0.052 0.258 0.714 0.286 CB 1.997 G: 6 3.115 1.445 0.448 0.338 0.800 0.600 N 0.993 Using tripeptides from previous cycle as seeds 72 residues left after pruning, divided into chains as follows: A: 15 B: 10 C: 18 D: 10 E: 13 F: 6 CC for partial structure against native data = 7.19 % ------------------------------------------------------------------------------ Global autotracing cycle 6 Phases from autotracing cycle 5 used as input for final density modification = 0.287, Contrast = 0.239, Connect. = 0.520 for dens.mod. cycle 1 = 0.287, Contrast = 0.242, Connect. = 0.512 for dens.mod. cycle 2 = 0.287, Contrast = 0.248, Connect. = 0.523 for dens.mod. cycle 3 = 0.287, Contrast = 0.248, Connect. = 0.531 for dens.mod. cycle 4 = 0.287, Contrast = 0.247, Connect. = 0.540 for dens.mod. cycle 5 Estimated mean FOM and mapCC as a function of resolution d inf - 3.71 - 2.94 - 2.56 - 2.32 - 2.16 - 2.03 - 1.92 - 1.84 - 1.77 - 1.71 0.212 0.212 0.244 0.276 0.232 0.209 0.262 0.278 0.217 0.209 0.287 0.273 0.305 0.354 0.309 0.293 0.374 0.414 0.335 0.332 N 2232 2224 2250 2277 2141 2250 2426 2134 2190 2096 Estimated mean FOM = 0.236 Pseudo-free CC = 28.20 % Anomalous density (in sigma units) at initial heavy atom sites Site x y z occ*Z density 1 0.3863 0.8584 0.6379 16.0000 32.38 2 0.4778 0.8443 0.8342 14.0000 27.55 3 0.1669 0.6732 0.3183 13.7952 25.83 4 0.6207 0.8341 0.8381 12.3808 23.19 5 0.6860 0.8146 0.6632 11.2240 20.99 6 0.3515 0.8414 0.2332 11.1744 21.71 7 0.5000 0.5000 0.6514 2.6160 21.03 8 0.3474 0.8555 0.4138 9.7888 17.90 9 0.0000 1.0000 0.4308 2.2272 17.77 10 0.5418 0.8660 0.9089 8.5568 16.44 11 0.7719 0.7472 0.8491 8.0848 16.81 12 0.3824 0.9533 0.4368 8.0288 16.78 13 0.4427 0.8364 0.1295 7.1888 13.43 14 0.0388 0.8489 0.0957 7.1840 12.37 15 0.4000 0.8854 0.6999 6.9600 13.39 16 0.3988 0.9122 0.2535 6.4736 12.92 17 0.5098 0.6460 0.3248 5.5072 10.14 18 0.4613 0.6562 0.8192 5.4336 9.02 19 0.4143 0.6888 0.9127 5.1968 10.92 Site x y z h(sig) near old near new 1 0.3859 0.8585 0.6379 32.4 1/0.03 14/3.51 8/10.89 2/11.10 11/11.36 2 0.4776 0.8447 0.8344 27.6 2/0.03 12/5.65 14/8.58 4/9.41 1/11.10 3 0.1668 0.6732 0.3183 25.8 3/0.01 11/15.55 18/16.33 7/16.35 5/16.93 4 0.6204 0.8342 0.8392 23.2 4/0.06 22/2.16 12/6.51 2/9.41 7/9.45 5 0.3520 0.8414 0.2328 21.7 6/0.04 15/5.61 13/7.79 8/8.57 11/12.26 6 0.5000 0.5000 0.6534 21.1 7/0.09 21/7.59 21/7.59 21/7.59 21/7.59 7 0.6862 0.8141 0.6634 21.0 5/0.04 22/8.03 4/9.45 10/11.35 12/15.39 8 0.3474 0.8560 0.4132 17.9 8/0.04 11/6.90 5/8.57 15/9.00 1/10.89 9 0.0000 1.0000 0.4324 17.8 9/0.08 16/18.90 16/18.90 16/18.90 16/18.90 10 0.7725 0.7465 0.8494 16.9 11/0.07 22/9.65 7/11.35 4/11.55 17/12.36 11 0.3831 0.9532 0.4363 16.8 12/0.05 8/6.90 15/9.08 1/11.36 5/12.26 12 0.5413 0.8666 0.9090 16.5 10/0.05 2/5.65 4/6.51 22/8.62 13/12.35 13 0.4438 0.8355 0.1287 13.5 13/0.10 5/7.79 15/8.31 12/12.35 2/14.07 14 0.3988 0.8845 0.7003 13.5 15/0.10 1/3.51 2/8.58 11/13.28 12/13.64 15 0.3978 0.9117 0.2539 13.0 16/0.07 5/5.61 13/8.31 8/9.00 11/9.08 16 0.0387 0.8490 0.0958 12.4 14/0.02 16/14.49 16/14.49 3/17.74 9/18.90 17 0.4151 0.6885 0.9146 11.0 19/0.11 19/5.85 2/11.69 10/12.36 13/14.10 18 0.5109 0.6453 0.3266 10.2 17/0.12 18/13.55 20/14.57 20/14.57 20/14.5 19 0.4633 0.6552 0.8211 9.2 18/0.17 17/5.85 21/10.49 21/10.49 21/10.49 20 0.5000 0.5000 0.0937 -5.9 17/14.54 21/13.18 21/13.18 21/13.18 21/13. 21 0.5000 0.5000 0.8142 -5.1 7/7.68 6/7.59 6/7.59 6/7.59 6/7.59 22 0.6506 0.8264 0.8253 -4.7 4/2.12 4/2.16 7/8.03 12/8.62 10/9.65 Best trace (cycle 5 with CC 7.19%) was saved as P4_visc-standard.pdb ============================================================================== CPU times required in seconds ----------------------------- 0.5 - Setup, data input and phasing 3.3 - FFTs and peak-searches 4.0 - Sphere of influence 0.1 - Rest of density modification 0.0 - Alpha-helix search 124.8 - Tripeptide search 24.3 - Chain tracing 0.0 - NCS analysis 2.9 - B-value refinement for trace 0.1 - Rest of tracing 0.0 - Comparison with known structure ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE finished at 12:26:06 Total time: 160.00 secs + ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++