++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE - PHASING AND DENSITY MODIFICATION - Version 2019/1 + + Copyright (c) George M. Sheldrick and Isabel Uson 2001-19 + + Started at 12:03:42 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: P2221_pse14 P2221_pse14_fa -i -h -a5 -s0.45 -z -e -m5 Cell, symmetry and heavy atoms from: P2221_pse14_fa.res FA and alpha from P2221_pse14_fa.hkl Native data from P2221_pse14.hkl Listing output to P2221_pse14_i.lst Phases output to P2221_pse14_i.phs Revised heavy atom sites output to P2221_pse14_i.hat Revised heavy atom phases output to P2221_pse14_i.pha Poly-Ala trace output to P2221_pse14_i.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 invert structure (and space group) -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 0.450 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 refine an unlimited number of heavy atoms Space group: P 2 2 21 Allowed origin shift code: 4 14 atoms read from file P2221_pse14_fa.res Trimmed to 13 atoms with occupancy > 0.2 11695 Reflections read from file P2221_pse14_fa.hkl 21043 Reflections read from file P2221_pse14.hkl 21024 Unique data, highest resolution = 2.289 Angstroms Anisotropic scaling: intensities multiplied by 0.003997h^2 -0.000434k^2 -0.000496l^2 +0.000000kl +0.000000hl +0.000000hk 26 Reflections with d > 2.489 and 0 in range 2.489 > d > 2.289 added Density sharpening factor set to 0.32 Fourier grid = 128 x 128 x 32 0.000 <= z <= 0.250 92 Point spherical net set up with radius 2.42A 24 Extra Fourier layers will be generated <|E^2-1|> = 0.781 ** Atom coordinates inverted ** 13 potential heavy atoms found Substructure optimization LOR = lowest occupancy retained, HOR = highest occupancy rejected Nats CC(HA) LOR HOR 13 19.48% 0.305 Overall CC between Eobs (from delF) and Ecalc (from heavy atoms) = 19.42% = 0.125 for phases from phiT = phiA + alpha = 0.200 after including heavy atoms = 0.183, Contrast = 0.036, Connect. = 0.525 for dens.mod. cycle 1 = 0.203, Contrast = 0.116, Connect. = 0.591 for dens.mod. cycle 2 = 0.217, Contrast = 0.163, Connect. = 0.614 for dens.mod. cycle 3 = 0.228, Contrast = 0.199, Connect. = 0.630 for dens.mod. cycle 4 = 0.238, Contrast = 0.228, Connect. = 0.644 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 13 heavy atoms with Occ*Z > 0.30 added to NOGO map 1964 peaks > 0.5 sigma used to seed fragment search Space for about 498 unique residues taking solvent into account 261 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 8 4.070 1.251 0.902 0.757 1.000 0.714 CB 0.515 B: 6 4.991 1.377 0.944 0.505 0.800 0.400 CB 1.037 C: 6 4.456 1.435 0.960 0.322 0.800 0.600 CB 1.077 D: 12 3.705 1.502 0.262 0.182 0.909 0.273 CB 0.951 E: 7 2.153 1.458 0.356 0.256 0.500 0.167 CB 1.175 F: 9 3.579 1.389 -0.051 0.461 0.875 0.375 CB 1.142 7 1.747 1.588 -0.154 0.248 0.667 0.667 CB 1.063 ? G: 9 2.245 1.391 0.274 0.210 0.750 0.375 CB 0.840 H: 6 3.693 1.540 0.248 0.297 0.800 0.800 CB 1.347 6 1.778 1.201 0.273 0.291 0.800 0.400 CB 0.822 ? I: 8 3.003 1.140 0.549 0.532 0.714 0.429 CB 0.872 J: 7 2.312 1.413 0.190 0.241 0.667 0.500 CB 1.137 K: 6 2.515 1.386 0.775 0.543 0.600 0.400 CB 0.731 L: 11 3.326 1.466 0.791 0.145 0.600 0.300 N 1.035 7 1.624 1.142 0.254 0.481 1.000 0.500 CB 0.472 ? M: 8 2.876 1.367 0.261 0.338 0.857 0.714 CB 0.878 N: 6 5.163 1.502 0.576 0.383 0.800 0.400 CB 1.381 O: 6 2.130 1.111 0.379 0.415 0.800 0.600 CB 0.850 P: 10 3.742 1.595 0.297 0.252 0.667 0.444 CB 1.203 6 0.770 1.231 0.368 0.298 0.400 0.400 CB 0.640 ? Q: 9 2.410 1.375 0.669 0.187 0.625 0.375 CB 0.865 R: 6 2.428 1.296 0.069 0.538 0.800 0.400 CB 0.944 7 1.288 1.013 0.266 0.256 0.833 0.500 N 0.651 ? S: 6 2.211 1.247 0.694 0.297 0.600 0.600 O 0.979 82 residues left after pruning, divided into chains as follows: A: 8 B: 6 C: 9 D: 9 E: 6 F: 6 G: 6 H: 6 I: 8 J: 6 K: 6 L: 6 CC for partial structure against native data = 7.11 % ------------------------------------------------------------------------------ Global autotracing cycle 2 = 0.281, Contrast = 0.231, Connect. = 0.593 for dens.mod. cycle 1 = 0.281, Contrast = 0.254, Connect. = 0.577 for dens.mod. cycle 2 = 0.281, Contrast = 0.298, Connect. = 0.619 for dens.mod. cycle 3 = 0.281, Contrast = 0.324, Connect. = 0.641 for dens.mod. cycle 4 = 0.281, Contrast = 0.343, Connect. = 0.656 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 13 heavy atoms with Occ*Z > 0.30 added to NOGO map 2256 peaks > 0.5 sigma used to seed fragment search Space for about 498 unique residues taking solvent into account 234 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 6 6.816 1.832 0.824 0.597 0.800 0.600 CB 1.039 B: 12 19.051 1.875 0.721 0.842 1.000 0.818 CB 1.327 C: 10 2.035 1.559 0.088 0.288 0.667 0.333 CB 0.762 D: 6 4.509 1.765 -0.055 0.619 0.800 0.600 N 1.350 E: 7 8.987 1.816 0.860 0.703 0.833 0.667 CB 1.083 F: 10 3.861 1.713 0.339 0.216 0.667 0.444 CB 1.175 G: 8 10.818 2.202 0.091 0.513 0.857 0.286 CA 1.960 H: 8 3.431 1.664 0.510 0.295 0.714 0.429 CB 0.909 I: 12 5.441 1.617 0.143 0.345 0.909 0.182 CB 1.155 J: 6 4.223 1.533 0.209 0.550 0.800 0.600 CB 1.213 K: 6 3.802 1.629 0.737 0.293 0.600 0.400 N 1.263 L: 6 2.023 1.132 0.499 0.294 1.000 0.400 O 0.672 M: 8 5.009 1.648 0.248 0.346 0.857 0.429 CB 1.270 N: 9 2.051 1.559 0.216 0.214 0.875 0.375 CB 0.612 Using tripeptides from previous cycle as seeds O: 10 3.436 1.760 -0.036 0.298 0.889 0.444 CB 0.952 7 residues pruned to eliminate duplicates 88 residues left after pruning, divided into chains as follows: A: 12 B: 10 C: 7 D: 9 E: 8 F: 8 G: 11 H: 6 I: 8 J: 9 CC for partial structure against native data = 9.06 % ------------------------------------------------------------------------------ Global autotracing cycle 3 = 0.281, Contrast = 0.293, Connect. = 0.606 for dens.mod. cycle 1 = 0.281, Contrast = 0.324, Connect. = 0.600 for dens.mod. cycle 2 = 0.281, Contrast = 0.368, Connect. = 0.642 for dens.mod. cycle 3 = 0.281, Contrast = 0.387, Connect. = 0.662 for dens.mod. cycle 4 = 0.281, Contrast = 0.406, Connect. = 0.678 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 13 heavy atoms with Occ*Z > 0.30 added to NOGO map 2122 peaks > 0.5 sigma used to seed fragment search Space for about 498 unique residues taking solvent into account 216 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 9 5.559 2.169 0.419 0.239 0.625 0.375 CB 1.382 B: 9 11.137 2.026 0.501 0.731 0.750 0.750 CB 1.403 C: 19 8.423 1.840 0.119 0.290 0.833 0.389 N 1.465 D: 7 11.284 1.880 0.185 0.823 1.000 0.833 N 1.563 E: 8 5.065 1.846 0.462 0.252 0.571 0.571 CA 1.650 F: 7 9.195 2.038 0.330 0.501 0.833 0.833 CB 1.661 G: 8 10.444 1.972 0.485 0.448 0.857 0.429 CB 1.658 H: 7 5.404 1.771 0.377 0.528 0.833 0.333 CB 1.057 I: 7 2.603 1.537 0.589 0.508 1.000 0.500 CB 0.431 J: 10 3.753 1.408 0.056 0.316 1.000 0.556 CB 1.031 6 1.828 1.278 0.257 0.298 0.600 0.400 N 1.063 ? K: 11 3.598 1.610 0.213 0.366 0.500 0.400 CB 1.345 Using tripeptides from previous cycle as seeds L: 10 5.010 1.780 0.026 0.303 0.889 0.444 CB 1.282 10 residues pruned to eliminate duplicates 83 residues left after pruning, divided into chains as follows: A: 8 B: 9 C: 19 D: 7 E: 8 F: 7 G: 8 H: 7 I: 10 CC for partial structure against native data = 8.86 % ------------------------------------------------------------------------------ Global autotracing cycle 4 = 0.281, Contrast = 0.317, Connect. = 0.612 for dens.mod. cycle 1 = 0.281, Contrast = 0.361, Connect. = 0.615 for dens.mod. cycle 2 = 0.281, Contrast = 0.409, Connect. = 0.659 for dens.mod. cycle 3 = 0.281, Contrast = 0.425, Connect. = 0.676 for dens.mod. cycle 4 = 0.281, Contrast = 0.438, Connect. = 0.689 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 13 heavy atoms with Occ*Z > 0.30 added to NOGO map 2056 peaks > 0.5 sigma used to seed fragment search Space for about 498 unique residues taking solvent into account 231 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 13 10.258 1.931 0.171 0.770 0.750 0.583 CB 1.375 B: 6 7.073 2.009 0.402 0.291 1.000 0.600 CB 1.420 C: 24 11.916 1.977 0.247 0.268 0.783 0.435 CB 1.676 D: 8 14.059 2.015 0.530 0.749 1.000 0.714 CB 1.380 E: 7 9.671 2.255 0.237 0.436 0.833 0.333 CB 1.815 F: 11 8.891 2.047 0.413 0.338 0.900 0.600 CB 1.290 G: 6 6.706 1.738 0.485 0.607 0.800 0.800 N 1.312 H: 8 5.235 1.558 0.617 0.331 0.857 0.571 CB 1.103 I: 8 5.446 1.921 0.076 0.409 0.857 0.714 CB 1.279 J: 8 3.091 1.433 0.476 0.408 0.571 0.429 N 1.064 K: 10 5.453 1.511 0.700 0.277 0.889 0.444 N 1.024 L: 6 3.081 1.557 -0.080 0.416 1.000 0.400 CB 1.050 Using tripeptides from previous cycle as seeds 84 residues left after pruning, divided into chains as follows: A: 13 B: 24 C: 8 D: 6 E: 11 F: 6 G: 8 H: 8 CC for partial structure against native data = 9.23 % ------------------------------------------------------------------------------ Global autotracing cycle 5 = 0.281, Contrast = 0.331, Connect. = 0.614 for dens.mod. cycle 1 = 0.281, Contrast = 0.375, Connect. = 0.619 for dens.mod. cycle 2 = 0.281, Contrast = 0.427, Connect. = 0.664 for dens.mod. cycle 3 = 0.281, Contrast = 0.442, Connect. = 0.683 for dens.mod. cycle 4 = 0.281, Contrast = 0.452, Connect. = 0.695 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 13 heavy atoms with Occ*Z > 0.30 added to NOGO map 2000 peaks > 0.5 sigma used to seed fragment search Space for about 498 unique residues taking solvent into account 227 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 11 16.402 2.027 0.354 0.772 1.000 0.700 CB 1.485 B: 8 6.765 2.208 0.018 0.350 0.714 0.571 CA 1.873 C: 25 13.623 2.027 0.247 0.311 0.833 0.333 CB 1.629 D: 9 13.596 1.859 0.583 0.754 0.875 0.750 CA 1.489 E: 9 5.112 1.984 0.176 0.375 0.625 0.500 CB 1.417 F: 12 11.032 1.960 0.460 0.431 0.818 0.455 CB 1.526 G: 7 4.404 1.695 0.579 0.366 0.833 0.333 CB 0.931 H: 6 2.327 1.638 0.205 0.294 0.600 0.400 CB 1.106 I: 7 2.625 1.385 0.375 0.413 0.667 0.500 CB 0.925 J: 10 3.693 1.791 0.379 0.234 0.667 0.444 CB 1.019 K: 7 2.095 1.472 0.688 0.309 0.500 0.500 CB 0.850 L: 7 3.680 1.573 0.359 0.380 0.667 0.333 CB 1.197 7 residues pruned to eliminate duplicates M: 9 4.709 1.230 0.664 0.479 0.875 0.625 N 0.949 N: 6 3.381 1.613 0.594 0.295 0.800 0.600 CB 0.925 O: 7 3.327 1.241 0.421 0.486 0.833 0.333 CB 0.937 Using tripeptides from previous cycle as seeds 99 residues left after pruning, divided into chains as follows: A: 11 B: 8 C: 24 D: 9 E: 9 F: 11 G: 7 H: 6 I: 7 J: 7 CC for partial structure against native data = 10.74 % ------------------------------------------------------------------------------ Global autotracing cycle 6 Phases from autotracing cycle 5 used as input for final density modification = 0.281, Contrast = 0.342, Connect. = 0.620 for dens.mod. cycle 1 = 0.282, Contrast = 0.395, Connect. = 0.630 for dens.mod. cycle 2 = 0.282, Contrast = 0.452, Connect. = 0.678 for dens.mod. cycle 3 = 0.282, Contrast = 0.465, Connect. = 0.694 for dens.mod. cycle 4 = 0.282, Contrast = 0.472, Connect. = 0.704 for dens.mod. cycle 5 Estimated mean FOM and mapCC as a function of resolution d inf - 5.07 - 3.99 - 3.47 - 3.14 - 2.91 - 2.74 - 2.60 - 2.48 - 2.38 - 2.29 0.441 0.541 0.554 0.596 0.524 0.469 0.438 0.410 0.376 0.313 0.571 0.717 0.751 0.806 0.744 0.705 0.672 0.674 0.642 0.550 N 2104 2106 2122 2159 2095 2055 2101 2201 2179 1902 Estimated mean FOM = 0.467 Pseudo-free CC = 51.45 % Anomalous density (in sigma units) at initial heavy atom sites Site x y z occ*Z density 1 0.5541 0.4339 0.8982 1.0000 31.34 2 0.6957 0.1815 1.1240 0.8698 26.61 3 0.7936 0.3155 0.9177 0.5934 18.75 4 0.9573 0.0725 1.1061 0.5767 16.88 5 0.7978 0.2645 1.0200 0.4406 13.29 6 0.6341 0.3256 0.9402 0.4350 11.49 7 0.5234 0.0150 1.1009 0.4013 10.90 8 0.8475 0.1709 1.1486 0.4010 11.48 9 0.4318 0.4287 1.0207 0.3726 11.01 10 0.7023 0.2346 1.2318 0.3686 10.18 11 0.9843 0.2490 1.0603 0.3567 10.55 12 0.8466 0.4560 0.9134 0.3156 9.80 13 0.5621 0.2416 1.1218 0.3051 9.29 Site x y z h(sig) near old near new 1 0.5544 0.4340 0.8980 31.4 1/0.03 15/2.50 16/2.59 7/10.78 8/13.66 2 0.6946 0.1820 1.1238 26.7 2/0.08 13/8.95 6/9.28 11/11.15 18/11.87 3 0.7939 0.3158 0.9183 18.8 3/0.06 7/9.49 5/10.67 12/12.08 16/16.52 4 0.9577 0.0742 1.1065 17.1 4/0.15 6/10.88 14/15.19 10/15.25 2/17.69 5 0.7985 0.2645 1.0212 13.5 5/0.12 18/9.76 3/10.67 10/11.32 7/13.29 6 0.8500 0.1711 1.1473 11.6 8/0.19 2/9.28 4/10.88 11/12.72 10/13.07 7 0.6339 0.3266 0.9397 11.5 6/0.09 16/9.20 3/9.49 1/10.78 15/12.48 8 0.4322 0.4276 1.0210 11.1 9/0.10 8/12.66 16/12.94 1/13.66 15/13.77 9 0.5212 0.0137 1.1010 11.0 7/0.17 2/17.31 13/19.22 9/19.34 6/23.44 10 0.9832 0.2490 1.0592 10.6 11/0.13 5/11.32 6/13.07 4/15.25 18/16.80 11 0.7050 0.2350 1.2314 10.3 10/0.17 2/11.15 6/12.72 17/12.91 13/13.10 12 0.8497 0.4561 0.9135 9.9 12/0.18 3/12.08 15/16.56 7/16.63 1/17.19 13 0.5656 0.2419 1.1228 9.5 13/0.23 2/8.95 18/11.23 11/13.10 5/16.64 14 1.0504 -0.0690 1.1885 -6.3 4/15.10 14/13.06 4/15.19 4/22.83 6/23.36 15 0.5632 0.4627 0.9038 -6.1 1/2.50 1/2.50 16/4.96 7/12.48 8/13.77 16 0.5392 0.4052 0.9029 -5.7 1/2.57 1/2.59 15/4.96 7/9.20 8/12.94 17 0.7999 0.3193 1.3299 -4.7 10/12.97 11/12.91 10/17.37 8/21.47 6/21.47 18 0.7182 0.3209 1.0971 4.7 5/9.83 5/9.76 13/11.23 2/11.87 11/14.64 Best trace (cycle 5 with CC 10.74%) was saved as P2221_pse14_i.pdb ============================================================================== CPU times required in seconds ----------------------------- 9.8 - Setup, data input and phasing 1.2 - FFTs and peak-searches 4.7 - Sphere of influence 0.6 - Rest of density modification 0.0 - Alpha-helix search 50.4 - Tripeptide search 45.0 - Chain tracing 0.0 - NCS analysis 3.9 - B-value refinement for trace 0.1 - Rest of tracing 0.0 - Comparison with known structure ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE finished at 12:05:38 Total time: 115.63 secs + ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++