++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE - PHASING AND DENSITY MODIFICATION - Version 2019/1 + + Copyright (c) George M. Sheldrick and Isabel Uson 2001-19 + + Started at 12:17:28 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: P22121_pse14 P22121_pse14_fa -i -h -a5 -s0.45 -z -e -m5 Cell, symmetry and heavy atoms from: P22121_pse14_fa.res FA and alpha from P22121_pse14_fa.hkl Native data from P22121_pse14.hkl Listing output to P22121_pse14_i.lst Phases output to P22121_pse14_i.phs Revised heavy atom sites output to P22121_pse14_i.hat Revised heavy atom phases output to P22121_pse14_i.pha Poly-Ala trace output to P22121_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 21 21 Allowed origin shift code: 4 14 atoms read from file P22121_pse14_fa.res Trimmed to 13 atoms with occupancy > 0.2 11695 Reflections read from file P22121_pse14_fa.hkl 21043 Reflections read from file P22121_pse14.hkl 21007 Unique data, highest resolution = 2.289 Angstroms Anisotropic scaling: intensities multiplied by 0.004007h^2 -0.000438k^2 -0.000495l^2 +0.000000kl +0.000000hl +0.000000hk 25 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 19.22% 0.266 Overall CC between Eobs (from delF) and Ecalc (from heavy atoms) = 18.81% = 0.123 for phases from phiT = phiA + alpha = 0.187 after including heavy atoms = 0.173, Contrast = 0.029, Connect. = 0.527 for dens.mod. cycle 1 = 0.190, Contrast = 0.083, Connect. = 0.589 for dens.mod. cycle 2 = 0.203, Contrast = 0.110, Connect. = 0.603 for dens.mod. cycle 3 = 0.216, Contrast = 0.139, Connect. = 0.619 for dens.mod. cycle 4 = 0.226, Contrast = 0.166, Connect. = 0.632 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 11 heavy atoms with Occ*Z > 0.30 added to NOGO map 1870 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 256 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 6 1.710 1.291 0.209 0.597 0.600 0.400 N 0.744 ? A: 8 4.469 1.325 0.320 0.504 0.857 0.429 N 1.122 B: 18 4.859 1.313 0.356 0.451 0.706 0.353 CB 0.987 C: 7 3.919 1.406 0.588 0.490 0.667 0.500 N 1.085 D: 7 4.860 1.228 0.570 0.557 1.000 0.500 CA 0.974 E: 7 4.451 1.304 0.847 0.504 1.000 0.667 CB 0.751 F: 8 2.787 1.465 0.858 0.224 0.714 0.429 CB 0.748 G: 6 4.211 1.525 0.295 0.473 0.800 0.400 N 1.226 6 1.996 1.362 -0.074 0.434 0.600 0.400 CB 1.263 ? H: 6 2.987 1.318 0.650 0.312 0.800 0.600 CB 0.945 I: 6 2.474 1.230 0.372 0.332 0.800 0.400 O 0.985 J: 9 4.101 1.443 0.238 0.458 0.750 0.500 CB 1.129 6 1.742 1.596 0.058 0.100 0.600 0.200 CB 1.282 ? K: 8 3.939 1.296 0.716 0.443 1.000 0.571 CB 0.710 L: 9 2.834 1.302 0.211 0.354 0.750 0.375 CB 0.992 M: 6 3.043 1.373 0.202 0.487 0.800 0.600 O 1.044 N: 7 3.253 1.382 0.487 0.264 0.667 0.667 CB 1.268 O: 6 5.358 1.478 0.622 0.487 0.800 0.400 CA 1.267 P: 11 3.135 1.162 0.080 0.376 0.900 0.300 CB 1.002 6 1.630 1.221 0.558 0.293 0.600 0.400 N 0.805 ? 8 1.949 1.161 0.151 0.357 0.714 0.429 CB 0.900 ? Q: 6 3.674 1.230 0.827 0.554 1.000 0.400 CB 0.694 R: 8 3.270 1.390 0.484 0.350 0.714 0.429 CB 0.987 118 residues left after pruning, divided into chains as follows: A: 8 B: 18 C: 7 D: 7 E: 7 F: 8 G: 6 H: 6 I: 8 J: 6 K: 6 L: 11 M: 6 N: 8 O: 6 CC for partial structure against native data = 9.14 % ------------------------------------------------------------------------------ Global autotracing cycle 2 = 0.281, Contrast = 0.205, Connect. = 0.591 for dens.mod. cycle 1 = 0.281, Contrast = 0.212, Connect. = 0.562 for dens.mod. cycle 2 = 0.281, Contrast = 0.241, Connect. = 0.595 for dens.mod. cycle 3 = 0.281, Contrast = 0.259, Connect. = 0.613 for dens.mod. cycle 4 = 0.281, Contrast = 0.280, Connect. = 0.630 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 11 heavy atoms with Occ*Z > 0.30 added to NOGO map 2438 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 271 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 7 7.657 1.975 0.712 0.498 0.667 0.333 CB 1.389 B: 9 9.533 2.187 0.214 0.354 1.000 0.625 CB 1.486 C: 17 4.760 1.630 0.300 0.222 0.688 0.375 CB 1.131 D: 9 6.820 1.684 0.717 0.268 0.750 0.500 O 1.448 E: 6 3.197 1.859 0.542 0.385 0.400 0.400 CB 1.409 6 1.038 1.688 -0.062 0.280 0.400 0.400 CB 0.940 ? F: 6 5.112 1.694 0.090 0.617 0.800 0.600 N 1.386 G: 6 4.949 1.345 0.515 0.764 1.000 0.200 CB 0.859 H: 7 5.316 1.786 0.447 0.284 0.833 0.333 CB 1.286 I: 8 4.526 1.431 0.406 0.425 0.714 0.571 C 1.287 J: 9 2.527 1.371 0.579 0.208 0.625 0.375 N 0.933 K: 7 4.754 1.766 0.169 0.251 0.833 0.500 CA 1.503 6 1.207 1.330 0.306 0.297 0.600 0.600 N 0.649 ? L: 6 3.479 1.505 0.325 0.292 0.800 0.800 C 1.231 M: 9 2.229 1.199 0.166 0.306 0.750 0.500 CB 0.932 N: 6 2.514 1.530 0.492 0.471 0.600 0.400 CB 0.845 O: 7 3.012 1.380 0.038 0.460 0.833 0.667 CB 1.073 P: 11 5.271 1.323 0.667 0.574 0.800 0.600 N 0.879 Q: 6 2.563 1.265 0.438 0.245 0.800 0.800 CA 1.057 6 1.990 1.603 -0.039 0.442 0.800 0.400 CB 0.766 ? R: 6 3.049 1.232 0.717 0.643 1.000 0.800 CB 0.564 7 1.154 1.224 0.114 0.134 0.667 0.333 CB 0.817 ? S: 9 3.082 1.499 0.185 0.317 0.750 0.625 CB 1.002 T: 8 4.717 1.541 0.243 0.487 0.857 0.571 CB 1.100 U: 9 5.037 1.482 0.404 0.349 0.875 0.375 CB 1.152 Using tripeptides from previous cycle as seeds V: 14 8.786 1.862 0.228 0.451 0.769 0.308 CB 1.457 6 residues pruned to eliminate duplicates 115 residues left after pruning, divided into chains as follows: A: 7 B: 9 C: 15 D: 9 E: 6 F: 6 G: 8 H: 9 I: 6 J: 6 K: 6 L: 8 M: 6 N: 14 CC for partial structure against native data = 9.83 % ------------------------------------------------------------------------------ Global autotracing cycle 3 = 0.281, Contrast = 0.252, Connect. = 0.597 for dens.mod. cycle 1 = 0.281, Contrast = 0.271, Connect. = 0.584 for dens.mod. cycle 2 = 0.281, Contrast = 0.308, Connect. = 0.621 for dens.mod. cycle 3 = 0.281, Contrast = 0.328, Connect. = 0.641 for dens.mod. cycle 4 = 0.281, Contrast = 0.343, Connect. = 0.654 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 11 heavy atoms with Occ*Z > 0.30 added to NOGO map 2298 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 236 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 16 13.616 2.118 0.218 0.467 0.800 0.333 CB 1.761 B: 7 9.975 1.900 0.739 0.453 1.000 0.500 CB 1.294 C: 9 12.526 2.396 0.377 0.346 1.000 0.625 CB 1.587 D: 7 6.285 1.520 0.453 0.668 1.000 0.667 CB 0.994 E: 10 6.570 1.682 0.330 0.278 0.889 0.556 CB 1.415 F: 6 4.209 1.827 0.598 0.289 0.600 0.600 CA 1.363 G: 11 4.763 1.761 0.526 0.201 0.600 0.400 N 1.332 H: 8 4.391 1.570 0.542 0.326 1.000 0.571 CB 0.830 I: 6 9.402 1.887 0.842 0.447 1.000 0.400 CB 1.277 J: 12 3.587 1.416 0.278 0.268 0.727 0.545 C 1.071 K: 6 2.393 1.411 -0.096 0.466 0.800 0.400 CB 1.086 6 1.984 1.375 0.556 0.458 0.600 0.600 CB 0.721 ? L: 15 6.029 1.588 0.360 0.360 0.786 0.643 CB 1.105 15 residues pruned to eliminate duplicates M: 16 6.053 1.726 0.263 0.306 0.667 0.333 N 1.334 N: 7 2.733 1.503 0.170 0.441 0.667 0.333 CB 1.013 O: 6 5.477 1.589 0.718 0.437 0.800 0.600 N 1.197 P: 7 3.236 1.373 0.081 0.548 0.833 0.667 CB 1.019 Q: 8 3.432 1.460 0.533 0.342 0.571 0.571 N 1.204 R: 15 5.666 1.590 0.434 0.234 0.786 0.357 CB 1.151 S: 9 3.052 1.403 0.405 0.277 0.625 0.375 CB 1.127 T: 9 6.897 1.504 1.116 0.580 0.750 0.125 CB 0.946 Using tripeptides from previous cycle as seeds U: 6 3.398 1.408 0.887 0.225 0.800 0.400 O 0.987 94 residues left after pruning, divided into chains as follows: A: 16 B: 7 C: 9 D: 7 E: 10 F: 6 G: 9 H: 9 I: 7 J: 14 CC for partial structure against native data = 8.00 % ------------------------------------------------------------------------------ Global autotracing cycle 4 = 0.281, Contrast = 0.282, Connect. = 0.604 for dens.mod. cycle 1 = 0.281, Contrast = 0.305, Connect. = 0.592 for dens.mod. cycle 2 = 0.281, Contrast = 0.346, Connect. = 0.632 for dens.mod. cycle 3 = 0.281, Contrast = 0.366, Connect. = 0.652 for dens.mod. cycle 4 = 0.281, Contrast = 0.380, Connect. = 0.666 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 11 heavy atoms with Occ*Z > 0.30 added to NOGO map 2225 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 262 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 10 9.430 2.092 0.481 0.399 0.667 0.444 CB 1.692 B: 15 15.476 2.224 0.327 0.414 0.786 0.429 CB 1.951 C: 8 6.859 1.680 0.564 0.681 1.000 0.714 CB 0.836 D: 7 4.486 1.931 0.464 0.247 0.667 0.500 CB 1.301 E: 7 8.080 1.814 0.607 0.492 0.833 0.500 CB 1.369 F: 10 4.510 1.866 0.283 0.234 0.667 0.222 CB 1.284 G: 12 7.492 2.238 0.160 0.241 0.818 0.455 CB 1.434 6 residues pruned to eliminate duplicates H: 10 3.867 1.867 -0.033 0.370 0.667 0.444 CB 1.231 I: 14 6.638 1.659 0.438 0.212 0.846 0.385 CB 1.282 J: 8 2.476 1.357 0.328 0.248 0.714 0.571 O 0.972 K: 6 3.119 1.353 0.517 0.488 0.600 0.600 O 1.147 L: 6 4.789 1.113 0.670 0.712 1.000 0.600 CB 0.951 M: 14 4.658 1.620 0.086 0.124 0.846 0.231 N 1.391 6 0.714 1.190 0.496 0.320 0.600 0.400 CB 0.365 ? N: 8 2.723 1.318 0.390 0.391 0.714 0.571 C 0.882 O: 6 2.909 1.391 0.308 0.303 0.800 0.800 N 1.114 P: 9 3.174 1.547 0.099 0.155 0.875 0.500 CB 1.151 8 1.387 1.395 -0.283 0.338 0.857 0.286 CB 0.730 ? Q: 10 2.544 1.142 0.057 0.441 0.889 0.444 O 0.840 R: 13 2.965 1.181 0.445 0.299 0.750 0.250 CB 0.838 Using tripeptides from previous cycle as seeds 125 residues left after pruning, divided into chains as follows: A: 10 B: 8 C: 6 D: 10 E: 7 F: 7 G: 13 H: 6 I: 6 J: 11 K: 6 L: 9 M: 26 CC for partial structure against native data = 12.81 % ------------------------------------------------------------------------------ Global autotracing cycle 5 = 0.281, Contrast = 0.291, Connect. = 0.609 for dens.mod. cycle 1 = 0.281, Contrast = 0.324, Connect. = 0.606 for dens.mod. cycle 2 = 0.281, Contrast = 0.375, Connect. = 0.649 for dens.mod. cycle 3 = 0.281, Contrast = 0.394, Connect. = 0.669 for dens.mod. cycle 4 = 0.281, Contrast = 0.409, Connect. = 0.682 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 11 heavy atoms with Occ*Z > 0.30 added to NOGO map 2102 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 250 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 7 6.043 1.912 0.492 0.621 1.000 0.667 CB 0.772 B: 19 12.275 1.947 0.327 0.367 0.778 0.333 CB 1.660 C: 9 8.098 2.110 0.354 0.418 0.875 0.500 CB 1.249 D: 8 5.894 1.716 0.385 0.297 0.857 0.429 CB 1.371 E: 8 4.215 2.052 0.306 0.313 0.571 0.429 CB 1.279 F: 9 3.577 1.675 0.067 0.351 0.750 0.375 CB 1.109 G: 8 6.437 1.628 0.440 0.311 0.857 0.571 O 1.492 H: 6 5.272 1.780 0.601 0.299 1.000 0.600 CB 1.035 I: 7 5.905 1.895 0.270 0.351 0.833 0.500 CB 1.412 J: 10 6.120 1.707 0.277 0.427 0.778 0.444 N 1.299 K: 6 2.907 1.326 0.570 0.418 0.600 0.400 C 1.133 L: 9 2.812 1.451 0.214 0.283 0.750 0.500 N 0.961 M: 18 4.932 1.541 0.408 0.249 0.647 0.176 CB 1.139 N: 10 6.115 1.545 0.474 0.369 0.889 0.556 CB 1.159 O: 8 3.605 1.613 0.179 0.290 0.714 0.286 CB 1.269 P: 9 2.977 1.561 0.244 0.338 0.750 0.375 CB 0.862 Using tripeptides from previous cycle as seeds Q: 7 5.957 1.354 0.569 0.772 0.833 0.667 CB 1.080 106 residues left after pruning, divided into chains as follows: A: 7 B: 19 C: 8 D: 8 E: 8 F: 8 G: 6 H: 7 I: 6 J: 13 K: 10 L: 6 CC for partial structure against native data = 12.16 % ------------------------------------------------------------------------------ Global autotracing cycle 6 Phases from autotracing cycle 4 used as input for final density modification = 0.281, Contrast = 0.298, Connect. = 0.609 for dens.mod. cycle 1 = 0.282, Contrast = 0.338, Connect. = 0.609 for dens.mod. cycle 2 = 0.282, Contrast = 0.394, Connect. = 0.653 for dens.mod. cycle 3 = 0.282, Contrast = 0.416, Connect. = 0.674 for dens.mod. cycle 4 = 0.282, Contrast = 0.430, Connect. = 0.688 for dens.mod. cycle 5 Estimated mean FOM and mapCC as a function of resolution d inf - 5.06 - 3.99 - 3.47 - 3.14 - 2.91 - 2.74 - 2.60 - 2.48 - 2.38 - 2.29 0.490 0.608 0.615 0.532 0.480 0.400 0.377 0.354 0.276 0.235 0.628 0.800 0.824 0.757 0.696 0.618 0.588 0.590 0.460 0.404 N 2110 2091 2120 2158 2094 2054 2100 2200 2179 1901 Estimated mean FOM = 0.438 Pseudo-free CC = 47.97 % Anomalous density (in sigma units) at initial heavy atom sites Site x y z occ*Z density 1 0.7013 0.9333 0.7549 1.0000 27.01 2 0.4478 0.8205 0.7676 0.8424 22.66 3 0.9533 0.8145 0.7383 0.7827 20.90 4 0.3582 0.9246 0.8080 0.6636 19.07 5 0.7075 0.9855 0.8616 0.6082 19.25 6 0.2025 0.9291 0.7836 0.5136 15.20 7 0.5268 0.9536 0.7911 0.4821 14.44 8 0.7072 0.8061 0.7047 0.4364 11.66 9 0.0177 0.9942 0.9259 0.4244 11.55 10 0.7105 0.6803 0.7600 0.4204 12.38 11 0.7312 1.1036 0.7770 0.3407 10.05 12 0.0772 0.9407 0.8132 0.2952 8.70 13 -0.0492 1.0874 0.8846 0.2657 7.08 Site x y z h(sig) near old near new 1 0.7000 0.9332 0.7555 27.1 1/0.09 15/2.65 14/8.39 7/10.72 4/11.00 2 0.4479 0.8205 0.7674 22.7 2/0.03 16/2.57 19/2.63 17/5.42 5/10.79 3 0.9530 0.8145 0.7389 20.9 3/0.06 10/14.54 12/14.63 15/17.40 1/17.67 4 0.7076 0.9851 0.8618 19.3 5/0.04 18/2.53 14/2.63 15/10.09 1/11.00 5 0.3578 0.9242 0.8085 19.1 4/0.07 19/9.20 6/9.23 7/10.12 2/10.79 6 0.2038 0.9292 0.7828 15.3 6/0.11 12/8.08 5/9.23 19/16.06 2/16.78 7 0.5261 0.9530 0.7914 14.5 7/0.07 19/9.58 5/10.12 15/10.34 1/10.72 8 0.7126 0.6799 0.7603 12.4 10/0.13 10/11.70 3/17.92 2/19.22 17/19.24 9 0.0191 0.9926 0.9262 11.7 9/0.16 13/9.51 12/11.87 6/18.08 4/18.99 10 0.7075 0.8057 0.7047 11.7 8/0.04 15/11.25 1/11.63 8/11.70 3/14.54 11 0.7304 1.1031 0.7776 10.2 11/0.09 18/10.94 14/12.29 4/12.71 1/14.36 12 0.0751 0.9430 0.8140 8.9 12/0.24 6/8.08 9/11.87 3/14.63 13/15.28 13 -0.0487 1.0850 0.8835 7.2 13/0.23 9/9.51 12/15.28 18/15.72 11/16.31 14 0.6998 0.9730 0.8367 -5.6 5/2.62 4/2.63 18/4.22 15/7.57 1/8.39 15 0.6940 0.9129 0.7768 -5.4 1/2.71 1/2.65 14/7.57 4/10.09 7/10.34 16 0.4397 0.8156 0.7411 -5.1 2/2.59 2/2.57 17/3.00 19/4.03 5/12.02 17 0.4534 0.8144 0.7107 -4.9 2/5.45 16/3.00 2/5.42 19/6.22 5/14.14 18 0.7004 1.0152 0.8615 -4.7 5/2.50 4/2.53 14/4.22 11/10.94 15/11.70 19 0.4576 0.8514 0.7676 -4.6 2/2.63 2/2.63 16/4.03 17/6.22 5/9.20 Best trace (cycle 4 with CC 12.81%) was saved as P22121_pse14_i.pdb ============================================================================== CPU times required in seconds ----------------------------- 9.8 - Setup, data input and phasing 1.3 - FFTs and peak-searches 4.7 - Sphere of influence 0.1 - Rest of density modification 0.0 - Alpha-helix search 54.3 - Tripeptide search 61.2 - Chain tracing 0.0 - NCS analysis 5.0 - B-value refinement for trace 0.1 - Rest of tracing 0.0 - Comparison with known structure ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE finished at 12:19:44 Total time: 136.35 secs + ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++