++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE - PHASING AND DENSITY MODIFICATION - Version 2019/1 + + Copyright (c) George M. Sheldrick and Isabel Uson 2001-19 + + Started at 12:14:46 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: P21221_pse14 P21221_pse14_fa -i -h -a5 -s0.45 -z -e -m5 Cell, symmetry and heavy atoms from: P21221_pse14_fa.res FA and alpha from P21221_pse14_fa.hkl Native data from P21221_pse14.hkl Listing output to P21221_pse14_i.lst Phases output to P21221_pse14_i.phs Revised heavy atom sites output to P21221_pse14_i.hat Revised heavy atom phases output to P21221_pse14_i.pha Poly-Ala trace output to P21221_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 21 2 21 Allowed origin shift code: 4 14 atoms read from file P21221_pse14_fa.res Trimmed to 11 atoms with occupancy > 0.2 11695 Reflections read from file P21221_pse14_fa.hkl 21043 Reflections read from file P21221_pse14.hkl 21014 Unique data, highest resolution = 2.289 Angstroms Anisotropic scaling: intensities multiplied by 0.003997h^2 -0.000434k^2 -0.000495l^2 +0.000000kl +0.000000hl +0.000000hk 23 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 ** 11 potential heavy atoms found Substructure optimization LOR = lowest occupancy retained, HOR = highest occupancy rejected Nats CC(HA) LOR HOR 11 20.07% 0.281 Overall CC between Eobs (from delF) and Ecalc (from heavy atoms) = 20.02% = 0.126 for phases from phiT = phiA + alpha = 0.210 after including heavy atoms = 0.176, Contrast = 0.040, Connect. = 0.541 for dens.mod. cycle 1 = 0.197, Contrast = 0.117, Connect. = 0.601 for dens.mod. cycle 2 = 0.211, Contrast = 0.160, Connect. = 0.619 for dens.mod. cycle 3 = 0.223, Contrast = 0.195, Connect. = 0.635 for dens.mod. cycle 4 = 0.233, Contrast = 0.220, Connect. = 0.646 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 9 heavy atoms with Occ*Z > 0.30 added to NOGO map 1954 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 265 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 6 6.198 1.156 1.121 0.856 0.800 0.600 CA 1.042 B: 10 3.617 1.436 0.369 0.179 0.778 0.333 CB 1.160 C: 11 4.086 1.389 0.364 0.458 0.800 0.200 CB 0.891 D: 8 4.846 1.523 0.909 0.482 0.857 0.571 CB 0.748 E: 7 3.093 1.459 0.455 0.292 0.667 0.333 CB 1.127 F: 10 4.691 1.274 0.541 0.603 0.778 0.444 CB 0.928 G: 9 4.203 1.521 0.211 0.461 0.875 0.375 CB 0.960 H: 9 2.117 1.319 0.788 0.305 0.375 0.250 CB 1.051 I: 10 3.747 1.164 0.325 0.571 0.889 0.556 N 0.851 J: 7 2.543 1.359 0.138 0.300 0.833 0.500 CB 1.007 K: 6 3.497 1.376 0.518 0.259 0.800 0.400 CA 1.234 L: 6 2.783 1.503 0.104 0.292 1.000 1.000 CB 0.947 M: 6 4.279 1.180 0.625 0.809 1.000 1.000 N 0.762 7 1.313 1.171 0.179 0.153 0.667 0.500 CB 0.892 ? 6 1.455 1.120 0.806 0.298 0.600 0.400 CB 0.672 ? N: 7 2.461 1.424 0.263 0.283 0.667 0.500 N 1.070 91 residues left after pruning, divided into chains as follows: A: 10 B: 11 C: 8 D: 7 E: 10 F: 9 G: 7 H: 10 I: 7 J: 6 K: 6 CC for partial structure against native data = 7.01 % ------------------------------------------------------------------------------ Global autotracing cycle 2 = 0.281, Contrast = 0.236, Connect. = 0.599 for dens.mod. cycle 1 = 0.281, Contrast = 0.257, Connect. = 0.577 for dens.mod. cycle 2 = 0.281, Contrast = 0.297, Connect. = 0.616 for dens.mod. cycle 3 = 0.281, Contrast = 0.321, Connect. = 0.637 for dens.mod. cycle 4 = 0.281, Contrast = 0.343, Connect. = 0.655 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 9 heavy atoms with Occ*Z > 0.30 added to NOGO map 2308 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 251 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 18 7.505 1.652 0.484 0.342 0.706 0.353 N 1.250 B: 6 7.484 1.731 0.488 0.681 0.800 0.600 CB 1.375 C: 6 3.014 1.714 -0.035 0.293 0.800 0.600 CB 1.283 6 1.650 1.645 -0.131 0.291 0.600 0.400 CB 1.088 ? D: 6 7.968 1.466 0.802 0.601 1.000 0.600 O 1.225 E: 6 3.088 1.511 0.047 0.462 0.800 0.400 CB 1.134 F: 8 3.131 1.291 0.339 0.370 0.857 0.286 CB 0.917 G: 11 8.492 1.705 0.325 0.475 0.800 0.200 CB 1.524 H: 7 3.956 1.519 0.126 0.539 0.833 0.667 CB 1.091 I: 6 2.482 1.407 0.011 0.324 1.000 0.400 N 0.948 J: 10 2.366 1.582 0.231 0.295 0.778 0.556 CB 0.655 K: 8 3.104 1.476 0.365 0.219 0.714 0.286 CB 1.134 L: 8 4.696 1.517 0.561 0.346 0.714 0.429 CA 1.242 6 1.948 1.340 0.708 0.298 0.600 0.400 N 0.795 ? M: 7 3.035 1.296 0.407 0.479 0.833 0.500 CB 0.833 N: 10 4.358 1.460 0.530 0.341 0.667 0.333 O 1.160 6 1.954 1.116 0.131 0.467 0.800 0.600 O 0.895 ? O: 7 4.859 1.424 0.698 0.400 0.833 0.333 CA 1.094 7 1.577 1.041 0.089 0.549 0.833 0.833 C 0.650 ? P: 14 3.510 1.356 0.398 0.171 0.769 0.231 CB 0.995 Using tripeptides from previous cycle as seeds Q: 6 3.284 1.582 0.027 0.393 0.800 0.400 CB 1.266 R: 6 5.003 1.489 0.698 0.218 1.000 0.200 CB 1.233 91 residues left after pruning, divided into chains as follows: A: 17 B: 6 C: 11 D: 7 E: 7 F: 10 G: 7 H: 14 I: 6 J: 6 CC for partial structure against native data = 9.21 % ------------------------------------------------------------------------------ Global autotracing cycle 3 = 0.281, Contrast = 0.286, Connect. = 0.599 for dens.mod. cycle 1 = 0.281, Contrast = 0.320, Connect. = 0.593 for dens.mod. cycle 2 = 0.281, Contrast = 0.367, Connect. = 0.638 for dens.mod. cycle 3 = 0.281, Contrast = 0.388, Connect. = 0.659 for dens.mod. cycle 4 = 0.281, Contrast = 0.405, Connect. = 0.675 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 9 heavy atoms with Occ*Z > 0.30 added to NOGO map 2149 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 253 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 15 14.616 2.316 0.320 0.414 0.786 0.357 CB 1.779 B: 9 7.532 1.556 0.564 0.675 0.750 0.625 O 1.242 C: 20 6.221 1.597 0.256 0.215 0.789 0.368 CB 1.262 D: 6 4.089 1.852 -0.048 0.625 0.800 0.600 N 1.152 E: 10 9.081 2.001 0.748 0.398 0.667 0.333 CB 1.446 F: 7 5.533 1.788 0.738 0.340 0.667 0.500 N 1.298 G: 8 4.515 1.726 0.342 0.440 0.714 0.429 CB 1.097 H: 7 2.902 1.334 0.474 0.284 0.667 0.333 CA 1.154 I: 8 2.907 1.616 0.246 0.354 0.714 0.429 CB 0.895 J: 7 3.896 1.351 0.126 0.419 1.000 0.833 N 1.138 6 1.046 1.337 0.516 0.178 0.400 0.200 CA 0.851 ? 7 1.284 1.276 0.282 0.344 0.500 0.500 N 0.759 ? K: 6 4.831 1.471 0.201 0.758 0.800 0.600 O 1.215 L: 12 11.786 1.972 0.486 0.385 0.818 0.545 N 1.674 13 residues pruned to eliminate duplicates M: 11 3.967 1.402 0.757 0.148 0.700 0.400 O 1.122 Using tripeptides from previous cycle as seeds N: 9 8.220 2.019 0.538 0.353 0.750 0.500 N 1.463 95 residues left after pruning, divided into chains as follows: A: 9 B: 13 C: 6 D: 10 E: 7 F: 8 G: 7 H: 6 I: 14 J: 8 K: 7 CC for partial structure against native data = 9.49 % ------------------------------------------------------------------------------ Global autotracing cycle 4 = 0.281, Contrast = 0.302, Connect. = 0.610 for dens.mod. cycle 1 = 0.281, Contrast = 0.345, Connect. = 0.613 for dens.mod. cycle 2 = 0.281, Contrast = 0.398, Connect. = 0.658 for dens.mod. cycle 3 = 0.281, Contrast = 0.417, Connect. = 0.675 for dens.mod. cycle 4 = 0.281, Contrast = 0.433, Connect. = 0.688 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 9 heavy atoms with Occ*Z > 0.30 added to NOGO map 2096 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 249 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 10 13.659 1.763 0.455 0.785 1.000 0.667 CA 1.381 B: 13 13.830 2.386 0.350 0.410 0.750 0.333 CB 1.816 C: 7 3.672 1.875 -0.134 0.347 0.833 0.500 CB 1.308 D: 11 4.711 1.571 0.463 0.403 0.700 0.500 CB 1.025 E: 7 6.163 1.566 0.434 0.818 0.833 0.667 CB 1.020 F: 14 4.083 1.420 0.424 0.326 0.692 0.462 CB 0.979 G: 12 6.757 1.957 0.405 0.329 0.727 0.182 CB 1.228 H: 7 3.714 1.891 0.426 0.253 0.667 0.667 CB 1.121 I: 6 6.659 1.512 0.637 0.675 1.000 1.000 CB 1.024 J: 6 2.637 1.496 0.041 0.357 0.800 0.600 CA 1.104 K: 6 3.921 1.389 0.741 0.300 0.800 0.400 CA 1.134 6 1.904 1.362 0.261 0.380 0.600 0.400 N 0.939 ? L: 11 2.209 1.771 0.081 0.131 0.500 0.200 CB 1.155 7 residues pruned to eliminate duplicates M: 8 2.044 1.168 0.555 0.207 0.714 0.286 C 0.842 7 1.957 1.297 0.261 0.249 0.833 0.667 CB 0.782 ? Using tripeptides from previous cycle as seeds N: 9 5.036 1.679 0.218 0.461 0.750 0.375 CB 1.208 O: 12 5.031 1.373 0.337 0.472 0.727 0.545 N 1.169 106 residues left after pruning, divided into chains as follows: A: 10 B: 13 C: 11 D: 13 E: 12 F: 7 G: 11 H: 12 I: 17 CC for partial structure against native data = 10.74 % ------------------------------------------------------------------------------ Global autotracing cycle 5 = 0.281, Contrast = 0.323, Connect. = 0.617 for dens.mod. cycle 1 = 0.281, Contrast = 0.368, Connect. = 0.617 for dens.mod. cycle 2 = 0.281, Contrast = 0.423, Connect. = 0.663 for dens.mod. cycle 3 = 0.281, Contrast = 0.439, Connect. = 0.681 for dens.mod. cycle 4 = 0.281, Contrast = 0.453, Connect. = 0.694 for dens.mod. cycle 5 NOGO map generated for regions about rotation axes (if any) 9 heavy atoms with Occ*Z > 0.30 added to NOGO map 2011 peaks > 0.5 sigma used to seed fragment search Space for about 499 unique residues taking solvent into account 246 potential tripeptides employed #(CA) CFOM DenFit NH...O SecStr Rama98 Rama80 A: 11 15.582 2.489 0.456 0.419 0.800 0.500 CB 1.849 B: 6 11.611 2.026 0.360 0.711 1.000 1.000 N 1.556 C: 7 11.176 2.027 0.271 0.894 1.000 0.500 CB 1.271 D: 6 4.333 1.761 0.399 0.481 0.800 0.600 CB 1.002 E: 8 5.444 1.956 0.491 0.441 0.714 0.571 CB 1.050 F: 14 3.549 1.545 0.649 0.285 0.462 0.385 N 1.066 6 1.181 1.313 0.027 0.327 0.800 0.400 CB 0.592 ? G: 6 3.565 1.581 0.672 0.376 0.600 0.400 CB 1.148 H: 9 4.075 1.487 0.563 0.376 0.625 0.500 N 1.131 I: 8 2.896 1.712 0.298 0.498 0.571 0.429 CB 0.864 J: 7 2.910 1.522 0.564 0.233 0.667 0.500 CB 1.021 K: 11 3.166 1.595 0.389 0.150 0.600 0.300 CB 1.159 L: 7 3.781 1.334 0.516 0.572 0.833 0.833 CA 0.854 M: 7 2.482 1.258 0.128 0.496 0.833 0.333 CB 0.861 N: 12 7.030 1.594 0.946 0.381 0.909 0.636 CB 0.853 O: 8 4.183 1.565 0.553 0.184 0.714 0.429 O 1.331 P: 7 2.268 1.522 0.216 0.278 0.500 0.333 O 1.286 Q: 7 3.159 1.577 0.365 0.250 0.833 0.333 CB 0.958 R: 9 2.322 1.467 0.178 0.191 0.750 0.250 CB 0.917 Using tripeptides from previous cycle as seeds S: 11 11.583 1.814 0.244 0.720 1.000 0.600 N 1.330 6 residues pruned to eliminate duplicates 93 residues left after pruning, divided into chains as follows: A: 11 B: 7 C: 6 D: 8 E: 14 F: 6 G: 9 H: 6 I: 7 J: 8 K: 11 CC for partial structure against native data = 9.92 % ------------------------------------------------------------------------------ Global autotracing cycle 6 Phases from autotracing cycle 4 used as input for final density modification = 0.281, Contrast = 0.324, Connect. = 0.613 for dens.mod. cycle 1 = 0.282, Contrast = 0.380, Connect. = 0.622 for dens.mod. cycle 2 = 0.282, Contrast = 0.442, Connect. = 0.671 for dens.mod. cycle 3 = 0.282, Contrast = 0.459, Connect. = 0.690 for dens.mod. cycle 4 = 0.282, Contrast = 0.469, Connect. = 0.702 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 0.523 0.586 0.607 0.556 0.515 0.417 0.372 0.372 0.326 0.275 0.723 0.769 0.818 0.766 0.729 0.622 0.566 0.619 0.555 0.494 N 2112 2093 2121 2158 2095 2054 2101 2200 2179 1901 Estimated mean FOM = 0.456 Pseudo-free CC = 51.00 % Anomalous density (in sigma units) at initial heavy atom sites Site x y z occ*Z density 1 1.0000 0.8192 1.0000 1.0000 51.03 2 1.2467 0.9324 1.0185 0.9174 21.16 3 0.8484 0.4312 1.0219 0.9103 21.84 4 0.6030 0.3143 1.0075 0.7025 16.31 5 0.8496 0.4814 1.1267 0.7022 17.38 6 1.2494 0.9599 1.1337 0.6510 14.37 7 1.0908 0.9224 1.0468 0.6484 16.08 8 0.9762 0.4462 0.9383 0.4877 10.52 9 1.1635 0.8442 0.9526 0.4329 10.74 10 0.9907 0.6588 0.9417 0.2869 8.14 11 0.6015 0.3171 0.8240 0.2807 4.66 Site x y z h(sig) near old near new 1 1.0000 0.8192 1.0000 51.0 1/0.00 11/2.67 11/2.67 12/2.71 12/2.71 2 0.8494 0.4312 1.0217 21.9 3/0.06 8/10.60 4/10.79 8/10.84 5/17.16 3 1.2464 0.9320 1.0186 21.2 2/0.04 15/2.79 6/9.55 9/10.72 7/11.17 4 0.8511 0.4810 1.1265 17.4 5/0.09 2/10.79 8/12.00 10/18.50 8/19.35 5 0.6057 0.3136 1.0078 16.4 4/0.17 5/12.28 2/17.16 4/22.82 8/25.09 6 1.0879 0.9216 1.0464 16.1 7/0.18 14/9.34 3/9.55 15/10.05 1/10.83 7 1.2477 0.9605 1.1335 14.4 6/0.11 3/11.17 6/12.80 15/13.91 9/20.28 8 0.9793 0.4483 0.9407 11.0 8/0.34 2/10.60 2/10.84 4/12.00 10/17.41 9 1.1645 0.8436 0.9530 10.8 9/0.09 15/8.59 14/9.74 11/10.51 1/10.68 10 0.9900 0.6579 0.9423 8.2 10/0.10 11/11.98 12/12.01 12/12.01 11/13.4 11 1.0034 0.7947 0.9819 -5.7 1/2.67 12/1.86 12/1.86 1/2.67 1/2.67 12 1.0000 0.7866 1.0000 -5.6 1/2.71 11/1.86 11/1.86 1/2.71 1/2.71 13 1.5000 0.8213 1.0000 -5.6 2/17.36 3/17.36 3/17.36 15/17.43 15/17.43 14 1.0000 0.8296 1.0276 -5.5 1/2.76 1/2.76 1/2.76 11/3.03 12/4.43 15 1.2362 0.9234 0.9909 -4.7 2/2.79 3/2.79 9/8.59 6/10.05 7/13.91 Best trace (cycle 4 with CC 10.74%) was saved as P21221_pse14_i.pdb ============================================================================== CPU times required in seconds ----------------------------- 7.9 - Setup, data input and phasing 1.2 - FFTs and peak-searches 4.7 - Sphere of influence 0.1 - Rest of density modification 0.0 - Alpha-helix search 50.3 - Tripeptide search 56.5 - Chain tracing 0.0 - NCS analysis 4.2 - B-value refinement for trace 0.1 - Rest of tracing 0.0 - Comparison with known structure ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++ + SHELXE finished at 12:16:51 Total time: 125.05 secs + ++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++