WHAT IF Check report

This file was created 2011-12-18 from WHAT_CHECK output by a conversion script. If you are new to WHAT_CHECK, please study the pdbreport pages. There also exists a legend to the output.

Please note that you are looking at an abridged version of the output (all checks that gave normal results have been removed from this report). You can have a look at the Full report instead.

Verification log for pdb6tmn.ent

Checks that need to be done early-on in validation

Warning: Ligands for which topology could not be determined

The ligands in the table below are too complicated for the automatic topology determination. WHAT IF uses a local copy of Daan van Aalten's Dundee PRODRG server to automatically generate topology information for ligands. Some molecules are too complicated for this software. If that happens, WHAT IF / WHAT-CHECK continue with a simplified topology that lacks certain information. Ligands with a simplified topology can, for example, not form hydrogen bonds, and that reduces the accuracy of all hydrogen bond related checking facilities.

The reason for topology generation failure is indicated. 'Atom types' indicates that the ligand contains atom types not known to PRODRUG. 'Attached' means that the ligand is covalently attached to a macromolecule. 'Size' indicates that the ligand has either too many atoms, or too many bonds, angles, or torsion angles. 'Fragmented' is written when the ligand is not one fully covalently connected molecule but consists of multiple fragments. 'N/O only' is given when the ligand contains only N and/or O atoms. 'OK' indicates that the automatic topology generation succeeded.

 318 0PI   ( 317-)  E  -         Atom types

Administrative problems that can generate validation failures

Warning: Duplicate atom names detected in ligands

The pairs of atoms listed in the table below sit in the same ligand, or co-factor, and have the same name.

The PDB holds a large series of co-factors that consist of multiple, identical building blocks. Often the names of atoms in these building blocks are made unique by adding an A,B,C, etc., in front of their name. For example, AO11, AO12, BO11, BO22 for 4 Oxygens. The limitations of the PDB format make it difficult (albeit not impossible) to avoid this. WHAT IF deals with this problem by stripping the leading A,B,C, etc., from the names. This way the atom names make chemical sense again, but they loose their uniqueness. Although this seems to generate few problems, it cannot be guaranteed that certain error messages are the direct or indirect result of this nomenclature problem. It is possible that these duplications are the result of earlier atom name corrections by WHAT IF.

 318 0PI   ( 317-)  E  -   O2   318 0PI   ( 317-)  E  -   O2

Non-validating, descriptive output paragraph

Note: Ramachandran plot

In this Ramachandran plot x-signs represent glycines, squares represent prolines, and plus-signs represent the other residues. If too many plus- signs fall outside the contoured areas then the molecule is poorly refined (or worse). Proline can only occur in the narrow region around phi=-60 that also falls within the other contour islands.

In a colour picture, the residues that are part of a helix are shown in blue, strand residues in red. Preferred regions for helical residues are drawn in blue, for strand residues in red, and for all other residues in green. A full explanation of the Ramachandran plot together with a series of examples can be found at the WHAT_CHECK website.

Chain identifier: E

Coordinate problems, unexpected atoms, B-factor and occupancy checks

Warning: What type of B-factor?

WHAT IF does not yet know well how to cope with B-factors in case TLS has been used. It simply assumes that the B-factor listed on the ATOM and HETATM cards are the total B-factors. When TLS refinement is used that assumption sometimes is not correct. TLS seems not mentioned in the header of the PDB file. But anyway, if WHAT IF complains about your B-factors, and you think that they are OK, then check for TLS related B-factor problems first.

Obviously, the temperature at which the X-ray data was collected has some importance too:

Temperature cannot be read from the PDB file. This most likely means that the temperature is listed as NULL (meaning unknown) in the PDB file.

Warning: More than 2 percent of buried atoms has low B-factor

For protein structures determined at room temperature, no more than about 1 percent of the B factors of buried atoms is below 5.0.

Percentage of buried atoms with B less than 5 : 4.06

Error: The B-factors of bonded atoms show signs of over-refinement

For each of the bond types in a protein a distribution was derived for the difference between the square roots of the B-factors of the two atoms. All bonds in the current protein were scored against these distributions. The number given below is the RMS Z-score over the structure. For a structure with completely restrained B-factors within residues, this value will be around 0.35, for extremely high resolution structures refined with free isotropic B-factors this number is expected to be near 1.0. Any value over 1.5 is sign of severe over-refinement of B-factors.

RMS Z-score : 1.799 over 2175 bonds
Average difference in B over a bond : 3.23
RMS difference in B over a bond : 4.33

Note: B-factor plot

The average atomic B-factor per residue is plotted as function of the residue number.

Chain identifier: E

Nomenclature related problems

Warning: Arginine nomenclature problem

The arginine residues listed in the table below have their N-H-1 and N-H-2 swapped.

  47 ARG   (  47-)  E

Warning: Tyrosine convention problem

The tyrosine residues listed in the table below have their chi-2 not between -90.0 and 90.0

  27 TYR   (  27-)  E
  28 TYR   (  28-)  E
  42 TYR   (  42-)  E
  46 TYR   (  46-)  E
  66 TYR   (  66-)  E
  75 TYR   (  75-)  E
  76 TYR   (  76-)  E
  81 TYR   (  81-)  E
  83 TYR   (  83-)  E
  84 TYR   (  84-)  E
 157 TYR   ( 157-)  E
 179 TYR   ( 179-)  E
 211 TYR   ( 211-)  E
 242 TYR   ( 242-)  E
 251 TYR   ( 251-)  E
 268 TYR   ( 268-)  E
 296 TYR   ( 296-)  E

Warning: Phenylalanine convention problem

The phenylalanine residues listed in the table below have their chi-2 not between -90.0 and 90.0.

  62 PHE   (  62-)  E
  63 PHE   (  63-)  E
 114 PHE   ( 114-)  E
 130 PHE   ( 130-)  E
 267 PHE   ( 267-)  E

Warning: Aspartic acid convention problem

The aspartic acid residues listed in the table below have their chi-2 not between -90.0 and 90.0, or their proton on OD1 instead of OD2.

  32 ASP   (  32-)  E
  37 ASP   (  37-)  E
  72 ASP   (  72-)  E
  94 ASP   (  94-)  E
 124 ASP   ( 124-)  E
 126 ASP   ( 126-)  E
 138 ASP   ( 138-)  E
 170 ASP   ( 170-)  E
 185 ASP   ( 185-)  E
 261 ASP   ( 261-)  E

Warning: Glutamic acid convention problem

The glutamic acid residues listed in the table below have their chi-3 outside the -90.0 to 90.0 range, or their proton on OE1 instead of OE2.

 119 GLU   ( 119-)  E
 166 GLU   ( 166-)  E
 187 GLU   ( 187-)  E

Geometric checks

Warning: Unusual bond lengths

The bond lengths listed in the table below were found to deviate more than 4 sigma from standard bond lengths (both standard values and sigmas for amino acid residues have been taken from Engh and Huber [REF], for DNA they were taken from Parkinson et al [REF]). In the table below for each unusual bond the bond length and the number of standard deviations it differs from the normal value is given.

Atom names starting with "-" belong to the previous residue in the chain. If the second atom name is "-SG*", the disulphide bridge has a deviating length.

  22 THR   (  22-)  E      CA   C     1.61    4.1
  53 SER   (  53-)  E      CB   OG    1.32   -4.9
  96 ASN   (  96-)  E      CG   OD1   1.32    4.6
 117 GLY   ( 117-)  E      CA   C     1.58    4.3
 150 ASP   ( 150-)  E      CG   OD2   1.37    6.6
 160 GLU   ( 160-)  E      CD   OE1   1.16   -4.7
 166 GLU   ( 166-)  E      CD   OE1   1.33    4.3
 170 ASP   ( 170-)  E      CG   OD1   1.35    5.2
 187 GLU   ( 187-)  E      CD   OE1   1.33    4.4
 203 ARG   ( 203-)  E      NE   CZ    1.39    4.1
 216 HIS   ( 216-)  E      CA   CB    1.62    4.4
 261 ASP   ( 261-)  E      CG   OD1   1.37    6.5
 308 GLN   ( 308-)  E      CG   CD    1.65    5.4
 308 GLN   ( 308-)  E      CD   OE1   1.37    6.9
 316 LYS   ( 316-)  E      CA   C     1.62    4.7

Warning: Possible cell scaling problem

Comparison of bond distances with Engh and Huber [REF] standard values for protein residues and Parkinson et al [REF] values for DNA/RNA shows a significant systematic deviation. It could be that the unit cell used in refinement was not accurate enough. The deformation matrix given below gives the deviations found: the three numbers on the diagonal represent the relative corrections needed along the A, B and C cell axis. These values are 1.000 in a normal case, but have significant deviations here (significant at the 99.99 percent confidence level)

There are a number of different possible causes for the discrepancy. First the cell used in refinement can be different from the best cell calculated. Second, the value of the wavelength used for a synchrotron data set can be miscalibrated. Finally, the discrepancy can be caused by a dataset that has not been corrected for significant anisotropic thermal motion.

Please note that the proposed scale matrix has NOT been restrained to obey the space group symmetry. This is done on purpose. The distortions can give you an indication of the accuracy of the determination.

If you intend to use the result of this check to change the cell dimension of your crystal, please read the extensive literature on this topic first. This check depends on the wavelength, the cell dimensions, and on the standard bond lengths and bond angles used by your refinement software.

Unit Cell deformation matrix

 |  0.992607  0.000308 -0.000676|
 |  0.000308  0.991525  0.000271|
 | -0.000676  0.000271  0.998741|
Proposed new scale matrix

 |  0.012362 -0.000004  0.000008|
 |  0.006178  0.010716  0.000001|
 |  0.000005 -0.000002  0.007620|
With corresponding cell

    A    =  93.356  B   =  93.302  C    = 131.241
    Alpha=  89.969  Beta=  90.083  Gamma= 119.948

The CRYST1 cell dimensions

    A    =  94.100  B   =  94.100  C    = 131.400
    Alpha=  90.000  Beta=  90.000  Gamma= 120.000

Variance: 373.258
(Under-)estimated Z-score: 14.239

Warning: Unusual bond angles

The bond angles listed in the table below were found to deviate more than 4 sigma from standard bond angles (both standard values and sigma for protein residues have been taken from Engh and Huber [REF], for DNA/RNA from Parkinson et al [REF]). In the table below for each strange angle the bond angle and the number of standard deviations it differs from the standard values is given. Please note that disulphide bridges are neglected. Atoms starting with "-" belong to the previous residue in the sequence.

   1 ILE   (   1-)  E      CA   CB   CG2 121.69    6.6
   1 ILE   (   1-)  E      CA   CB   CG1 102.76   -4.5
   2 THR   (   2-)  E     -O   -C    N   113.95   -5.7
   4 THR   (   4-)  E      C    CA   CB  118.98    4.7
   4 THR   (   4-)  E      CG2  CB   OG1  85.13  -12.1
   5 SER   (   5-)  E      N    CA   CB  120.65    6.0
   5 SER   (   5-)  E      CA   CB   OG  101.15   -5.0
   6 THR   (   6-)  E      CG2  CB   OG1  91.65   -8.8
   9 VAL   (   9-)  E      CA   CB   CG1 117.70    4.2
  13 VAL   (  13-)  E     -O   -C    N   116.47   -4.1
  14 LEU   (  14-)  E      CD1  CG   CD2  96.22   -6.6
  16 ASP   (  16-)  E      CB   CG   OD2 105.80   -5.5
  17 GLN   (  17-)  E      CG   CD   NE2 106.86   -6.4
  17 GLN   (  17-)  E      NE2  CD   OE1 128.34    5.7
  19 ASN   (  19-)  E      CA   CB   CG  108.06   -4.5
  21 ASN   (  21-)  E      CB   CG   ND2 123.38    4.7
  21 ASN   (  21-)  E      ND2  CG   OD1 114.07   -8.5
  26 THR   (  26-)  E      N    CA   CB  120.86    6.1
  26 THR   (  26-)  E      C    CA   CB  121.19    5.8
  26 THR   (  26-)  E      CA   CB   OG1 120.61    7.3
  26 THR   (  26-)  E      CG2  CB   OG1 119.42    5.1
  28 TYR   (  28-)  E      CA   C    O   113.98   -4.0
  29 TYR   (  29-)  E      CB   CG   CD1 114.35   -4.3
  32 ASP   (  32-)  E      CA   CB   CG  117.25    4.6
  33 ASN   (  33-)  E      ND2  CG   OD1 129.45    6.8
And so on for a total of 176 lines.

Warning: High bond angle deviations

Bond angles were found to deviate more than normal from the mean standard bond angles (normal values for protein residues were taken from Engh and Huber [REF], for DNA/RNA from Parkinson et al [REF]). The RMS Z-score given below is expected to be near 1.0 for a normally restrained data set, and this is indeed observed for very high resolution X-ray structures. The fact that it is higher than 2.0 in this structure might indicate that the restraints used in the refinement were not strong enough. This will also occur if a different bond angle dictionary is used.

RMS Z-score for bond angles: 2.017
RMS-deviation in bond angles: 3.675

Error: Nomenclature error(s)

Checking for a hand-check. WHAT IF has over the course of this session already corrected the handedness of atoms in several residues. These were administrative corrections. These residues are listed here.

  32 ASP   (  32-)  E
  37 ASP   (  37-)  E
  47 ARG   (  47-)  E
  72 ASP   (  72-)  E
  94 ASP   (  94-)  E
 119 GLU   ( 119-)  E
 124 ASP   ( 124-)  E
 126 ASP   ( 126-)  E
 138 ASP   ( 138-)  E
 166 GLU   ( 166-)  E
 170 ASP   ( 170-)  E
 185 ASP   ( 185-)  E
 187 GLU   ( 187-)  E
 261 ASP   ( 261-)  E

Warning: Chirality deviations detected

The atoms listed in the table below have an improper dihedral value that is deviating from expected values. As the improper dihedral values are all getting very close to ideal values in recent X-ray structures, and as we actually do not know how big the spread around these values should be, this check only warns for 6 sigma deviations.

Improper dihedrals are a measure of the chirality/planarity of the structure at a specific atom. Values around -35 or +35 are expected for chiral atoms, and values around 0 for planar atoms. Planar side chains are left out of the calculations, these are better handled by the planarity checks.

Three numbers are given for each atom in the table. The first is the Z-score for the improper dihedral. The second number is the measured improper dihedral. The third number is the expected value for this atom type. A final column contains an extra warning if the chirality for an atom is opposite to the expected value.

Please also see the previous table that lists a series of administrative chirality problems that were corrected automatically upon reading-in the PDB file.

   4 THR   (   4-)  E      CB     6.0    47.58    34.09
   6 THR   (   6-)  E      CB     6.7    49.14    34.09
   9 VAL   (   9-)  E      CB     6.9   -23.91   -32.96
  14 LEU   (  14-)  E      CG    -8.1   -47.35   -33.01
  26 THR   (  26-)  E      CA   -10.2    16.80    33.84
  26 THR   (  26-)  E      CB   -10.0    11.64    34.09
  65 SER   (  65-)  E      CA    -7.2    20.91    34.32
 156 ILE   ( 156-)  E      CB    -8.3    21.56    32.31
 197 ILE   ( 197-)  E      CB    -7.4    22.72    32.31
 202 LEU   ( 202-)  E      CG    -7.7   -46.55   -33.01
 220 ARG   ( 220-)  E      CA     6.7    44.93    33.91
 224 THR   ( 224-)  E      CB   -10.6    10.25    34.09
 230 VAL   ( 230-)  E      CB     9.7   -20.30   -32.96
 260 ARG   ( 260-)  E      CA   -10.3    16.97    33.91
 278 THR   ( 278-)  E      CB    -8.9    14.23    34.09
 316 LYS   ( 316-)  E      CA   -10.6    16.25    33.92
The average deviation= 2.427

Warning: High improper dihedral angle deviations

The RMS Z-score for the improper dihedrals in the structure is high. For well refined structures this number is expected to be near 1.0. The fact that it is higher than 2.0 worries us a bit. However, we determined the improper normal distribution from 500 high-resolution X-ray structures, rather than from CSD data, so we cannot be 100 percent certain about these numbers.

Improper dihedral RMS Z-score : 2.159

Error: Tau angle problems

The side chains of the residues listed in the table below contain a tau angle (N-Calpha-C) that was found to deviate from te expected value by more than 4.0 times the expected standard deviation. The number in the table is the number of standard deviations this RMS value deviates from the expected value.

 260 ARG   ( 260-)  E    6.50
 133 LEU   ( 133-)  E    6.36
 176 VAL   ( 176-)  E    4.79
 312 ALA   ( 312-)  E    4.34
 295 LEU   ( 295-)  E    4.23
   1 ILE   (   1-)  E    4.14
  68 ALA   (  68-)  E    4.10

Warning: High tau angle deviations

The RMS Z-score for the tau angles (N-Calpha-C) in the structure is too high. For well refined structures this number is expected to be near 1.0. The fact that it is higher than 1.5 worries us. However, we determined the tau normal distributions from 500 high-resolution X-ray structures, rather than from CSD data, so we cannot be 100 percent certain about these numbers.

Tau angle RMS Z-score : 1.572

Error: Side chain planarity problems

The side chains of the residues listed in the table below contain a planar group that was found to deviate from planarity by more than 4.0 times the expected value. For an amino acid residue that has a side chain with a planar group, the RMS deviation of the atoms to a least squares plane was determined. The number in the table is the number of standard deviations this RMS value deviates from the expected value. Not knowing better yet, we assume that planarity of the groups analyzed should be perfect.

  21 ASN   (  21-)  E   12.23
  74 HIS   (  74-)  E    8.22
  89 ASN   (  89-)  E    7.86
 142 HIS   ( 142-)  E    6.23
 200 ASP   ( 200-)  E    5.46
  37 ASP   (  37-)  E    5.11
 227 ASN   ( 227-)  E    4.98
 150 ASP   ( 150-)  E    4.52
  31 GLN   (  31-)  E    4.47
 213 ASP   ( 213-)  E    4.42
 226 ASP   ( 226-)  E    4.35
 251 TYR   ( 251-)  E    4.31
 105 HIS   ( 105-)  E    4.28
 211 TYR   ( 211-)  E    4.21
 122 TYR   ( 122-)  E    4.01

Torsion-related checks

Warning: Torsion angle evaluation shows unusual residues

The residues listed in the table below contain bad or abnormal torsion angles.

These scores give an impression of how `normal' the torsion angles in protein residues are. All torsion angles except omega are used for calculating a `normality' score. Average values and standard deviations were obtained from the residues in the WHAT IF database. These are used to calculate Z-scores. A residue with a Z-score of below -2.0 is poor, and a score of less than -3.0 is worrying. For such residues more than one torsion angle is in a highly unlikely position.

  26 THR   (  26-)  E    -3.5
 157 TYR   ( 157-)  E    -2.5
  20 ILE   (  20-)  E    -2.4
 278 THR   ( 278-)  E    -2.4
  46 TYR   (  46-)  E    -2.3
 182 LYS   ( 182-)  E    -2.3
  18 LYS   (  18-)  E    -2.3
 183 ASN   ( 183-)  E    -2.2
  92 SER   (  92-)  E    -2.2
 185 ASP   ( 185-)  E    -2.1
 107 SER   ( 107-)  E    -2.1

Warning: Backbone evaluation reveals unusual conformations

The residues listed in the table below have abnormal backbone torsion angles.

Residues with `forbidden' phi-psi combinations are listed, as well as residues with unusual omega angles (deviating by more than 3 sigma from the normal value). Please note that it is normal if about 5 percent of the residues is listed here as having unusual phi-psi combinations.

  26 THR   (  26-)  E  Poor phi/psi
  45 LYS   (  45-)  E  Poor phi/psi
  46 TYR   (  46-)  E  Poor phi/psi
  50 LEU   (  50-)  E  PRO omega poor
  60 ASN   (  60-)  E  Poor phi/psi
  89 ASN   (  89-)  E  Poor phi/psi
  92 SER   (  92-)  E  Poor phi/psi
  97 ASN   (  97-)  E  Poor phi/psi
 105 HIS   ( 105-)  E  Poor phi/psi
 107 SER   ( 107-)  E  Poor phi/psi
 118 SER   ( 118-)  E  Poor phi/psi
 130 PHE   ( 130-)  E  omega poor
 152 THR   ( 152-)  E  Poor phi/psi
 157 TYR   ( 157-)  E  Poor phi/psi
 159 ASN   ( 159-)  E  Poor phi/psi
 181 ASN   ( 181-)  E  Poor phi/psi
 188 ILE   ( 188-)  E  omega poor
 315 VAL   ( 315-)  E  omega poor
 chi-1/chi-2 correlation Z-score : -1.589

Warning: Unusual rotamers

The residues listed in the table below have a rotamer that is not seen very often in the database of solved protein structures. This option determines for every residue the position specific chi-1 rotamer distribution. Thereafter it verified whether the actual residue in the molecule has the most preferred rotamer or not. If the actual rotamer is the preferred one, the score is 1.0. If the actual rotamer is unique, the score is 0.0. If there are two preferred rotamers, with a population distribution of 3:2 and your rotamer sits in the lesser populated rotamer, the score will be 0.667. No value will be given if insufficient hits are found in the database.

It is not necessarily an error if a few residues have rotamer values below 0.3, but careful inspection of all residues with these low values could be worth it.

 291 SER   ( 291-)  E    0.36

Warning: Unusual backbone conformations

For the residues listed in the table below, the backbone formed by itself and two neighbouring residues on either side is in a conformation that is not seen very often in the database of solved protein structures. The number given in the table is the number of similar backbone conformations in the database with the same amino acid in the centre.

For this check, backbone conformations are compared with database structures using C-alpha superpositions with some restraints on the backbone oxygen positions.

A residue mentioned in the table can be part of a strange loop, or there might be something wrong with it or its directly surrounding residues. There are a few of these in every protein, but in any case it is worth looking at!

   5 SER   (   5-)  E      0
  14 LEU   (  14-)  E      0
  19 ASN   (  19-)  E      0
  24 TYR   (  24-)  E      0
  25 SER   (  25-)  E      0
  26 THR   (  26-)  E      0
  30 LEU   (  30-)  E      0
  34 THR   (  34-)  E      0
  37 ASP   (  37-)  E      0
  45 LYS   (  45-)  E      0
  46 TYR   (  46-)  E      0
  49 THR   (  49-)  E      0
  51 PRO   (  51-)  E      0
  53 SER   (  53-)  E      0
  55 TRP   (  55-)  E      0
  60 ASN   (  60-)  E      0
  61 GLN   (  61-)  E      0
  62 PHE   (  62-)  E      0
  63 PHE   (  63-)  E      0
  88 HIS   (  88-)  E      0
  89 ASN   (  89-)  E      0
  91 LEU   (  91-)  E      0
  92 SER   (  92-)  E      0
  93 TYR   (  93-)  E      0
  96 ASN   (  96-)  E      0
And so on for a total of 120 lines.

Warning: Unusual PRO puckering amplitudes

The proline residues listed in the table below have a puckering amplitude that is outside of normal ranges. Puckering parameters were calculated by the method of Cremer and Pople [REF]. Normal PRO rings have a puckering amplitude Q between 0.20 and 0.45 Angstrom. If Q is lower than 0.20 Angstrom for a PRO residue, this could indicate disorder between the two different normal ring forms (with C-gamma below and above the ring, respectively). If Q is higher than 0.45 Angstrom something could have gone wrong during the refinement. Be aware that this is a warning with a low confidence level. See: Who checks the checkers? Four validation tools applied to eight atomic resolution structures [REF]

 184 PRO   ( 184-)  E    0.17 LOW
 195 PRO   ( 195-)  E    0.20 LOW
 208 PRO   ( 208-)  E    0.18 LOW

Bump checks

Error: Abnormally short interatomic distances

The pairs of atoms listed in the table below have an unusually short interactomic distance; each bump is listed in only one direction.

The contact distances of all atom pairs have been checked. Two atoms are said to `bump' if they are closer than the sum of their Van der Waals radii minus 0.40 Angstrom. For hydrogen bonded pairs a tolerance of 0.55 Angstrom is used. The first number in the table tells you how much shorter that specific contact is than the acceptable limit. The second distance is the distance between the centres of the two atoms. Although we believe that two water atoms at 2.4 A distance are too close, we only report water pairs that are closer than this rather short distance.

The last text-item on each line represents the status of the atom pair. If the final column contains the text 'HB', the bump criterion was relaxed because there could be a hydrogen bond. Similarly relaxed criteria are used for 1-3 and 1-4 interactions (listed as 'B2' and 'B3', respectively). BL indicates that the B-factors of the clashing atoms have a low B-factor thereby making this clash even more worrisome. INTRA and INTER indicate whether the clashes are between atoms in the same asymmetric unit, or atoms in symmetry related asymmetric units, respectively.

 269 ARG   ( 269-)  E      NH1 <->  294 ASP   ( 294-)  E      OD2    0.54    2.16  INTRA
 116 ASN   ( 116-)  E      O   <->  324 HOH   ( 323 )  E      O      0.34    2.06  INTRA
 194 THR   ( 194-)  E      CA  <->  195 PRO   ( 195-)  E      CD     0.20    2.60  INTRA BL
 115 TRP   ( 115-)  E      NE1 <->  150 ASP   ( 150-)  E      OD2    0.17    2.53  INTRA
 262 LYS   ( 262-)  E      NZ  <->  302 GLU   ( 302-)  E      OE2    0.16    2.54  INTRA
  81 TYR   (  81-)  E      OH  <->   97 ASN   (  97-)  E      ND2    0.16    2.54  INTRA BL
   5 SER   (   5-)  E      C   <->    6 THR   (   6-)  E      CG2    0.15    2.95  INTRA
 149 THR   ( 149-)  E      O   <->  154 GLY   ( 154-)  E      N      0.14    2.56  INTRA BL
 251 TYR   ( 251-)  E      N   <->  324 HOH   ( 409 )  E      O      0.13    2.57  INTRA BL
 220 ARG   ( 220-)  E      CD  <->  324 HOH   ( 480 )  E      O      0.12    2.68  INTRA
 126 ASP   ( 126-)  E      OD1 <->  128 GLN   ( 128-)  E      N      0.12    2.58  INTRA BL
 308 GLN   ( 308-)  E      O   <->  312 ALA   ( 312-)  E      N      0.11    2.59  INTRA BL
 274 TYR   ( 274-)  E      OH  <->  294 ASP   ( 294-)  E      OD2    0.11    2.29  INTRA
 194 THR   ( 194-)  E      N   <->  195 PRO   ( 195-)  E      CD     0.10    2.90  INTRA BL
 308 GLN   ( 308-)  E      NE2 <->  324 HOH   ( 370 )  E      O      0.09    2.61  INTRA BF
  66 TYR   (  66-)  E      O   <->  105 HIS   ( 105-)  E      NE2    0.08    2.62  INTRA BL
 194 THR   ( 194-)  E      O   <->  197 ILE   ( 197-)  E      N      0.08    2.62  INTRA
 167 ALA   ( 167-)  E      O   <->  171 ILE   ( 171-)  E      N      0.07    2.63  INTRA BL
 189 GLY   ( 189-)  E      N   <->  190 GLU   ( 190-)  E      N      0.06    2.54  INTRA BL
 285 ARG   ( 285-)  E      CD  <->  316 LYS   ( 316-)  E      CD     0.06    3.14  INTRA
 182 LYS   ( 182-)  E      NZ  <->  191 ASP   ( 191-)  E      OD2    0.06    2.64  INTRA BL
 196 GLY   ( 196-)  E      N   <->  324 HOH   ( 428 )  E      O      0.05    2.65  INTRA
 194 THR   ( 194-)  E      C   <->  196 GLY   ( 196-)  E      N      0.04    2.86  INTRA
 206 SER   ( 206-)  E      O   <->  239 LYS   ( 239-)  E      NZ     0.04    2.66  INTRA BL
 250 HIS   ( 250-)  E      ND1 <->  324 HOH   ( 410 )  E      O      0.04    2.66  INTRA
 126 ASP   ( 126-)  E      N   <->  127 GLY   ( 127-)  E      N      0.03    2.57  INTRA BL
 252 GLY   ( 252-)  E      N   <->  324 HOH   ( 408 )  E      O      0.03    2.67  INTRA BL
 195 PRO   ( 195-)  E      C   <->  196 GLY   ( 196-)  E      CA     0.03    2.27  INTRA B2
 257 GLY   ( 257-)  E      N   <->  324 HOH   ( 379 )  E      O      0.02    2.68  INTRA
  45 LYS   (  45-)  E      NZ  <->   53 SER   (  53-)  E      OG     0.02    2.68  INTRA
 224 THR   ( 224-)  E      C   <->  225 GLN   ( 225-)  E      CA     0.01    2.29  INTRA B2
 269 ARG   ( 269-)  E      NH1 <->  294 ASP   ( 294-)  E      CG     0.01    3.09  INTRA
 126 ASP   ( 126-)  E      OD1 <->  129 THR   ( 129-)  E      N      0.01    2.69  INTRA BL
 186 TRP   ( 186-)  E      O   <->  205 MET   ( 205-)  E      N      0.01    2.69  INTRA BL

Packing, accessibility and threading

Note: Inside/Outside RMS Z-score plot

The Inside/Outside distribution normality RMS Z-score over a 15 residue window is plotted as function of the residue number. High areas in the plot (above 1.5) indicate unusual inside/outside patterns.

Chain identifier: E

Warning: Abnormal packing environment for some residues

The residues listed in the table below have an unusual packing environment.

The packing environment of the residues is compared with the average packing environment for all residues of the same type in good PDB files. A low packing score can indicate one of several things: Poor packing, misthreading of the sequence through the density, crystal contacts, contacts with a co-factor, or the residue is part of the active site. It is not uncommon to see a few of these, but in any case this requires further inspection of the residue.

 108 GLN   ( 108-)  E      -5.92
  88 HIS   (  88-)  E      -5.87
 251 TYR   ( 251-)  E      -5.56
 246 GLN   ( 246-)  E      -5.52
 273 GLN   ( 273-)  E      -5.50
 221 TYR   ( 221-)  E      -5.36
 225 GLN   ( 225-)  E      -5.07

Warning: Abnormal packing environment for sequential residues

A stretch of at least three sequential residues with a questionable packing environment was found. This could indicate that these residues are part of a strange loop. It might also be an indication of misthreading in the density. However, it can also indicate that one or more residues in this stretch have other problems such as, for example, missing atoms, very weird angles or bond lengths, etc.

The table below lists the first and last residue in each stretch found, as well as the average residue score of the series.

 225 GLN   ( 225-)  E       228 - GLY    228- ( E)         -4.48

Note: Quality value plot

The quality value smoothed over a 10 residue window is plotted as function of the residue number. Low areas in the plot (below -2.0) indicate unusual packing.

Chain identifier: E

Warning: Low packing Z-score for some residues

The residues listed in the table below have an unusual packing environment according to the 2nd generation packing check. The score listed in the table is a packing normality Z-score: positive means better than average, negative means worse than average. Only residues scoring less than -2.50 are listed here. These are the unusual residues in the structure, so it will be interesting to take a special look at them.

  44 ALA   (  44-)  E   -2.76

Note: Second generation quality Z-score plot

The second generation quality Z-score smoothed over a 10 residue window is plotted as function of the residue number. Low areas in the plot (below -1.3) indicate unusual packing.

Chain identifier: E

Water, ion, and hydrogenbond related checks

Warning: Water molecules need moving

The water molecules listed in the table below were found to be significantly closer to a symmetry related non-water molecule than to the ones given in the coordinate file. For optimal viewing convenience revised coordinates for these water molecules should be given.

The number in brackets is the identifier of the water molecule in the input file. Suggested coordinates are also given in the table. Please note that alternative conformations for protein residues are not taken into account for this calculation. If you are using WHAT IF / WHAT-CHECK interactively, then the moved waters can be found in PDB format in the file: MOVEDH2O.pdb.

 324 HOH   ( 391 )  E      O     69.27    6.86   -5.46
 324 HOH   ( 427 )  E      O     74.44    4.99  -11.57
 324 HOH   ( 443 )  E      O     29.66   38.98   13.61
 324 HOH   ( 445 )  E      O     52.40   30.55   20.18

Error: Water molecules without hydrogen bonds

The water molecules listed in the table below do not form any hydrogen bonds, neither with the protein or DNA/RNA, nor with other water molecules. This is a strong indication of a refinement problem. The last number on each line is the identifier of the water molecule in the input file.

 324 HOH   ( 492 )  E      O
Metal-coordinating Histidine residue 142 fixed to   1
Metal-coordinating Histidine residue 146 fixed to   1

Error: HIS, ASN, GLN side chain flips

Listed here are Histidine, Asparagine or Glutamine residues for which the orientation determined from hydrogen bonding analysis are different from the assignment given in the input. Either they could form energetically more favourable hydrogen bonds if the terminal group was rotated by 180 degrees, or there is no assignment in the input file (atom type 'A') but an assignment could be made. Be aware, though, that if the topology could not be determined for one or more ligands, then this option will make errors.

  31 GLN   (  31-)  E
  33 ASN   (  33-)  E
  96 ASN   (  96-)  E
 181 ASN   ( 181-)  E
 280 ASN   ( 280-)  E
 290 GLN   ( 290-)  E
 301 GLN   ( 301-)  E

Warning: Buried unsatisfied hydrogen bond donors

The buried hydrogen bond donors listed in the table below have a hydrogen atom that is not involved in a hydrogen bond in the optimized hydrogen bond network.

Hydrogen bond donors that are buried inside the protein normally use all of their hydrogens to form hydrogen bonds within the protein. If there are any non hydrogen bonded buried hydrogen bond donors in the structure they will be listed here. In very good structures the number of listed atoms will tend to zero.

Waters are not listed by this option.

  35 ARG   (  35-)  E      N
  49 THR   (  49-)  E      N
  49 THR   (  49-)  E      OG1
  97 ASN   (  97-)  E      ND2
 112 ASN   ( 112-)  E      N
 155 LEU   ( 155-)  E      N
 203 ARG   ( 203-)  E      NH2
 206 SER   ( 206-)  E      N
 216 HIS   ( 216-)  E      N
 237 ILE   ( 237-)  E      N
 269 ARG   ( 269-)  E      NH1
 302 GLU   ( 302-)  E      N
Only metal coordination for  142 HIS  ( 142-) E      NE2
Only metal coordination for  146 HIS  ( 146-) E      NE2
Only metal coordination for  190 GLU  ( 190-) E      OE2

Warning: Buried unsatisfied hydrogen bond acceptors

The buried side-chain hydrogen bond acceptors listed in the table below are not involved in a hydrogen bond in the optimized hydrogen bond network.

Side-chain hydrogen bond acceptors buried inside the protein normally form hydrogen bonds within the protein. If there are any not hydrogen bonded in the optimized hydrogen bond network they will be listed here.

Waters are not listed by this option.

 143 GLU   ( 143-)  E      OE2
 231 HIS   ( 231-)  E      NE2
 238 ASN   ( 238-)  E      OD1

Warning: No crystallisation information

No, or very inadequate, crystallisation information was observed upon reading the PDB file header records. This information should be available in the form of a series of REMARK 280 lines. Without this information a few things, such as checking ions in the structure, cannot be performed optimally.

Warning: Unusual ion packing

We implemented the ion valence determination method of Brown and Wu [REF] similar to Nayal and Di Cera [REF]. See also Mueller, Koepke and Sheldrick [REF]. It must be stated that the validation of ions in PDB files is very difficult. Ideal ion-ligand distances often differ no more than 0.1 Angstrom, and in a 2.0 Angstrom resolution structure 0.1 Angstrom is not very much. Nayal and Di Cera showed that this method has great potential, but the method has not been validated. Part of our implementation (comparing ion types) is even fully new and despite that we see it work well in the few cases that are trivial, we must emphasize that this validation method is untested. See: swift.cmbi.ru.nl/teach/theory/ for a detailed explanation.

The output gives the ion, the valency score for the ion itself, the valency score for the suggested alternative ion, and a series of possible comments *1 indicates that the suggested alternate atom type has been observed in the PDB file at another location in space. *2 indicates that WHAT IF thinks to have found this ion type in the crystallisation conditions as described in the REMARK 280 cards of the PDB file. *S Indicates that this ions is located at a special position (i.e. at a symmetry axis). N4 stands for NH4+.

 319  CA   ( 318-)  E   -.-  -.-  Part of ionic cluster
 319  CA   ( 318-)  E     0.73   0.96 Scores about as good as NA
 320  CA   ( 319-)  E   -.-  -.-  Part of ionic cluster
 321  CA   ( 320-)  E     0.89   1.13 Scores about as good as NA

Warning: Unusual water packing

We implemented the ion valence determination method of Brown and Wu [REF] similar to Nayal and Di Cera [REF] and Mueller, Koepke and Sheldrick [REF]. It must be stated that the validation of ions in PDB files is very difficult. Ideal ion-ligand distances often differ no more than 0.1 Angstrom, and in a 2.0 Angstrom resolution structure 0.1 Angstrom is not very much. Nayal and Di Cera showed that this method nevertheless has great potential for detecting water molecules that actually should be metal ions. The method has not been extensively validated, though. Part of our implementation (comparing waters with multiple ion types) is even fully new and despite that we see it work well in the few cases that are trivial, we must emphasize that this method is untested.

The score listed is the valency score. This number should be close to (preferably a bit above) 1.0 for the suggested ion to be a likely alternative for the water molecule. Ions listed in brackets are good alternate choices. *1 indicates that the suggested ion-type has been observed elsewhere in the PDB file too. *2 indicates that the suggested ion-type has been observed in the REMARK 280 cards of the PDB file. Ion-B and ION-B indicate that the B-factor of this water is high, or very high, respectively. H2O-B indicates that the B-factors of atoms that surround this water/ion are suspicious. See: swift.cmbi.ru.nl/teach/theory/ for a detailed explanation.

 324 HOH   ( 474 )  E      O  0.88  K  4 ION-B

Warning: Possible wrong residue type

The residues listed in the table below have a weird environment that cannot be improved by rotamer flips. This can mean one of three things, non of which WHAT CHECK really can do much about. 1) The side chain has actually another rotamer than is present in the PDB file; 2) A counter ion is present in the structure but is not given in the PDB file; 3) The residue actually is another amino acid type. The annotation 'Alt-rotamer' indicates that WHAT CHECK thinks you might want to find an alternate rotamer for this residue. The annotation 'Sym-induced' indicates that WHAT CHECK believes that symmetry contacts might have something to do with the difficulties of this residue's side chain. Determination of these two annotations is difficult, so their absence is less meaningful than their presence. The annotation Ligand-bound indicates that a ligand seems involved with this residue. In nine of ten of these cases this indicates that the ligand is causing the weird situation rather than the residue.

  37 ASP   (  37-)  E   H-bonding suggests Asn
 119 GLU   ( 119-)  E   H-bonding suggests Gln
 294 ASP   ( 294-)  E   H-bonding suggests Asn; but Alt-Rotamer

Final summary

Note: Summary report for users of a structure

This is an overall summary of the quality of the structure as compared with current reliable structures. This summary is most useful for biologists seeking a good structure to use for modelling calculations.

The second part of the table mostly gives an impression of how well the model conforms to common refinement restraint values. The first part of the table shows a number of global quality indicators.


Structure Z-scores, positive is better than average:

  1st generation packing quality :  -1.085
  2nd generation packing quality :  -1.468
  Ramachandran plot appearance   :  -0.475
  chi-1/chi-2 rotamer normality  :  -1.589
  Backbone conformation          :  -0.418

RMS Z-scores, should be close to 1.0:
  Bond lengths                   :   1.334
  Bond angles                    :   2.017 (loose)
  Omega angle restraints         :   0.823
  Side chain planarity           :   2.827 (loose)
  Improper dihedral distribution :   2.159 (loose)
  B-factor distribution          :   1.799 (loose)
  Inside/Outside distribution    :   1.022

Note: Summary report for depositors of a structure

This is an overall summary of the quality of the X-ray structure as compared with structures solved at similar resolutions. This summary can be useful for a crystallographer to see if the structure makes the best possible use of the data. Warning. This table works well for structures solved in the resolution range of the structures in the WHAT IF database, which is presently (summer 2008) mainly 1.1 - 1.3 Angstrom. The further the resolution of your file deviates from this range the more meaningless this table becomes.

The second part of the table mostly gives an impression of how well the model conforms to common refinement restraint values. The first part of the table shows a number of global quality indicators, which have been calibrated against structures of similar resolution.

Resolution found in PDB file : 1.60


Structure Z-scores, positive is better than average:

  1st generation packing quality :  -0.9
  2nd generation packing quality :  -1.4
  Ramachandran plot appearance   :  -0.9
  chi-1/chi-2 rotamer normality  :  -2.0
  Backbone conformation          :  -0.8

RMS Z-scores, should be close to 1.0:
  Bond lengths                   :   1.334
  Bond angles                    :   2.017 (loose)
  Omega angle restraints         :   0.823
  Side chain planarity           :   2.827 (loose)
  Improper dihedral distribution :   2.159 (loose)
  B-factor distribution          :   1.799 (loose)
  Inside/Outside distribution    :   1.022
==============

WHAT IF
    G.Vriend,
      WHAT IF: a molecular modelling and drug design program,
    J. Mol. Graph. 8, 52--56 (1990).

WHAT_CHECK (verification routines from WHAT IF)
    R.W.W.Hooft, G.Vriend, C.Sander and E.E.Abola,
      Errors in protein structures
    Nature 381, 272 (1996).
    (see also http://swift.cmbi.ru.nl/gv/whatcheck for a course and extra inform

Bond lengths and angles, protein residues
    R.Engh and R.Huber,
      Accurate bond and angle parameters for X-ray protein structure
      refinement,
    Acta Crystallogr. A47, 392--400 (1991).

Bond lengths and angles, DNA/RNA
    G.Parkinson, J.Voitechovsky, L.Clowney, A.T.Bruenger and H.Berman,
      New parameters for the refinement of nucleic acid-containing structures
    Acta Crystallogr. D52, 57--64 (1996).

DSSP
    W.Kabsch and C.Sander,
      Dictionary of protein secondary structure: pattern
      recognition of hydrogen bond and geometrical features
    Biopolymers 22, 2577--2637 (1983).

Hydrogen bond networks
    R.W.W.Hooft, C.Sander and G.Vriend,
      Positioning hydrogen atoms by optimizing hydrogen bond networks in
      protein structures
    PROTEINS, 26, 363--376 (1996).

Matthews' Coefficient
    B.W.Matthews
      Solvent content of Protein Crystals
    J. Mol. Biol. 33, 491--497 (1968).

Protein side chain planarity
    R.W.W. Hooft, C. Sander and G. Vriend,
      Verification of protein structures: side-chain planarity
    J. Appl. Cryst. 29, 714--716 (1996).

Puckering parameters
    D.Cremer and J.A.Pople,
      A general definition of ring puckering coordinates
    J. Am. Chem. Soc. 97, 1354--1358 (1975).

Quality Control
    G.Vriend and C.Sander,
      Quality control of protein models: directional atomic
      contact analysis,
    J. Appl. Cryst. 26, 47--60 (1993).

Ramachandran plot
    G.N.Ramachandran, C.Ramakrishnan and V.Sasisekharan,
      Stereochemistry of Polypeptide Chain Conformations
    J. Mol. Biol. 7, 95--99 (1963).

Symmetry Checks
    R.W.W.Hooft, C.Sander and G.Vriend,
      Reconstruction of symmetry related molecules from protein
      data bank (PDB) files
    J. Appl. Cryst. 27, 1006--1009 (1994).

Ion Checks
    I.D.Brown and K.K.Wu,
      Empirical Parameters for Calculating Cation-Oxygen Bond Valences
    Acta Cryst. B32, 1957--1959 (1975).

    M.Nayal and E.Di Cera,
      Valence Screening of Water in Protein Crystals Reveals Potential Na+
      Binding Sites
    J.Mol.Biol. 256 228--234 (1996).

    P.Mueller, S.Koepke and G.M.Sheldrick,
      Is the bond-valence method able to identify metal atoms in protein
      structures?
    Acta Cryst. D 59 32--37 (2003).

Checking checks
    K.Wilson, C.Sander, R.W.W.Hooft, G.Vriend, et al.
      Who checks the checkers
    J.Mol.Biol. (1998) 276,417-436.