WHAT IF Check report

This file was created 2012-10-23 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 pdb4aph.ent

Checks that need to be done early-on in validation

Warning: Ligands for which a topology was generated automatically

The topology for the ligands in the table below were determined automatically. WHAT IF uses a local copy of Daan van Aalten's Dundee PRODRG server to automatically generate topology information for ligands. For this PDB file that seems to have gone fine, but be aware that automatic topology generation is a complicated task. So, if you get messages that you fail to understand or that you believe are wrong, and one of these ligands is involved, then check the ligand topology first.

 603 ACT   (1634-)  A  -
 604 ACT   (1635-)  A  -
 605 ACT   (1636-)  A  -
 606 ACT   (1637-)  A  -
 607 ACT   (1638-)  A  -
 608 BMA   (1633-)  A  -
 609 PE4   (1629-)  A  -

Administrative problems that can generate validation failures

Warning: Overlapping residues or molecules

This molecule contains residues or molecules that overlap too much while not being (administrated as) alternate atom/residue pairs. The residues or molecules listed in the table below have been removed before the validation continued.

Overlapping residues or molecules (for short entities) are occasionally observed in the PDB. Often these are cases like, for example, two sugars that bind equally well in the same active site, are both seen overlapping in the density, and are both entered in the PDB file as separate entities. This can cause some false positive error messsages further down the validation path, and therefore the second of the overlapping entities has been deleted before the validation continued. If you want to validate both situations, make it two PDB files, one for each sugar. And fudge reality a bit by making the occupancy of the sugar atoms 1.0 in both cases, because many validation options are not executed on atoms with low occupancy. If you go for this two-file option, please make sure that any side chains that have alternate locations depending on the sugar bound are selected in each of the two cases in agreement with the sugar that you keep for validation in that particular file.

 583 ASP   (   1-)  P  -
 591 VAL   (   3-)  P  -
 592 TYR   (   4-)  P  -
 593 ILE   (   5-)  P  -
 594 HIS   (   6-)  P  -
 595 PRO   (   7-)  P  -

Warning: Groups attached to potentially hydrogenbonding atoms

Residues were observed with groups attached to (or very near to) atoms that potentially can form hydrogen bonds. WHAT IF is not very good at dealing with such exceptional cases (Mainly because it's author is not...). So be warned that the hydrogenbonding-related analyses of these residues might be in error.

For example, an aspartic acid can be protonated on one of its delta oxygens. This is possible because the one delta oxygen 'helps' the other one holding that proton. However, if a delta oxygen has a group bound to it, then it can no longer 'help' the other delta oxygen bind the proton. However, both delta oxygens, in principle, can still be hydrogen bond acceptors. Such problems can occur in the amino acids Asp, Glu, and His. I have opted, for now to simply allow no hydrogen bonds at all for any atom in any side chain that somewhere has a 'funny' group attached to it. I know this is wrong, but there are only 12 hours in a day.

 585 VAL   (   3-)  P  - B N   bound to  583 ASP   (   1-)  P  - A C
 592 NAG   (1631-)  A  -   O4  bound to  593 NAG   (1632-)  A  -   C1
 593 NAG   (1632-)  A  -   O4  bound to  602 BMA   (1633-)  A  -   C1

Warning: Residues with missing backbone atoms.

Residues were detected with missing backbone atoms. This can be a normal result of poor or missing density, but it can also be an error.

In X-ray the coordinates must be located in density. Mobility or disorder sometimes cause this density to be so poor that the positions of the atoms cannot be determined. Crystallographers tend to leave out the atoms in such cases. This is not an error, albeit that we would prefer them to give it their best shot and provide coordinates with an occupancy of zero in cases where only a few atoms are involved. Anyway, several checks depend on the presence of the backbone atoms, so if you find errors in, or directly adjacent to, residues with missing backbone atoms, then please check by hand what is going on.

 574 PRO   ( 617-)  A  -

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: A

Note: Ramachandran plot

Chain identifier: P

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

Warning: Missing atoms

The atoms listed in the table below are missing from the entry. If many atoms are missing, the other checks can become less sensitive. Be aware that it often happens that groups at the termini of DNA or RNA are really missing, so that the absence of these atoms normally is neither an error nor the result of poor electron density. Some of the atoms listed here might also be listed by other checks, most noticeably by the options in the previous section that list missing atoms in several categories. The plausible atoms with zero occupancy are not listed here, as they already got assigned a non-zero occupancy, and thus are no longer 'missing'.

 574 PRO   ( 617-)  A      O
 583 ASP   (   1-)  P      CG
 583 ASP   (   1-)  P      OD1
 583 ASP   (   1-)  P      OD2

Warning: C-terminal nitrogen atoms detected.

It is becoming habit to indicate that a residue is not the true C-terminus by including only the backbone N of the next residue. This has been observed in this PDB file.

In X-ray the coordinates must be located in density. Mobility or disorder sometimes cause this density to be so poor that the positions of the atoms cannot be determined. Crystallographers tend to leave out the atoms in such cases. In many cases the N- or C-terminal residues are too disordered to see. In case of the N-terminus, you can see from the residue numbers if there are missing residues, but at the C-terminus this is impossible. Therefore, often the position of the backbone nitrogen of the first residue missing at the C-terminal end is calculated and added to indicate that there are missing residues. As a single N causes validation trouble, we remove these single-N-residues before doing the validation. But, if you get weird errors at, or near, the left-over incomplete C-terminal residue, please check by hand if a missing Oxt or removed N is the cause.

 590 PHE   (   8-)  P
 584 PRO   (   7-)  P

Warning: Occupancies atoms do not add up to 1.0.

In principle, the occupancy of all alternates of one atom should add up till 1.0. A valid exception is the missing atom (i.e. an atom not seen in the electron density) that is allowed to have a 0.0 occupancy. Sometimes this even happens when there are no alternate atoms given...

Atoms want to move. That is the direct result of the second law of thermodynamics, in a somewhat weird way of thinking. Any way, many atoms seem to have more than one position where they like to sit, and they jump between them. The population difference between those sites (which is related to their energy differences) is seen in the occupancy factors. As also for atoms it is 'to be or not to be', these occupancies should add up to 1.0. Obviously, it is possible that they add up to a number less than 1.0, in cases where there are yet more, but undetected' rotamers/positions in play, but also in those cases a warning is in place as the information shown in the PDB file is less certain than it could have been. The residues listed below contain atoms that have an occupancy greater than zero, but all their alternates do not add up to one.

WARNING. Presently WHAT CHECK only deals with a maximum of two alternate positions. A small number of atoms in the PDB has three alternates. In those cases the warning given here should obviously be neglected! In a next release we will try to fix this.

 583 ASP   (   1-)  P    0.50
 584 VAL   (   3-)  P    0.50
 585 TYR   (   4-)  P    0.50
 586 ILE   (   5-)  P    0.50
 587 HIS   (   6-)  P    0.50
 588 PRO   (   7-)  P    0.50
 589 NAG   (1630-)  A    0.50

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. The header of the PDB file states that TLS groups were used. So, if WHAT IF complains about your B-factors, while 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:


Number of TLS groups mentione in PDB file header: 1

Crystal temperature (K) :100.000

Note: B-factor plot

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

Chain identifier: A

Geometric checks

Warning: Low bond length variability

Bond lengths were found to deviate less than normal from the mean Engh and Huber [REF] and/or Parkinson et al [REF] standard bond lengths. The RMS Z-score given below is expected to be near 1.0 for a normally restrained data set. The fact that it is lower than 0.667 in this structure might indicate that too-strong restraints have been used in the refinement. This can only be a problem for high resolution X-ray structures.

RMS Z-score for bond lengths: 0.356
RMS-deviation in bond distances: 0.008

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

 |  0.997120 -0.000013 -0.000073|
 | -0.000013  0.997322  0.000432|
 | -0.000073  0.000432  0.997123|
Proposed new scale matrix

 |  0.017750  0.000000  0.000001|
 |  0.000000  0.011844 -0.000005|
 |  0.000000 -0.000003  0.007486|
With corresponding cell

    A    =  56.338  B   =  84.433  C    = 133.591
    Alpha=  89.950  Beta=  90.005  Gamma=  90.001

The CRYST1 cell dimensions

    A    =  56.500  B   =  84.660  C    = 133.970
    Alpha=  90.000  Beta=  90.000  Gamma=  90.000

Variance: 162.816
(Under-)estimated Z-score: 9.404

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.

 425 ARG   ( 468-)  A      CB   CG   CD  105.26   -4.4

Warning: Low bond angle variability

Bond angles were found to deviate less than normal from the 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. The fact that it is lower than 0.667 in this structure might indicate that too-strong restraints have been used in the refinement. This can only be a problem for high resolution X-ray structures.

RMS Z-score for bond angles: 0.556
RMS-deviation in bond angles: 1.140

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.

 542 PRO   ( 585-)  A    -2.9
 438 TYR   ( 481-)  A    -2.7
 124 PRO   ( 163-)  A    -2.5
 586 ILE   (   5-)  P    -2.3
 355 TYR   ( 394-)  A    -2.3
 478 ILE   ( 521-)  A    -2.3
  84 GLU   ( 123-)  A    -2.1
 474 SER   ( 517-)  A    -2.1
 201 LEU   ( 240-)  A    -2.0

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.

  33 ASN   (  72-)  A  Poor phi/psi
  84 GLU   ( 123-)  A  Poor phi/psi
 113 CYS   ( 152-)  A  omega poor
 123 GLU   ( 162-)  A  PRO omega poor
 124 PRO   ( 163-)  A  omega poor
 157 PHE   ( 196-)  A  omega poor
 232 PRO   ( 271-)  A  omega poor
 317 ALA   ( 356-)  A  omega poor
 321 TYR   ( 360-)  A  Poor phi/psi
 366 ALA   ( 405-)  A  omega poor
 392 ASN   ( 431-)  A  Poor phi/psi
 511 GLN   ( 554-)  A  Poor phi/psi
 538 ILE   ( 581-)  A  omega poor
 543 ASN   ( 586-)  A  omega poor
 chi-1/chi-2 correlation Z-score : -1.093

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.

 180 SER   ( 219-)  A    0.37
  39 SER   (  78-)  A    0.39

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!

  32 THR   (  71-)  A      0
  69 GLN   ( 108-)  A      0
  84 GLU   ( 123-)  A      0
  85 ARG   ( 124-)  A      0
 114 HIS   ( 153-)  A      0
 115 PRO   ( 154-)  A      0
 116 ASN   ( 155-)  A      0
 122 LEU   ( 161-)  A      0
 123 GLU   ( 162-)  A      0
 125 ASP   ( 164-)  A      0
 133 SER   ( 172-)  A      0
 149 LYS   ( 188-)  A      0
 175 VAL   ( 214-)  A      0
 186 GLU   ( 225-)  A      0
 220 TYR   ( 259-)  A      0
 224 HIS   ( 263-)  A      0
 234 HIS   ( 273-)  A      0
 235 LEU   ( 274-)  A      0
 240 TRP   ( 279-)  A      0
 242 GLN   ( 281-)  A      0
 243 THR   ( 282-)  A      0
 244 TRP   ( 283-)  A      0
 245 SER   ( 284-)  A      0
 247 ILE   ( 286-)  A      0
 251 VAL   ( 290-)  A      0
And so on for a total of 163 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]

 454 PRO   ( 497-)  A    0.18 LOW

Warning: Unusual PRO puckering phases

The proline residues listed in the table below have a puckering phase that is not expected to occur in protein structures. Puckering parameters were calculated by the method of Cremer and Pople [REF]. Normal PRO rings approximately show a so-called envelope conformation with the C-gamma atom above the plane of the ring (phi=+72 degrees), or a half-chair conformation with C-gamma below and C-beta above the plane of the ring (phi=-90 degrees). If phi deviates strongly from these values, this is indicative of a very strange conformation for a PRO residue, and definitely requires a manual check of the data. 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].

 124 PRO   ( 163-)  A   -52.6 half-chair C-beta/C-alpha (-54 degrees)
 258 PRO   ( 297-)  A   -65.1 envelop C-beta (-72 degrees)
 273 PRO   ( 312-)  A   111.4 envelop C-beta (108 degrees)
 294 PRO   ( 333-)  A   106.3 envelop C-beta (108 degrees)
 542 PRO   ( 585-)  A   -38.6 envelop C-alpha (-36 degrees)

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.

 591 NAG   (1632-)  A      O4  <->  600 BMA   (1633-)  A      C1     0.95    1.45  INTRA B3
 591 NAG   (1632-)  A      C4  <->  600 BMA   (1633-)  A      C1     0.85    2.35  INTRA
 425 ARG   ( 468-)  A      NH2 <->  470 HIS   ( 513-)  A      O      0.27    2.43  INTRA BL
 348 HIS   ( 387-)  A      ND1 <->  371 HIS   ( 410-)  A      ND1    0.24    2.76  INTRA BL
 156 GLN   ( 195-)  A      NE2 <->  602 HOH   (2091 )  A      O      0.20    2.50  INTRA
 226 ASN   ( 265-)  A      ND2 <->  575 GLN   ( 618-)  A      NE2    0.18    2.67  INTRA
  33 ASN   (  72-)  A      ND2 <->  589 NAG   (1630-)  A      C2     0.15    2.05  INTRA B2
 464 ASP   ( 507-)  A      N   <->  465 PRO   ( 508-)  A      CD     0.15    2.85  INTRA BL
 468 LYS   ( 511-)  A      NZ  <->  596 ACT   (1635-)  A      CH3    0.11    2.99  INTRA
 400 ASP   ( 443-)  A      OD1 <->  505 HIS   ( 548-)  A      NE2    0.10    2.60  INTRA
 121 GLN   ( 160-)  A      NE2 <->  602 HOH   (2058 )  A      O      0.09    2.61  INTRA
  46 ASN   (  85-)  A      ND2 <->  598 ACT   (1637-)  A      O2     0.08    2.62  INTRA
 469 PHE   ( 512-)  A      N   <->  596 ACT   (1635-)  A      CH3    0.08    3.02  INTRA
 531 TRP   ( 574-)  A      N   <->  532 PRO   ( 575-)  A      CD     0.08    2.92  INTRA BL
 399 HIS   ( 442-)  A      ND1 <->  602 HOH   (2170 )  A      O      0.06    2.64  INTRA
 263 THR   ( 302-)  A      OG1 <->  602 HOH   (2173 )  A      O      0.06    2.34  INTRA
 123 GLU   ( 162-)  A      O   <->  304 LYS   ( 343-)  A      NZ     0.06    2.64  INTRA
 203 PRO   ( 242-)  A      O   <->  207 ASN   ( 246-)  A      ND2    0.06    2.64  INTRA BL
  33 ASN   (  72-)  A      ND2 <->  589 NAG   (1630-)  A      N2     0.05    2.55  INTRA B3
 427 ARG   ( 470-)  A      NH2 <->  441 GLU   ( 484-)  A      OE1    0.05    2.65  INTRA
 439 ASN   ( 482-)  A      N   <->  602 HOH   (2233 )  A      O      0.05    2.65  INTRA BL
 411 LYS   ( 454-)  A      NZ  <->  602 HOH   (2218 )  A      O      0.05    2.65  INTRA BL
 264 GLU   ( 303-)  A      O   <->  268 LYS   ( 307-)  A      N      0.04    2.66  INTRA
 240 TRP   ( 279-)  A      CH2 <->  596 ACT   (1635-)  A      C      0.04    3.16  INTRA
 446 ARG   ( 489-)  A      NH1 <->  450 GLN   ( 493-)  A      OE1    0.03    2.67  INTRA BL
 217 HIS   ( 256-)  A      ND1 <->  602 HOH   (2144 )  A      O      0.03    2.67  INTRA
 581 ASN   ( 624-)  A      N   <->  602 HOH   (2152 )  A      O      0.02    2.68  INTRA
 321 TYR   ( 360-)  A      OH  <->  583 ASP   (   1-)  P    A N      0.02    2.68  INTRA
 121 GLN   ( 160-)  A      OE1 <->  304 LYS   ( 343-)  A      NZ     0.02    2.68  INTRA
 443 TRP   ( 486-)  A      NE1 <->  464 ASP   ( 507-)  A      OD2    0.02    2.68  INTRA BL
 596 ACT   (1635-)  A      O2  <->  602 HOH   (2070 )  A      O      0.02    2.38  INTRA
 224 HIS   ( 263-)  A      ND1 <->  602 HOH   (2153 )  A      O      0.01    2.69  INTRA BL
 314 HIS   ( 353-)  A      NE2 <->  587 HIS   (   6-)  P    B O      0.01    2.69  INTRA
 214 ARG   ( 253-)  A      NE  <->  569 GLU   ( 612-)  A      OE2    0.01    2.69  INTRA
 244 TRP   ( 283-)  A      N   <->  409 LEU   ( 452-)  A      O      0.01    2.69  INTRA BL
 213 ARG   ( 252-)  A      NH2 <->  570 LYS   ( 613-)  A      O      0.01    2.69  INTRA
 581 ASN   ( 624-)  A      N   <->  582 SER   ( 625-)  A      N      0.01    2.59  INTRA B3

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: A

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.

 567 HIS   ( 610-)  A      -6.16
 570 LYS   ( 613-)  A      -5.87
 116 ASN   ( 155-)  A      -5.82
 460 GLN   ( 503-)  A      -5.76
 581 ASN   ( 624-)  A      -5.70
 576 TYR   ( 619-)  A      -5.69
 219 HIS   ( 258-)  A      -5.55
 223 GLN   ( 262-)  A      -5.51
  67 GLN   ( 106-)  A      -5.49
 450 GLN   ( 493-)  A      -5.16
 587 HIS   (   6-)  P      -5.15
 227 LEU   ( 266-)  A      -5.02

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.

 218 ARG   ( 257-)  A       220 - TYR    259- ( A)         -4.75

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: A

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.

 184 MET   ( 223-)  A   -3.05
  83 LEU   ( 122-)  A   -2.91

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: A

Water, ion, and hydrogenbond related checks

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.

 602 HOH   (2019 )  A      O
 602 HOH   (2113 )  A      O
 602 HOH   (2176 )  A      O
 602 HOH   (2177 )  A      O
 602 HOH   (2223 )  A      O
 602 HOH   (2276 )  A      O
 602 HOH   (2277 )  A      O
 602 HOH   (2280 )  A      O
Bound group on Asn; dont flip   33 ASN  (  72-) A
Bound to:  589 NAG  (1630-) A
Bound group on Asn; dont flip   70 ASN  ( 109-) A
Bound to:  590 NAG  (1631-) A
Marked this atom as acceptor  592  CL  (1626-) A     CL
Marked this atom as acceptor  594  CL  (1628-) A     CL
Metal-coordinating Histidine residue 344 fixed to   1
Metal-coordinating Histidine residue 348 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.

  52 HIS   (  91-)  A
  67 GLN   ( 106-)  A
 209 HIS   ( 248-)  A
 389 HIS   ( 428-)  A
 567 HIS   ( 610-)  A
 575 GLN   ( 618-)  A
 581 ASN   ( 624-)  A

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.

  46 ASN   (  85-)  A      ND2
  75 ARG   ( 114-)  A      NH2
  85 ARG   ( 124-)  A      NH1
 176 ASP   ( 215-)  A      N
 226 ASN   ( 265-)  A      ND2
 234 HIS   ( 273-)  A      N
 234 HIS   ( 273-)  A      ND1
 238 ASN   ( 277-)  A      N
 243 THR   ( 282-)  A      N
 245 SER   ( 284-)  A      N
 272 THR   ( 311-)  A      N
 309 ARG   ( 348-)  A      NE
 324 LYS   ( 363-)  A      N
 325 ASP   ( 364-)  A      N
 361 ALA   ( 400-)  A      N
 367 ASN   ( 406-)  A      N
 376 ASP   ( 415-)  A      N
 399 HIS   ( 442-)  A      N
 406 LYS   ( 449-)  A      NZ
 417 PHE   ( 460-)  A      N
 434 THR   ( 477-)  A      OG1
 446 ARG   ( 489-)  A      NE
 456 VAL   ( 499-)  A      N
 458 ARG   ( 501-)  A      NH2
 479 ARG   ( 522-)  A      N
 481 PHE   ( 524-)  A      N
 483 SER   ( 526-)  A      OG
 536 GLN   ( 579-)  A      NE2
 579 THR   ( 622-)  A      N
Only metal coordination for  344 HIS  ( 383-) A      NE2
Only metal coordination for  348 HIS  ( 387-) A      NE2
Only metal coordination for  372 GLU  ( 411-) A      OE1

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.

 148 ASP   ( 187-)  A      OD1
 234 HIS   ( 273-)  A      NE2
 264 GLU   ( 303-)  A      OE2
 314 HIS   ( 353-)  A      NE2
 330 GLN   ( 369-)  A      OE1
 345 GLU   ( 384-)  A      OE1
 345 GLU   ( 384-)  A      OE2
 587 HIS   (   6-)  P    B ND1

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+.

 593  ZN   (1627-)  A   -.-  -.-  Too few ligands (3)

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.

 148 ASP   ( 187-)  A   H-bonding suggests Asn; but Alt-Rotamer
 193 ASP   ( 232-)  A   H-bonding suggests Asn; but Alt-Rotamer
 264 GLU   ( 303-)  A   H-bonding suggests Gln
 282 ASP   ( 321-)  A   H-bonding suggests Asn; but Alt-Rotamer
 376 ASP   ( 415-)  A   H-bonding suggests Asn; but Alt-Rotamer; Ligand-contact
 430 ASP   ( 473-)  A   H-bonding suggests Asn

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 :  -0.065
  2nd generation packing quality :  -1.075
  Ramachandran plot appearance   :   0.458
  chi-1/chi-2 rotamer normality  :  -1.093
  Backbone conformation          :   0.014

RMS Z-scores, should be close to 1.0:
  Bond lengths                   :   0.356 (tight)
  Bond angles                    :   0.556 (tight)
  Omega angle restraints         :   0.906
  Side chain planarity           :   0.288 (tight)
  Improper dihedral distribution :   0.516
  B-factor distribution          :   0.335
  Inside/Outside distribution    :   1.005

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 : 1.99


Structure Z-scores, positive is better than average:

  1st generation packing quality :   0.3
  2nd generation packing quality :  -0.7
  Ramachandran plot appearance   :   0.9
  chi-1/chi-2 rotamer normality  :  -0.3
  Backbone conformation          :  -0.2

RMS Z-scores, should be close to 1.0:
  Bond lengths                   :   0.356 (tight)
  Bond angles                    :   0.556 (tight)
  Omega angle restraints         :   0.906
  Side chain planarity           :   0.288 (tight)
  Improper dihedral distribution :   0.516
  B-factor distribution          :   0.335
  Inside/Outside distribution    :   1.005
==============

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.