WHAT IF Check report

This file was created 2013-03-27 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.

Verification log for /data/tmp/pdb\_redo/1kyt/wctemp/1kyt\_besttls.pdb

Checks that need to be done early-on in validation

Note: Non crystallographic symmetry RMS plot

The plot shows the RMS differences between two similar chains on a residue- by-residue basis. Individual "spikes" can be indicative of interesting or wrong residues. If all residues show a high RMS value, the structure could be incorrectly refined.

Chain identifiers of the two chains: A and B

All-atom RMS fit for the two chains : 0.931
CA-only RMS fit for the two chains : 0.519

Note: Non crystallographic symmetry backbone difference plot

The plot shows the differences in backbone torsion angles between two similar chains on a residue-by-residue basis. Individual "spikes" can be indicative of interesting or wrong residues. If all residues show high differences, the structure could be incorrectly refined.

Chain identifiers of the two chains: A and B

Note: No crystallographic symmetry between molecules

No extra crystallographic symmetry was observed between the independent molecules.

Warning: Problem detected upon counting molecules and matrices

The parameter Z as given on the CRYST card represents the molecular multiplicity in the crystallographic cell. Normally, Z equals the number of matrices of the space group multiplied by the number of NCS relations. The value of Z is multiplied by the integrated molecular weight of the molecules in the file to determine the Matthews coefficient. This relation is being validated in this option. Be aware that the validation can get confused if both multiple copies of the molecule are present in the ATOM records and MTRIX records are present in the header of the PDB file.

Space group as read from CRYST card: I 2 2 2
Number of matrices in space group: 8
Highest polymer chain multiplicity in structure: 2
Highest polymer chain multiplicity according to SEQRES: 1
No explicit MTRIX NCS matrices found in the input file
Value of Z as found on the CRYST1 card: 0
Polymer chain multiplicity and SEQRES multiplicity disagree 2 1
Z agrees neither with the 3D multiplicity, nor with the SEQRES multiplicity
It could be that Z must be one of: 8 16

Error: Matthews Coefficient (Vm) very high

The Matthews coefficient [REF] is defined as the density of the protein structure in cubic Angstroms per Dalton. Normal values are between 1.5 (tightly packed, little room for solvent) and 4.0 (loosely packed, much space for solvent). Some very loosely packed structures can get values a bit higher than that.

Numbers this high are almost always caused by giving the wrong value for Z on the CRYST1 card (or not giving this number at all).

Molecular weight of all polymer chains: 48733.262
Volume of the Unit Cell V= 1001255.8
Space group multiplicity: 8
No NCS symmetry matrices (MTRIX records) found in PDB file
Matthews coefficient for observed atoms and Z is high: Vm= 20.546
Matthews coefficient read from REMARK 280 Vm= 2.400
Vm by authors and this calculated Vm do not agree very well
SEQRES and ATOM multiplicities disagree. Error-reasoning thus is difficult.
(and the absence of MTRIX records doesn't help)
Could it be that Z must be: 8
This number is the multiplication of the spacegroup and NCS symmetry count
Matthews coefficient for observed atoms and corrected Z: Vm= 2.568

Note: Z missing on CRYST1 card

The messages above seem likely caused by the fact that Z is missing from the CRYST1 card.

Note: No atoms with high occupancy detected at special positions

Either there were no atoms at special positions, or all atoms at special positions have adequately reduced occupancies. An atom is considered to be located at a special position if it is within 0.3 Angstrom from one of its own symmetry copies. See also the next check...

Note: All atoms are sufficiently far away from symmetry axes

None of the atoms in the structure is closer than 0.77 Angstrom to a proper symmetry axis.

Note: Chain identifiers OK

WHAT CHECK has not detected any serious chain identifier problems. But be aware that WHAT CHECK doesn't care about the chain identifiers of waters.

Administrative problems that can generate validation failures

Note: No strange inter-chain connections detected

No covalent bonds have been detected between molecules with non-identical chain identifiers.

Note: No duplicate atom names in ligands

All atom names in ligands (if any) seem adequately unique.

Note: In all cases the primary alternate atom was used

WHAT CHECK saw no need to make any alternate atom corrections (which means they are all correct, or there are none).

Note: No residues detected inside ligands

Either this structure does not contain ligands with amino acid groups inside it, or their naming is proper (enough).

Note: No attached groups interfere with hydrogen bond calculations

It seems there are no sugars, lipids, etc., bound (or very close) to atoms that otherwise could form hydrogen bonds.

Note: No probable side chain atoms with zero occupancy detected.

Either there are no side chain atoms with zero occupancy, or the side chain atoms with zero occupancy were not present in the input PDB file (in which case they are listed as missing atoms), or their positions are sufficiently improbable to warrant a zero occupancy.

Note: No probable backbone atoms with zero occupancy detected.

Either there are no backbone atoms with zero occupancy, or the backbone atoms with zero occupancy were not present in the input PDB file (in which case they are listed as missing atoms), or their positions are sufficiently improbable to warrant a zero occupancy.

Note: All residues have a complete backbone.

No residues have missing backbone atoms.

Note: No C-alpha only residues

There are no residues that consist of only an alpha carbon atom.

Warning: Non-canonical residue(s)

WHAT CHECK has detected non-canonical residue(s). OK indicates that WHAT CHECK does have topology information for it, but be aware that many options were callibrated using only the twenty canonical residue types. The label HARD is used for non-canonical residues not yet known to WHAT CHECK.

   2 MSE   (   1-)  A  -         OK
  71 MSE   (  70-)  A  -         OK
  94 MSE   (  93-)  A  -         OK
 100 MSE   (  99-)  A  -         OK
 146 MSE   ( 145-)  A  -         OK
 162 MSE   ( 161-)  A  -         OK
 180 MSE   ( 179-)  A  -         OK
 182 MSE   ( 181-)  A  -         OK
 225 MSE   ( 224-)  A  -         OK
 227 MSE   (   1-)  B  -         OK
 296 MSE   (  70-)  B  -         OK
 319 MSE   (  93-)  B  -         OK
 325 MSE   (  99-)  B  -         OK
 371 MSE   ( 145-)  B  -         OK
 387 MSE   ( 161-)  B  -         OK
 405 MSE   ( 179-)  B  -         OK
 407 MSE   ( 181-)  B  -         OK
 450 MSE   ( 224-)  B  -         OK

Non-validating, descriptive output paragraph

Note: Content of the PDB file as interpreted by WHAT CHECK

Content of the PDB file as interpreted by WHAT CHECK. WHAT CHECK has read your PDB file, and stored it internally in what is called 'the soup'. The content of this soup is listed here. An extensive explanation of all frequently used WHAT CHECK output formats can be found at swift.cmbi.ru.nl. Look under output formats. A course on reading this 'Molecules' table is part of the WHAT CHECK website.

     1     1 (    0)   225 (  224) A Protein             1kyt.besttls.pdb
     2   226 (    0)   450 (  224) B Protein             1kyt.besttls.pdb
     3   451 (  224)   451 (  224) A M O2 <-   225       1kyt.besttls.pdb
     4   452 (  502)   452 (  502) A CA                  1kyt.besttls.pdb
     5   453 (  503)   453 (  503) A CA                  1kyt.besttls.pdb
     6   454 (  505)   454 (  505) A CA                  1kyt.besttls.pdb
     7   455 (  506)   455 (  506) A CA                  1kyt.besttls.pdb
     8   456 (  501)   456 (  501) B CA                  1kyt.besttls.pdb
     9   457 (  504)   457 (  504) B CA                  1kyt.besttls.pdb
    10   458 (  224)   458 (  224) B M O2 <-   450       1kyt.besttls.pdb
    11   459 ( HOH )   459 ( HOH ) A water   (  223)     1kyt.besttls.pdb
    12   460 ( HOH )   460 ( HOH ) B water   (  214)     1kyt.besttls.pdb

Note: Some notes regarding the PDB file contents

The numbers and remarks listed below have no explicit validation purpose, they are merely meant for the crystallographer or NMR spectroscopists to perhaps pinpoint something unexpected. See the WHAT_CHECK course for an explanation of terms like 'poor', 'missing', etcetera (in case those words pop up in the lines underneath this message).

The total number of amino acids found is 450
of which 6 have poor or missing atoms
Number of water molecules: 437
of which one is poor
2 Residues (protein, nucleic acid, sugars) have residue number zero

Note: Chain identifiers seem OK

All ions seem to have a logical chain identifier, or there are no ions present in the input file.

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: B

Note: Secondary structure

This is the secondary structure according to DSSP. Only helix (H), overwound or 3/10-helix (3), strand (S), turn (T) and coil (blank) are shown [REF]. All DSSP related information can be found at the DSSP page This is not really a structure validation option, but a very scattered secondary structure (i.e. many strands of only a few residues length, many Ts inside helices, etc) tends to indicate a poor structure. A full explanation of the DSSP secondary structure determination program together with a series of examples can be found at the WHAT_CHECK website.
                     10        20        30        40        50        60
                      |         |         |         |         |         |
    1 -   60 HMIRLAAIDVDGNLTDRDRLISTKAIESIRSAEKKGLTVSLLSGNVIPVVYALKIFLGIN
(   0)-(  59)    SSSSSHHHHT  TTT   HHHHHHHHHHHHTT SSSSS TT HHHHHHHHHHHT
                     70        80        90       100       110       120
                      |         |         |         |         |         |
   61 -  120 GPVFGENGGIMFDNDGSIKKFFSNEGTNKFLEEMSKRTSMRSILTNRWREASTGFDIDPE
(  60)-( 119)T SSS333TSSS TTT SSS T THHHHHHHHHHTTTTT    33333 TTTSSS   33
                    130       140       150       160       170       180
                      |         |         |         |         |         |
  121 -  180 DVDYVRKEAESRGFVIFYSGYSWHLMNRGEDKAFAVNKLKEMYSLEYDEILVIGDSNNDM
( 120)-( 179)3HHHHHHHHHTTTSSSSSSTTSSSSSSTT THHHHHHHHHHHTT  333SSSS  T333H
                    190       200       210       220
                      |         |         |         |
  181 -  225 PMFQLPVRKACPANATDNIKAVSDFVSDYSYGEEIGQIFKHFELM
( 180)-( 224)HHHTTTTSSSS TTT HHHHHH TSS T  TTHHHHHHHHHTT
 
               230       240       250       260       270       280
                 |         |         |         |         |         |
  226 -  285 HMIRLAAIDVDGNLTDRDRLISTKAIESIRSAEKKGLTVSLLSGNVIPVVYALKIFLGIN
(   0)-(  59)    SSSSSTTTTT  TTT   HHHHHHHHHHHHTT SSSSS TT HHHHHHHHHHHT
               290       300       310       320       330       340
                 |         |         |         |         |         |
  286 -  345 GPVFGENGGIMFDNDGSIKKFFSNEGTNKFLEEMSKRTSMRSILTNRWREASTGFDIDPE
(  60)-( 119)T SSS333TSSS TTT SSS T THHHHHHHHHHTTTTT    33333 TTTSSS   33
               350       360       370       380       390       400
                 |         |         |         |         |         |
  346 -  405 DVDYVRKEAESRGFVIFYSGYSWHLMNRGEDKAFAVNKLKEMYSLEYDEILVIGDSNNDM
( 120)-( 179)3HHHHHHHHHTTTSSSSSSTTSSSSSSTT THHHHHHHHHHHTT  333SSSS  T333H
               410       420       430       440       450
                 |         |         |         |         |
  406 -  450 PMFQLPVRKACPANATDNIKAVSDFVSDYSYGEEIGQIFKHFELM
( 180)-( 224)HHHTTTTSSSS TTT HHHHHH TSS T  TTHHHHHHHHHTT
 
 
 

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

Note: No rounded coordinates detected

No significant rounding of atom coordinates has been detected.

Note: No artificial side chains detected

No artificial side-chain positions characterized by chi-1=0.0 or chi-1=180.0 have been detected.

Note: No missing atoms detected in residues

All expected atoms are present in residues. This validation option has not looked at 'things' that can or should be attached to the elemantary building blocks (amino acids, nucleotides). Even the C-terminal oxygens are treated separately.

Warning: B-factors outside the range 0.0 - 100.0

In principle, B-factors can have a very wide range of values, but in practice, B-factors should not be zero while B-factors above 100.0 are a good indicator that the location of that atom is meaningless. Be aware that the cutoff at 100.0 is arbitrary. 'High' indicates that atoms with a B-factor > 100.0 were observed; 'Zero' indicates that atoms with a B-factor of zero were observed.

  35 LYS   (  34-)  A    High
  92 GLU   (  91-)  A    High
 146 MSE   ( 145-)  A    High
 220 LYS   ( 219-)  A    High
 244 ARG   (  18-)  B    High
 448 GLU   ( 222-)  B    High

Note: No C-terminal nitrogen detected

The PDB indicates that a residue is not the true C-terminus by including only the backbone N of the next residue. This has not been observed in this PDB file.

Note: C-terminus capping

The residues listed in the table below either are pseudo C-terminal residues, or have two groups attached of which neither is the normal C-terminal O. In this table REAL means that the C-terminal residue is likely to be the real C-terminus of its chain; OX means that an incorrect second oxygen (OXT) was detected that should not be there; -O indicates that the 'normal' oxygen (i.e. not the OXT) is missing; OT indicates the detection of any other capping group. C-terminal nitrogen atoms, if any, have already been dealt with in a previous check and are indicated here by -N. PSEUDO means that this is the last visible residue in the chain, but not the real C-terminus, i.e. all residues after this one are missing in this chain. BREAK means that this is the last residue before a chain-break, i.e. the chain continues but after this residue a number of residues is missing. In case a break is observed the number of residues that seems to be missing is shown in brackets. OK means that given the status (REAL, PSEUDO, BREAK), no problems were found.

Be aware that we cannot easily see the difference between these errors and errors in the chain and residue numbering schemes. So do not blindly trust the table below.

 225 MSE   ( 224-)  A          : Unknown problem
 450 MSE   ( 224-)  B          : Unknown problem

Note: No OXT found in the middle of chains

No OXT groups were found in the middle of protein chains.

Note: Weights administratively correct

All atomic occupancy factors ('weights') fall in the 0.0-1.0 range, which makes them administratively correct.

Note: Normal distribution of occupancy values

The distribution of the occupancy values in this file seems 'normal'.

Be aware that this evaluation is merely the result of comparing this file with about 500 well-refined high-resolution files in the PDB. If this file has much higher or much lower resolution than the PDB files used in WHAT CHECK's training set, non-normal values might very well be perfectly fine, or normal values might actually be not so normal. So, this check is actually more an indicator and certainly not a check in which I have great confidence.

Note: All occupancies seem to add up to 0.0 - 1.0.

In principle, the occupancy of all alternates of one atom should add up till 0.0 - 1.0. 0.0 is used for the missing atom (i.e. an atom not seen in the electron density). Obviously, there is nothing terribly wrong when a few occupancies add up to a bit more than 1.0, because the mathematics of refinement allow for that. However, if it happens often, it seems worth evaluating this in light of the refinement protocol used.

Warning: What type of B-factor?

WHAT CHECK 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 CHECK complains about your B-factors, while you think that they are OK, then check for TLS related B-factor problems first.


Number of TLS groups mentione in PDB file header: 0

Temperature not mentioned in PDB file. This most likely means that the temperature record is absent.
Room temperature assumed

Warning: More than 5 percent of buried atoms has low B-factor

For normal protein structures, no more than about 1 percent of the B factors of buried atoms is below 5.0. The fact that this value is much higher in the current structure could be a signal that the B-factors were restraints or constraints to too-low values, misuse of B-factor field in the PDB file, or a TLS/scaling problem. If the average B factor is low too, it is probably a low temperature structure determination.

Percentage of buried atoms with B less than 5 : 84.17

Note: B-factor plot

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

Chain identifier: A

Note: B-factor plot

Chain identifier: B

Nomenclature related problems

Note: Introduction to the nomenclature section.

Nomenclature problems seem, at first, rather unimportant. After all who cares if we call the delta atoms in leucine delta2 and delta1 rather than the other way around. Chemically speaking that is correct. But structures have not been solved and deposited just for chemists to look at them. Most times a structure is used, it is by software in a bioinformatics lab. And if they compare structures in which the one used C delta1 and delta2 and the other uses C delta2 and delta1, then that comparison will fail. Also, we recalculate all structures every so many years to make sure that everybody always can get access to the best coordinates that can be obtained from the (your?) experimental data. These recalculations will be troublesome if there are nomenclature problems.

Several nomenclature problems actually are worse than that. At the WHAT_CHECK website you can get an overview of the importance of all nomenclature problems that we list.

Note: Valine nomenclature OK

No errors were detected in valine nomenclature.

Note: Threonine nomenclature OK

No errors were detected in threonine nomenclature.

Note: Isoleucine nomenclature OK

No errors were detected in isoleucine nomenclature.

Note: Leucine nomenclature OK

No errors were detected in leucine nomenclature.

Note: Arginine nomenclature OK

No errors were detected in arginine nomenclature.

Note: Tyrosine torsion conventions OK

No errors were detected in tyrosine torsion angle conventions.

Warning: Phenylalanine convention problem

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

 137 PHE   ( 136-)  A
 379 PHE   ( 153-)  B

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.

  11 ASP   (  10-)  A
 168 ASP   ( 167-)  A
 422 ASP   ( 196-)  B

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.

 128 GLU   ( 127-)  A
 213 GLU   ( 212-)  A
 310 GLU   (  84-)  B

Note: Phosphate group names OK in DNA/RNA

No errors were detected in nucleic acid phosphate group naming conventions.

Note: Heavy atom naming OK

No errors were detected in the atom names for non-hydrogen atoms. Please be aware that the PDB wants us to deliberately make some nomenclature errors; especially in non-canonical amino acids.

Note: No decreasing residue numbers

All residue numbers are strictly increasing within each chain.

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.

 175 ASP   ( 174-)  A      CG   OD1   1.16   -4.4
 400 ASP   ( 174-)  B      CG   OD1   1.16   -4.4

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: 0.622
RMS-deviation in bond distances: 0.021

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

 |  1.000189 -0.000839  0.000226|
 | -0.000839  0.998057 -0.000332|
 |  0.000226 -0.000332  0.997863|
Proposed new scale matrix

 |  0.011294  0.000009 -0.000003|
 |  0.000008  0.010089  0.000003|
 | -0.000002  0.000003  0.008800|
With corresponding cell

    A    =  88.544  B   =  99.122  C    = 113.639
    Alpha=  90.000  Beta=  90.000  Gamma=  90.000

The CRYST1 cell dimensions

    A    =  88.526  B   =  99.316  C    = 113.886
    Alpha=  90.000  Beta=  90.000  Gamma=  90.000

Variance: 33.568
(Under-)estimated Z-score: 4.270

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.

 225 MSE   ( 224-)  A      N    CA   CB  100.06   -6.1
 332 ARG   ( 106-)  B      CG   CD   NE  117.41    4.1
 400 ASP   ( 174-)  B      CB   CG   OD2 127.65    4.0

Note: Normal bond angle variability

Bond angles were found to deviate normally 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.

RMS Z-score for bond angles: 0.746
RMS-deviation in bond angles: 1.555

Error: Nomenclature error(s)

Checking for a hand-check. WHAT CHECK 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.

  11 ASP   (  10-)  A
 128 GLU   ( 127-)  A
 168 ASP   ( 167-)  A
 213 GLU   ( 212-)  A
 310 GLU   (  84-)  B
 422 ASP   ( 196-)  B

Note: Chirality OK

All protein atoms have proper chirality. But, look at the previous table to see a series of administrative chirality problems that were corrected automatically upon reading-in the PDB file.

Note: Improper dihedral angle distribution OK

The RMS Z-score for all improper dihedrals in the structure is within normal ranges.

Improper dihedral RMS Z-score : 0.904

Note: Tau angles OK

All of the tau angles (N-Calpha-C) of amino acids fall within expected RMS deviations.

Note: Normal tau angle deviations

The RMS Z-score for the tau angles (N-Calpha-C) in the structure falls within the normal range that we guess to be 0.5 - 1.5. Be aware, 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 : 0.958

Note: Side chain planarity OK

All of the side chains of residues that have an intact planar group are planar within expected RMS deviations.

Note: Atoms connected to aromatic rings OK

All of the atoms that are connected to planar aromatic rings in side chains of amino-acid residues are in the plane within expected RMS deviations.

Torsion-related checks

Note: Ramachandran Z-score OK

The score expressing how well the backbone conformations of all residues correspond to the known allowed areas in the Ramachandran plot is within expected ranges for well-refined structures.

Ramachandran Z-score : -0.193

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 CHECK 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.

 448 GLU   ( 222-)  B    -2.5
 223 GLU   ( 222-)  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.

  19 ARG   (  18-)  A  Poor phi/psi
  67 ASN   (  66-)  A  Poor phi/psi
 110 GLU   ( 109-)  A  Poor phi/psi
 142 SER   ( 141-)  A  omega poor
 165 LEU   ( 164-)  A  omega poor
 175 ASP   ( 174-)  A  Poor phi/psi
 211 TYR   ( 210-)  A  Poor phi/psi
 223 GLU   ( 222-)  A  Poor phi/psi
 244 ARG   (  18-)  B  Poor phi/psi
 292 ASN   (  66-)  B  Poor phi/psi
 335 GLU   ( 109-)  B  Poor phi/psi
 367 SER   ( 141-)  B  omega poor
 400 ASP   ( 174-)  B  Poor phi/psi
 413 ARG   ( 187-)  B  omega poor
 436 TYR   ( 210-)  B  Poor phi/psi
 448 GLU   ( 222-)  B  Poor phi/psi
 chi-1/chi-2 correlation Z-score : -0.099

Note: chi-1/chi-2 angle correlation Z-score OK

The score expressing how well the chi-1/chi-2 angles of all residues correspond to the populated areas in the database is within expected ranges for well-refined structures.

chi-1/chi-2 correlation Z-score : -0.099

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.

 152 LYS   ( 151-)  A    0.36
 342 ILE   ( 116-)  B    0.36
 117 ILE   ( 116-)  A    0.37
  31 SER   (  30-)  A    0.39
 256 SER   (  30-)  B    0.39
/data/zata/whatcheck10/dssp/DSSP.EXE TAPEIN.DAT TAPEOUT.DAT > /dev/null
/data/zata/whatcheck10/dssp/DSSP.EXE TAPEIN.DAT TAPEOUT.DAT > /dev/null

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, especially if a regular DSSP secondary structure (H or S for helix or strand) is indicated!

Please be aware that the seleno-methionines have been validated as if they were methionines. This is not entirely correct, of course, but we do not believe that this influences the final outcome significantly.

 141 TYR   ( 140-)  A        0
 150 GLU   ( 149-)  A        0
 224 LEU   ( 223-)  A        0
 225 MSE   ( 224-)  A        0
 226 HIS   (   0-)  B        0
 227 MSE   (   1-)  B        0
 366 TYR   ( 140-)  B        0
 375 GLU   ( 149-)  B        0
 291 GLU   (  65-)  B  H     1
  66 GLU   (  65-)  A  H     2
 175 ASP   ( 174-)  A        2
 211 TYR   ( 210-)  A        2
 336 ALA   ( 110-)  B        2
 436 TYR   ( 210-)  B        2

Note: Backbone conformation Z-score OK

The backbone conformation analysis gives a score that is normal for well refined protein structures.

Backbone conformation Z-score : -0.275

Note: Omega angle restraint OK

The omega angles for trans-peptide bonds in a structure is expected to give a gaussian distribution with the average around +178 degrees, and a standard deviation around 5.5. In the current structure the standard deviation agrees with this expectation.

Omega average and std. deviation= 178.980 5.519

Note: PRO puckering amplitude OK

Puckering amplitudes for all PRO residues are within normal ranges.

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].

 411 PRO   ( 185-)  B    51.9 half-chair C-delta/C-gamma (54 degrees)

Note: Backbone oxygen evaluation OK

All residues for which the local backbone conformation could be found in the WHAT CHECK database have a normal backbone oxygen position.

Note: Peptide bond conformations

There are not enough (intact) amino acids in the file to analyse peptide bond conformations.

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.

 403 ASN   ( 177-)  B      ND2 <->  460 HOH   ( 681 )  B      O      0.90    1.80  INTRA
 107 ARG   ( 106-)  A      NH1 <->  459 HOH   ( 649 )  A      O      0.38    2.32  INTRA
 459 HOH   ( 603 )  A      O   <->  459 HOH   ( 724 )  A      O      0.33    1.87  INTRA
  17 ARG   (  16-)  A    A NH2 <->  459 HOH   ( 692 )  A      O      0.32    2.38  INTRA
 357 ARG   ( 131-)  B      NE  <->  460 HOH   ( 591 )  B      O      0.31    2.39  INTRA
 400 ASP   ( 174-)  B      OD2 <->  460 HOH   ( 688 )  B      O      0.30    2.10  INTRA
 459 HOH   ( 513 )  A      O   <->  459 HOH   ( 708 )  A      O      0.26    1.94  INTRA
 403 ASN   ( 177-)  B      CG  <->  460 HOH   ( 681 )  B      O      0.25    2.55  INTRA
 401 SER   ( 175-)  B      OG  <->  403 ASN   ( 177-)  B      ND2    0.25    2.45  INTRA BL
 260 LYS   (  34-)  B      NZ  <->  460 HOH   ( 560 )  B      O      0.24    2.46  INTRA
  35 LYS   (  34-)  A      NZ  <->  225 MSE   ( 224-)  A      O      0.23    2.47  INTRA BF
 305 LYS   (  79-)  B      NZ  <->  460 HOH   ( 686 )  B      O      0.22    2.48  INTRA
 305 LYS   (  79-)  B      NZ  <->  460 HOH   ( 663 )  B      O      0.21    2.49  INTRA
 144 HIS   ( 143-)  A      CB  <->  146 MSE   ( 145-)  A      CE     0.20    3.00  INTRA BF
 459 HOH   ( 706 )  A      O   <->  459 HOH   ( 724 )  A      O      0.19    2.01  INTRA
And so on for a total of 51 lines.

Note: Some notes regarding these bumps

The bumps have been binned in 5 categories ranging from 'should deal with' till 'must fix'. Additionally, the integrated sum of all bumps, the squared sum of all bumps, and these latter two values normalized by the number of contacts are listed too for comparison purposes between, for example, small and large proteins.

Total bump value: 7.499
Total bump value per residue: 0.113
Total number of bumps: 51
Total squared bump value: 2.149
Total number of bumps in the mildest bin: 43
Total number of bumps in the second bin: 7
Total number of bumps in the middle bin: 0
Total number of bumps in the fourth bin: 1
Total number of bumps in the worst bin: 0

Packing, accessibility and threading

Note: Inside/Outside residue distribution normal

The distribution of residue types over the inside and the outside of the protein is normal.

inside/outside RMS Z-score : 1.075

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

Note: Inside/Outside RMS Z-score plot

Chain identifier: B

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. Please be aware that the seleno-methionines have been validated as if they were methionines. This is not entirely correct, of course, but we do not believe that this influences the final outcome significantly

   2 MSE   (   1-)  A   -8.56
  71 MSE   (  70-)  A   -8.56
 100 MSE   (  99-)  A   -8.56
 146 MSE   ( 145-)  A   -8.56
 227 MSE   (   1-)  B   -8.56
 296 MSE   (  70-)  B   -8.56
 325 MSE   (  99-)  B   -8.56
 371 MSE   ( 145-)  B   -8.56
 366 TYR   ( 140-)  B   -8.13
 141 TYR   ( 140-)  A   -7.46
  97 ARG   (  96-)  A   -6.21
 322 ARG   (  96-)  B   -5.67
 108 TRP   ( 107-)  A   -5.45
 333 TRP   ( 107-)  B   -5.42
  56 PHE   (  55-)  A   -5.07
 281 PHE   (  55-)  B   -5.05
 209 TYR   ( 208-)  A   -5.04

Note: No series of residues with bad packing environment

There are no stretches of three or more residues each having a packing score worse than -4.0.

Note: Structural average packing environment OK

The structural average packing score is within normal ranges.

Average for range 1 - 450 : -0.660

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

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: B

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.

 365 GLY   ( 139-)  B   -2.97
 366 TYR   ( 140-)  B   -2.62

Warning: Abnormal packing Z-score for sequential residues

A stretch of at least four sequential residues with a 2nd generation packing Z-score below -1.75 was found. This could indicate that these residues are part of a strange loop or that the residues in this range are incomplete, but it might also be an indication of misthreading.

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

 363 TYR   ( 137-)  B     -  366 TYR   ( 140-)  B        -1.83

Note: Structural average packing Z-score OK

The structural average for the second generation packing score is within normal ranges.

All contacts : Average = -0.303 Z-score = -1.74
BB-BB contacts : Average = -0.205 Z-score = -1.44
BB-SC contacts : Average = -0.088 Z-score = -0.75
SC-BB contacts : Average = -0.259 Z-score = -1.57
SC-SC contacts : Average = -0.274 Z-score = -1.48

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

Note: Second generation quality Z-score plot

Chain identifier: B

Water, ion, and hydrogenbond related checks

Note: Water contacts OK

All water clusters make at least one contact with a non-water atom.

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.

 459 HOH   ( 702 )  A      O     11.05   18.22   13.34
 460 HOH   ( 649 )  B      O     -2.06   22.28  -20.45
 460 HOH   ( 660 )  B      O     -1.75   39.34   -0.70

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.

 459 HOH   ( 670 )  A      O
 460 HOH   ( 673 )  B      O
Number of ambiguities touching ambiguities: 0

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.

  13 ASN   (  12-)  A
 144 HIS   ( 143-)  A
 369 HIS   ( 143-)  B

Note: Histidine type assignments

For all complete HIS residues in the structure a tentative assignment to HIS-D (protonated on ND1), HIS-E (protonated on NE2), or HIS-H (protonated on both ND1 and NE2, positively charged) is made based on the hydrogen bond network. A second assignment is made based on which of the Engh and Huber [REF] histidine geometries fits best to the structure.

In the table below all normal histidine residues are listed. The assignment based on the geometry of the residue is listed first, together with the RMS Z-score for the fit to the Engh and Huber parameters. For all residues where the H-bond assignment is different, the assignment is listed in the last columns, together with its RMS Z-score to the Engh and Huber parameters.

As always, the RMS Z-scores should be close to 1.0 if the residues were restrained to the Engh and Huber parameters during refinement, and if enough (high resolution) data is available.

Please note that because the differences between the geometries of the different types are small it is possible that the geometric assignment given here does not correspond to the type used in refinement. This is especially true if the RMS Z-scores are much higher than 1.0.

If the two assignments differ, or the `geometry' RMS Z-score is high, it is advisable to verify the hydrogen bond assignment, check the HIS type used during the refinement and possibly adjust it.

   1 HIS   (   0-)  A     HIS-H   0.10 HIS-D   0.57
 144 HIS   ( 143-)  A     HIS-H   0.55 HIS-E   0.73
 221 HIS   ( 220-)  A     HIS-H   0.19 HIS-E   0.64
 226 HIS   (   0-)  B     HIS-H   0.08 HIS-E   0.61
 369 HIS   ( 143-)  B     HIS-H   0.45 HIS-E   0.78
 446 HIS   ( 220-)  B     HIS-H   0.18 HIS-E   0.54

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.

  15 THR   (  14-)  A      N
  17 ARG   (  16-)  A    A NE
  47 ILE   (  46-)  A      N
  67 ASN   (  66-)  A      ND2
 109 ARG   ( 108-)  A      NE
 123 ASP   ( 122-)  A      N
 136 ILE   ( 135-)  A      N
 153 ALA   ( 152-)  A      N
 187 VAL   ( 186-)  A      N
 236 ASP   (  10-)  B      N
 242 ARG   (  16-)  B      N
 268 SER   (  42-)  B      OG
 272 ILE   (  46-)  B      N
 307 PHE   (  81-)  B      N
 334 ARG   ( 108-)  B      NE
 345 GLU   ( 119-)  B      N
 361 ILE   ( 135-)  B      N
 378 ALA   ( 152-)  B      N
 412 VAL   ( 186-)  B      N
 436 TYR   ( 210-)  B      N
 450 MSE   ( 224-)  B      N

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.

  13 ASN   (  12-)  A      OD1
 144 HIS   ( 143-)  A      ND1
 150 GLU   ( 149-)  A      OE2
 221 HIS   ( 220-)  A      ND1
 238 ASN   (  12-)  B      OD1
 292 ASN   (  66-)  B      OD1
 369 HIS   ( 143-)  B      ND1
 375 GLU   ( 149-)  B      OE2
 439 GLU   ( 213-)  B      OE2
 446 HIS   ( 220-)  B      ND1

Note: Some notes regarding these donors and acceptors

The donors and acceptors have been counted, also as function of their accessibility. The buried donors and acceptors have been binned in five categories ranging from not forming any hydrogen bond till forming a poor till perfect hydrogen bond. Obviously, the buried donors and acceptors with no or just a poor hydrogen bond should be a topic of concern.

Note: Crystallisation conditions from REMARK 280

Crystallisation conditions as found in the PDB file header.

CRYSTAL
SOLVENT CONTENT, VS   (%): 48.84
MATTHEWS COEFFICIENT, VM (ANGSTROMS**3/DA): 2.40
CRYSTALLIZATION CONDITIONS: PEG 3350, SODIUM HEPES, CALCIUM
       CHLORIDE, GLYCEROL, PH 7.5, VAPOR DIFFUSION, HANGING DROP,
       TEMPERATURE 298.0K

Note: Overview of ions

When ions are located at special positions, their occupancy should be reduce by a factor that is the same as the multiplicity of that special position. This seems to have been done OK in this PDB file.

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 CHECK 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+.

 452 CA    ( 502-)  A  -    0.55   0.76 Scores about as good as NA *2
 453 CA    ( 503-)  A  -  -.-  -.-  Too few ligands (2)
 454 CA    ( 505-)  A  -  -.-  -.-  Too few ligands (1)
 455 CA    ( 506-)  A  -  -.-  -.-  Too few ligands (0)
 456 CA    ( 501-)  B  -    0.54   0.75 Scores about as good as NA *2
 457 CA    ( 504-)  B  -  -.-  -.-  Too few ligands (2)

Note: Water packing OK

All waters seem properly packed, or at least not packed like an ion. This method is experimental. See: swift.cmbi.ru.nl/teach/theory/

Note: Content of the PDB file as interpreted by WHAT CHECK

Content of the PDB file as interpreted by WHAT CHECK. WHAT CHECK has read your PDB file, and stored it internally in what is called 'the soup'. The content of this soup is listed here. An extensive explanation of all frequently used WHAT CHECK output formats can be found at swift.cmbi.ru.nl. Look under output formats. A course on reading this 'Molecules' table is part of the WHAT CHECK website.

     1     1 (    0)   225 (  224) A Protein             1kyt.besttls.pdb
     2   226 (    0)   450 (  224) B Protein             1kyt.besttls.pdb
     3   451 (  224)   451 (  224) A M O2 <-   225       1kyt.besttls.pdb
     4   452 (  502)   452 (  502) A CA                  1kyt.besttls.pdb
     5   453 (  503)   453 (  503) A CA                  1kyt.besttls.pdb
     6   454 (  505)   454 (  505) A CA                  1kyt.besttls.pdb
     7   455 (  506)   455 (  506) A CA                  1kyt.besttls.pdb
     8   456 (  501)   456 (  501) B CA                  1kyt.besttls.pdb
     9   457 (  504)   457 (  504) B CA                  1kyt.besttls.pdb
    10   458 (  224)   458 (  224) B M O2 <-   450       1kyt.besttls.pdb
    11   459 ( HOH )   459 ( HOH ) B water   (  212)     /data/tmp/pdb_red...
    12   460 ( HOH )   460 ( HOH ) A water   (  221)     /data/tmp/pdb_red...

Final summary

Note: Summary report

This is an overall summary of the quality of the structure as compared with current reliable structures. Numbers in brackets are the average and standard deviation observed for a large number of files determined with a similar resolution.

The second table mostly gives an impression of how well the model conforms to common refinement restraint values.


Structure Z-scores, positive is better than average:

  Resolution read from PDB file  :   1.700
  1st generation packing quality :  -0.400 (          (  0.0, 2.5))
  2nd generation packing quality :  -1.743 (          ( -0.3, 1.4))
  Ramachandran plot appearance   :  -0.193 (          (  0.0, 1.1))
  chi-1/chi-2 rotamer normality  :  -0.099
  Backbone conformation          :  -0.275
  Inside/Outside distribution    :   1.075
RMS Z-scores, should be close to 1.0:
Bond lengths : 0.622 (tight)
  Bond angles                    :   0.746
  Omega angle restraints         :   1.003
  Side chain planarity           :   0.934
  Improper dihedral distribution :   0.904
# 106 # Note: Introduction to refinement recommendations
First, be aware that the recommendations for crystallographers listed below
are produced by a computer program that was written by a guy who got his
PhD in NMR...
 
We have tried to convert the messages written in this report into a small
set of things you can do with your refinement software to get a better
structure. The things you should do first are listed first. And in some
cases you should first fix that problem, then refine a bit further, and
then run WHAT CHECK again before looking at other problems. If, for example,
WHAT CHECK has found a problem with the SCALE and CRYST cards, then you must
first fix that problem, refine the structure a bit further, and run WHAT
CHECK again because errors in the SCALE and or CRYST card can lead to many
problems elsewhere in the validation process.
 
It is also important to keep in mind that WHAT CHECK is software and that it
occasionally totally misunderstands what is the cause of a problem. But, if
WHAT CHECK lists a problem there normally is a problem albeit that it not
always is the actual problem that gets listed.
 
# 107 # Note: Matthews coefficient problem
WHAT CHECK detected a Matthews coefficient problem. Most times this is an
administrative problem caused by typing the wrong cell multiplicity number
on the CRYST card (or not typing it at all). Occasionally it is caused by
typing the wrong space group on the CRYST card. You better fix this problem,
but normally this problem does not cause WHAT CHECK to give any erroneous
error messages further down in the report.
 
# 108 # Note: Cell parameter anomaly
WHAT CHECK has compared the observed bond lengths with the Engh and Huber
parameters, and has done this as function of the direction of the bond
relative to the cell axes. From this analysis it was concluded that the
cell dimensions are probably not entirely perfect. The problem is not very
big, so you do not need to fix this before you start dealing with the other
suggestions, but you better fix this.
 
If this problem is caused by refining with another set of target values
than the Engh and Huber values, then I cannot help you because systematic
target value deviations can also cause this message to pop up.
 
# 109 # Note: Non-canonical amino acids detected
WHAT CHECK has detected a high percentage of non-canonical amino acids in
your PDB file. Be aware
that the validation of non-canonical residues is meaningless, unless you
provide a topology entry for the non-canonical residue(s). Validation of
residues near non-canonical residues is also troublesome. You must make
one (or more) new entries for the topology file (or ask Gert very friendly
to help you with this). The percentage of non-canonical residues is so
high that all but this warning can be neglected.
 
# 110 # Error: Bumps in your structure
Upon analysing the bumps in your structure, WHAT CHECK got very
worried. Often this means that you have forgotten to lower the
occupancy of overlapping ligands, residues, or water molecules. But,
whatever is the origin of this problem, you have to analyse it and
fix it.
 
# 111 # Error: Water bumps in your structure
WHAT CHECK had to delete several water molecules because they overlapped
so badly that things like hydrogen bond calculations were not possible.
Obviously WHAT CHECK is not as smart as you in dealing with this problem,
so you better do something about the overlapping waters.
Often this problem is caused by having alternate water locations that
did not get a reduced occupancy.
 
# 112 # Note: His, Asn, Gln side chain flips.
His, Asn, and Gln have an asymmetry in their side chain that is hard to
detect unless you have data at much better than 1.0 Angstrom resolution.
WHAT CHECK thinks that your structure contains His, Asn, or Gln residues that
will make better hydrogen bonds when flipped around their chi-2, chi-2, or
chi-3 side chain torsion angle, respectively. You better
check these Asn, His, and Gln residues, and if you use a refinement program
that includes molecular dynamics, then you must (after the
flips were made) refine a bit further before running WHAT CHECK again.
 
# 113 # Note: Free floating waters
Your structure contains a few water molecules that make no hydrogen bonds at
all. These waters must be removed, and you must then refine a bit further
before running WHAT CHECK again.
 
# 114 # Warning: Troublesome residues
The residues listed in the table below need to be inspected
 
This table is a very rough attempt to sort the residues according to how
badly they need your attention. The idea is that when you sit in  in front
of the graphics screen and study the residues with the electron density
present that you improve the structure most by dealing with the top residues
in this list first.
 
 225 MSE   ( 224-)  A      24.29
 403 ASN   ( 177-)  B      22.61
 146 MSE   ( 145-)  A      19.31
 450 MSE   ( 224-)  B      18.23
   2 MSE   (   1-)  A      17.13
  71 MSE   (  70-)  A      17.13
 100 MSE   (  99-)  A      17.13
 227 MSE   (   1-)  B      17.13
 296 MSE   (  70-)  B      17.13
 325 MSE   (  99-)  B      17.13
 371 MSE   ( 145-)  B      17.13
 322 ARG   (  96-)  B      16.69
 366 TYR   ( 140-)  B      16.26
 141 TYR   ( 140-)  A      14.91
  97 ARG   (  96-)  A      14.00
And so on for a total of    55 lines.
==============


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.