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.
260 DMS ( 264-) A - 261 DMS ( 265-) A - 262 E36 ( 262-) A -
Plausible side chain atoms were detected with (near) zero occupancy
When crystallographers do not see an atom they either leave it out completely, or give it an occupancy of zero or a very high B-factor. WHAT IF neglects these atoms. In this case some atoms were found with zero occupancy, but with coordinates that place them at a plausible position. Although WHAT IF knows how to deal with missing side chain atoms, validation will go more reliable if all atoms are presnt. So, please consider manually setting the occupancy of the listed atoms at 1.0.
6 LYS ( 9-) A - CD 6 LYS ( 9-) A - CE 6 LYS ( 9-) A - NZ 42 LYS ( 45-) A - CE 42 LYS ( 45-) A - NZ 77 LYS ( 80-) A - CD 77 LYS ( 80-) A - CE 77 LYS ( 80-) A - NZ 129 LYS ( 133-) A - CD 129 LYS ( 133-) A - CE 129 LYS ( 133-) A - NZ 155 LYS ( 159-) A - CD 155 LYS ( 159-) A - CE 155 LYS ( 159-) A - NZ 168 LYS ( 172-) A - CD 168 LYS ( 172-) A - CE 168 LYS ( 172-) A - NZ 249 ASN ( 253-) A - CB 249 ASN ( 253-) A - CG 249 ASN ( 253-) A - OD1 249 ASN ( 253-) A - ND2 253 LYS ( 257-) A - CD 253 LYS ( 257-) A - CE 253 LYS ( 257-) A - NZ 257 LYS ( 261-) A - CB 257 LYS ( 261-) A - CG 257 LYS ( 261-) A - CD 257 LYS ( 261-) A - CE 257 LYS ( 261-) A - NZ
Plausible backbone atoms were detected with (near) zero occupancy
When crystallographers do not see an atom they either leave it out completely, or give it an occupancy of zero or a very high B-factor. WHAT IF neglects these atoms. However, if a backbone atom is present in the PDB file, and its position seems 'logical' (i.e. normal bond lengths with all atoms it should be bound to, and those atoms exist normally) WHAT IF will set the occupancy to 1.0 if it believes that the full presence of this atom will be beneficial to the rest of the validation process. If you get weird errors at, or near, these atoms, please check by hand what is going on, and repair things intelligently before running this validation again.
249 ASN ( 253-) A - CA
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
Coordinate problems, unexpected atoms, B-factor and occupancy checks
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.
11 GLU ( 14-) A 0.50 100 GLN ( 103-) A 0.50 257 LYS ( 261-) A 0.50
Obviously, the temperature at which the X-ray data was collected has some importance too:
Number of TLS groups mentione in PDB file header: 0
Crystal temperature (K) :100.000
Note: B-factor plot
The average atomic B-factor per residue is plotted as function of the residue
Chain identifier: A
Nomenclature related problems
Warning: Phenylalanine convention problem
The phenylalanine residues listed in the table below have their chi-2 not
between -90.0 and 90.0.
63 PHE ( 66-) A
161 ASP ( 165-) A 171 ASP ( 175-) A
210 GLU ( 214-) A
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.
5 GLY ( 8-) A N CA 1.52 4.5 20 ALA ( 23-) A N CA 1.54 4.3 24 ARG ( 27-) A CG CD 1.65 4.5 84 THR ( 87-) A CA CB 1.61 4.1 123 LYS ( 127-) A N -C 1.47 6.9 212 ILE ( 216-) A CB CG2 1.68 4.3
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
| 0.992717 -0.000428 0.001214| | -0.000428 0.989745 -0.000481| | 0.001214 -0.000481 0.993165|Proposed new scale matrix
| 0.023992 0.000013 0.006144| | 0.000011 0.024639 0.000012| | -0.000018 0.000007 0.014504|With corresponding cell
A = 41.667 B = 40.587 C = 71.148 Alpha= 90.042 Beta= 104.293 Gamma= 90.049
The CRYST1 cell dimensions
A = 41.972 B = 41.007 C = 71.681 Alpha= 90.000 Beta= 104.430 Gamma= 90.000
(Under-)estimated Z-score: 18.071
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.
61 HIS ( 64-) A CG ND1 CE1 109.99 4.4 116 HIS ( 119-) A CG ND1 CE1 109.65 4.0 119 HIS ( 122-) A CG ND1 CE1 109.94 4.3 123 LYS ( 127-) A -O -C N 111.93 -6.9 123 LYS ( 127-) A -CA -C N 124.32 4.1 173 THR ( 177-) A N CA CB 103.33 -4.2
161 ASP ( 165-) A 171 ASP ( 175-) A 210 GLU ( 214-) A
31 ASP ( 34-) A 5.83
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.
19 ILE ( 22-) A -2.3 36 LYS ( 39-) A -2.3 159 VAL ( 163-) A -2.2 27 PRO ( 30-) A -2.2 80 PRO ( 83-) A -2.1 172 PHE ( 176-) A -2.1 147 GLY ( 151-) A -2.1 97 LEU ( 100-) A -2.0 47 SER ( 50-) A -2.0
Residues with `forbidden' phi-psi combinations are listed, as well as residues with unusual omega angles (deviating by more than 3 sigma from the normal value). Please note that it is normal if about 5 percent of the residues is listed here as having unusual phi-psi combinations.
26 SER ( 29-) A PRO omega poor 31 ASP ( 34-) A omega poor 80 PRO ( 83-) A omega poor 83 GLY ( 86-) A omega poor 89 GLN ( 92-) A omega poor 108 LYS ( 111-) A Poor phi/psi 125 GLY ( 129-) A Poor phi/psi 172 PHE ( 176-) A omega poor 174 ASN ( 178-) A Poor phi/psi 187 TYR ( 191-) A omega poor 193 SER ( 197-) A omega poor 197 PRO ( 201-) A PRO omega poor 199 LEU ( 203-) A Poor phi/psi 203 VAL ( 207-) A omega poor 211 PRO ( 215-) A omega poor 239 ASP ( 243-) A Poor phi/psi 248 LYS ( 252-) A Poor phi/psi 255 SER ( 259-) A omega poor chi-1/chi-2 correlation Z-score : -2.166
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!
4 TYR ( 7-) A 0 7 HIS ( 10-) A 0 12 HIS ( 15-) A 0 13 TRP ( 16-) A 0 16 ASP ( 19-) A 0 17 PHE ( 20-) A 0 21 LYS ( 24-) A 0 23 GLU ( 26-) A 0 24 ARG ( 27-) A 0 25 GLN ( 28-) A 0 26 SER ( 29-) A 0 47 SER ( 50-) A 0 59 ASN ( 62-) A 0 61 HIS ( 64-) A 0 62 ALA ( 65-) A 0 69 ASP ( 72-) A 0 70 SER ( 73-) A 0 72 ASP ( 75-) A 0 73 LYS ( 76-) A 0 74 ALA ( 77-) A 0 77 LYS ( 80-) A 0 80 PRO ( 83-) A 0 82 ASP ( 85-) A 0 88 ILE ( 91-) A 0 89 GLN ( 92-) A 0And so on for a total of 119 lines.
Standard deviation of omega values : 7.000
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]
197 PRO ( 201-) A 0.47 HIGH
39 PRO ( 42-) A -119.2 half-chair C-delta/C-gamma (-126 degrees)
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.
173 THR ( 177-) A CG2 <-> 174 ASN ( 178-) A ND2 0.38 2.72 INTRA 73 LYS ( 76-) A O <-> 263 HOH ( 314 ) A O 0.27 2.13 INTRA 24 ARG ( 27-) A CG <-> 201 GLU ( 205-) A CD 0.20 3.00 INTRA BL 107 ASP ( 110-) A OD2 <-> 263 HOH ( 335 ) A O 0.18 2.22 INTRA 109 LYS ( 112-) A NZ <-> 263 HOH ( 368 ) A O 0.16 2.54 INTRA 55 ARG ( 58-) A NH2 <-> 66 GLU ( 69-) A OE2 0.15 2.55 INTRA 89 GLN ( 92-) A NE2 <-> 261 DMS ( 265-) A O 0.10 2.60 INTRA 12 HIS ( 15-) A ND1 <-> 15 LYS ( 18-) A NZ 0.09 2.91 INTRA 231 GLY ( 235-) A O <-> 263 HOH ( 357 ) A O 0.08 2.32 INTRA 100 GLN ( 103-) A OE1 <-> 263 HOH ( 343 ) A O 0.06 2.34 INTRA 48 TYR ( 51-) A OH <-> 119 HIS ( 122-) A NE2 0.05 2.65 INTRA BL 230 GLU ( 234-) A N <-> 263 HOH ( 398 ) A O 0.04 2.66 INTRA 4 TYR ( 7-) A N <-> 263 HOH ( 320 ) A O 0.04 2.66 INTRA 72 ASP ( 75-) A OD1 <-> 86 ARG ( 89-) A NH2 0.04 2.66 INTRA 239 ASP ( 243-) A O <-> 260 DMS ( 264-) A C1 0.03 2.77 INTRA 114 GLU ( 117-) A OE2 <-> 116 HIS ( 119-) A NE2 0.03 2.67 INTRA BL 248 LYS ( 252-) A N <-> 263 HOH ( 403 ) A O 0.03 2.67 INTRA 7 HIS ( 10-) A A C <-> 12 HIS ( 15-) A CD2 0.03 3.17 INTRA 248 LYS ( 252-) A NZ <-> 263 HOH ( 386 ) A O 0.02 2.68 INTRA 24 ARG ( 27-) A CG <-> 201 GLU ( 205-) A CB 0.02 3.18 INTRA BL 2 TRP ( 5-) A CZ2 <-> 61 HIS ( 64-) A CE1 0.01 3.19 INTRA 243 PRO ( 247-) A O <-> 245 GLN ( 249-) A NE2 0.01 2.69 INTRA BL
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.
7 HIS ( 10-) A -6.21 97 LEU ( 100-) A -5.27 6 LYS ( 9-) A -5.19 33 HIS ( 36-) A -5.18 132 GLN ( 136-) A -5.09
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.
251 GLN ( 255-) A -2.64
Chain identifier: A
Water, ion, and hydrogenbond related checks
Warning: Water molecules need moving
The water molecules listed in the table below were found to be significantly
closer to a symmetry related non-water molecule than to the ones given in the
coordinate file. For optimal viewing convenience revised coordinates for
these water molecules should be given.
The number in brackets is the identifier of the water molecule in the input file. Suggested coordinates are also given in the table. Please note that alternative conformations for protein residues are not taken into account for this calculation. If you are using WHAT IF / WHAT-CHECK interactively, then the moved waters can be found in PDB format in the file: MOVEDH2O.pdb.
263 HOH ( 318 ) A O 8.21 -5.81 -10.48 263 HOH ( 362 ) A O -12.27 12.86 29.10 263 HOH ( 370 ) A O 12.79 -3.57 13.44 263 HOH ( 374 ) A O -15.40 11.46 34.68 263 HOH ( 381 ) A O -15.07 19.97 17.72 263 HOH ( 385 ) A O -12.21 16.20 8.02 263 HOH ( 391 ) A O -12.77 13.18 31.70 263 HOH ( 392 ) A O 10.12 -2.36 14.02 263 HOH ( 395 ) A O -4.85 20.21 21.24 263 HOH ( 406 ) A O 14.05 -4.07 10.39 263 HOH ( 411 ) A O -7.04 1.61 35.81
263 HOH ( 375 ) A O 263 HOH ( 405 ) A O Marked this atom as acceptor 262 E36 ( 262-) A CL26 Metal-coordinating Histidine residue 91 fixed to 1 Metal-coordinating Histidine residue 93 fixed to 1 Metal-coordinating Histidine residue 116 fixed to 1
1 HIS ( 4-) A 50 GLN ( 53-) A 133 GLN ( 137-) A
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.
28 VAL ( 31-) A N 71 GLN ( 74-) A N 89 GLN ( 92-) A NE2 97 LEU ( 100-) A N 200 LEU ( 204-) A N 226 ASN ( 230-) A ND2 240 ASN ( 244-) A ND2 241 TRP ( 245-) A N 256 PHE ( 260-) A N Only metal coordination for 91 HIS ( 94-) A NE2 Only metal coordination for 93 HIS ( 96-) A NE2 Only metal coordination for 116 HIS ( 119-) A ND1
103 GLU ( 106-) A H-bonding suggests Gln 161 ASP ( 165-) A H-bonding suggests Asn
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 : -0.242 2nd generation packing quality : 0.545 Ramachandran plot appearance : -1.563 chi-1/chi-2 rotamer normality : -2.166 Backbone conformation : -1.089
Bond lengths : 1.178 Bond angles : 1.016 Omega angle restraints : 1.273 (loose) Side chain planarity : 1.496 Improper dihedral distribution : 1.288 B-factor distribution : 0.865 Inside/Outside distribution : 0.947
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 : 1.80
Structure Z-scores, positive is better than average:
1st generation packing quality : 0.1 2nd generation packing quality : -0.2 Ramachandran plot appearance : -1.5 chi-1/chi-2 rotamer normality : -1.8 Backbone conformation : -1.4
Bond lengths : 1.178 Bond angles : 1.016 Omega angle restraints : 1.273 (loose) Side chain planarity : 1.496 Improper dihedral distribution : 1.288 B-factor distribution : 0.865 Inside/Outside distribution : 0.947 ==============
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.