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.
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: 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: 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: Tyrosine convention problem
The tyrosine residues listed in the table below have their chi-2 not between
-90.0 and 90.0
19 TYR ( 23-) A
43 PHE ( 47-) A 72 PHE ( 76-) A 77 PHE ( 81-) A 102 PHE ( 106-) A 103 PHE ( 107-) A 114 PHE ( 118-) A 119 PHE ( 123-) A
65 GLU ( 69-) A
RMS Z-score for bond lengths: 0.541
RMS-deviation in bond distances: 0.013
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
| 0.996400 0.000158 0.000207| | 0.000158 0.995925 0.000281| | 0.000207 0.000281 0.995876|Proposed new scale matrix
| 0.031201 -0.000008 0.010072| | -0.000003 0.017835 -0.000005| | -0.000006 -0.000007 0.026571|With corresponding cell
A = 32.048 B = 56.071 C = 39.545 Alpha= 89.975 Beta= 107.879 Gamma= 89.982
The CRYST1 cell dimensions
A = 32.164 B = 56.301 C = 39.712 Alpha= 90.000 Beta= 107.890 Gamma= 90.000
(Under-)estimated Z-score: 6.668
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.
82 HIS ( 86-) A CG ND1 CE1 109.65 4.0
65 GLU ( 69-) A
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.
54 THR ( 58-) A -2.9 86 PHE ( 90-) A -2.5 50 PHE ( 54-) A -2.2 51 MET ( 55-) A -2.2
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.
40 ASN ( 44-) A Poor phi/psi 49 HIS ( 53-) A Poor phi/psi 50 PHE ( 54-) A Poor phi/psi 86 PHE ( 90-) A Poor phi/psi 114 PHE ( 118-) A Poor phi/psi 141 ASP ( 145-) A Poor phi/psi 148 LYS ( 152-) A omega poor 151 ASN ( 155-) A omega poor chi-1/chi-2 correlation Z-score : -0.429
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.
33 SER ( 37-) A 0.36
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!
10 MET ( 14-) A 0 18 PHE ( 22-) A 0 19 TYR ( 23-) A 0 22 CYS ( 26-) A 0 40 ASN ( 44-) A 0 43 PHE ( 47-) A 0 44 HIS ( 48-) A 0 45 ARG ( 49-) A 0 48 LYS ( 52-) A 0 49 HIS ( 53-) A 0 50 PHE ( 54-) A 0 51 MET ( 55-) A 0 54 THR ( 58-) A 0 58 SER ( 62-) A 0 68 TRP ( 72-) A 0 70 ASN ( 74-) A 0 73 GLU ( 77-) A 0 79 HIS ( 83-) A 0 80 LEU ( 84-) A 0 82 HIS ( 86-) A 0 83 SER ( 87-) A 0 86 PHE ( 90-) A 0 90 MET ( 94-) A 0 91 ALA ( 95-) A 0 92 ASN ( 96-) A 0And so on for a total of 71 lines.
For each of the residues in the structure, a search was performed to find 5-residue stretches in the WHAT IF database with superposable C-alpha coordinates, and some restraining on the neighbouring backbone oxygens.
In the following table the RMS distance between the backbone oxygen positions of these matching structures in the database and the position of the backbone oxygen atom in the current residue is given. If this number is larger than 1.5 a significant number of structures in the database show an alternative position for the backbone oxygen. If the number is larger than 2.0 most matching backbone fragments in the database have the peptide plane flipped. A manual check needs to be performed to assess whether the experimental data can support that alternative as well. The number in the last column is the number of database hits (maximum 80) used in the calculation. It is "normal" that some glycine residues show up in this list, but they are still worth checking!
69 GLY ( 73-) A 1.64 23
1 PRO ( 5-) A 50.9 half-chair C-delta/C-gamma (54 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.
1 PRO ( 5-) A N <-> 156 HOH ( 339 ) A O 0.37 2.33 INTRA 60 ASP ( 64-) A OD2 <-> 62 THR ( 66-) A OG1 0.30 2.10 INTRA 111 TRP ( 115-) A NE1 <-> 156 HOH ( 233 ) A O 0.21 2.49 INTRA 60 ASP ( 64-) A OD1 <-> 156 HOH ( 276 ) A O 0.10 2.30 INTRA BF 48 LYS ( 52-) A NZ <-> 156 HOH ( 227 ) A O 0.08 2.62 INTRA 40 ASN ( 44-) A ND2 <-> 156 HOH ( 278 ) A O 0.08 2.62 INTRA 40 ASN ( 44-) A ND2 <-> 148 LYS ( 152-) A NZ 0.07 2.78 INTRA 20 LYS ( 24-) A O <-> 23 LYS ( 27-) A NZ 0.06 2.64 INTRA 56 ASP ( 60-) A OD2 <-> 61 GLY ( 65-) A N 0.05 2.65 INTRA 145 GLU ( 149-) A CG <-> 156 HOH ( 345 ) A O 0.04 2.76 INTRA BF 40 ASN ( 44-) A N <-> 149 ILE ( 153-) A O 0.04 2.66 INTRA 70 ASN ( 74-) A OD1 <-> 156 HOH ( 330 ) A O 0.01 2.39 INTRA 32 HIS ( 36-) A NE2 <-> 65 GLU ( 69-) A O 0.01 2.69 INTRA 81 ASN ( 85-) A ND2 <-> 156 HOH ( 314 ) A O 0.01 2.69 INTRA 155 ASN ( 159-) A ND2 <-> 156 HOH ( 341 ) A O 0.01 2.69 INTRA BF
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.
93 CYS ( 97-) A -5.39 136 ARG ( 140-) A -5.21
Chain identifier: A
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: 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.
40 ASN ( 44-) A 155 ASN ( 159-) 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.
107 VAL ( 111-) A N 111 TRP ( 115-) A N 152 ILE ( 156-) A N
The score listed is the valency score. This number should be close to (preferably a bit above) 1.0 for the suggested ion to be a likely alternative for the water molecule. Ions listed in brackets are good alternate choices. *1 indicates that the suggested ion-type has been observed elsewhere in the PDB file too. *2 indicates that the suggested ion-type has been observed in the REMARK 280 cards of the PDB file. Ion-B and ION-B indicate that the B-factor of this water is high, or very high, respectively. H2O-B indicates that the B-factors of atoms that surround this water/ion are suspicious. See: swift.cmbi.ru.nl/teach/theory/ for a detailed explanation.
156 HOH ( 182 ) A O 0.85 K 4 156 HOH ( 217 ) A O 1.02 K 4 156 HOH ( 243 ) A O 1.03 K 4
75 GLU ( 79-) A H-bonding suggests Gln; but Alt-Rotamer
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 : -0.201 2nd generation packing quality : -2.226 Ramachandran plot appearance : -0.319 chi-1/chi-2 rotamer normality : -0.429 Backbone conformation : -0.438
Bond lengths : 0.541 (tight) Bond angles : 0.685 Omega angle restraints : 1.120 Side chain planarity : 0.606 (tight) Improper dihedral distribution : 0.701 B-factor distribution : 0.411 Inside/Outside distribution : 1.020
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 : 0.2 2nd generation packing quality : -1.6 Ramachandran plot appearance : -0.2 chi-1/chi-2 rotamer normality : -0.2 Backbone conformation : -0.7
Bond lengths : 0.541 (tight) Bond angles : 0.685 Omega angle restraints : 1.120 Side chain planarity : 0.606 (tight) Improper dihedral distribution : 0.701 B-factor distribution : 0.411 Inside/Outside distribution : 1.020 ==============
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.