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. TLS seems not mentioned in the header of the PDB
file. But anyway, if WHAT IF complains about your B-factors, and you think
that they are OK, then check for TLS related B-factor problems first.
Obviously, the temperature at which the X-ray data was collected has some importance too:
Crystal temperature (K) :287.000
Note: B-factor plot
The average atomic B-factor per residue is plotted as function of the residue
Chain identifier: A
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
| 0.996642 0.000519 -0.000940| | 0.000519 0.998283 0.000165| | -0.000940 0.000165 0.998967|Proposed new scale matrix
| 0.023504 -0.000013 0.006128| | -0.000013 0.024022 -0.000004| | 0.000013 -0.000002 0.014171|With corresponding cell
A = 42.557 B = 41.628 C = 72.946 Alpha= 89.997 Beta= 104.668 Gamma= 89.940
The CRYST1 cell dimensions
A = 42.700 B = 41.700 C = 73.000 Alpha= 90.000 Beta= 104.600 Gamma= 90.000
(Under-)estimated Z-score: 4.668
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.
203 THR ( 208-) A N CA C 99.85 -4.1
202 VAL ( 207-) A 4.75 149 LYS ( 154-) A 4.31 123 TYR ( 128-) A 4.14 203 THR ( 208-) A 4.05
Tau angle RMS Z-score : 1.633
Error: Connections to aromatic rings out of plane
The atoms listed in the table below are connected to a planar aromatic group
in the sidechain of a protein residue but were found to deviate from the
least squares plane.
For all atoms that are connected to an aromatic side chain in a protein residue the distance of the atom to the least squares plane through the aromatic system was determined. This value was divided by the standard deviation from a distribution of similar values from a database of small molecule structures.
103 HIS ( 107-) A CB 5.03 Since there is no DNA and no protein with hydrogens, no uncalibrated planarity check was performed. Ramachandran Z-score : -1.804
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.
79 PRO ( 83-) A -2.8 76 LYS ( 80-) A -2.5 56 LEU ( 60-) A -2.2 88 GLN ( 92-) A -2.1 158 VAL ( 163-) A -2.1 18 ILE ( 22-) 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.
25 SER ( 29-) A PRO omega poor 59 GLY ( 63-) A Poor phi/psi 71 ASP ( 75-) A Poor phi/psi 88 GLN ( 92-) A Poor phi/psi 107 LYS ( 111-) A Poor phi/psi 112 ALA ( 116-) A Poor phi/psi 173 ASN ( 178-) A Poor phi/psi 196 PRO ( 201-) A PRO omega poor 247 LYS ( 252-) A Poor phi/psi chi-1/chi-2 correlation Z-score : -2.076
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!
3 TYR ( 7-) A 0 15 ASP ( 19-) A 0 16 PHE ( 20-) A 0 20 LYS ( 24-) A 0 23 ARG ( 27-) A 0 24 GLN ( 28-) A 0 25 SER ( 29-) A 0 46 SER ( 50-) A 0 48 ASP ( 52-) A 0 50 ALA ( 54-) A 0 58 ASN ( 62-) A 0 60 HIS ( 64-) A 0 68 ASP ( 72-) A 0 69 SER ( 73-) A 0 71 ASP ( 75-) A 0 72 LYS ( 76-) A 0 73 ALA ( 77-) A 0 75 LEU ( 79-) A 0 76 LYS ( 80-) A 0 79 PRO ( 83-) A 0 80 LEU ( 84-) A 0 81 ASP ( 85-) A 0 87 ILE ( 91-) A 0 88 GLN ( 92-) A 0 91 PHE ( 95-) A 0And so on for a total of 117 lines.
Standard deviation of omega values : 1.947
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]
79 PRO ( 83-) A 0.45 HIGH
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.
11 HIS ( 15-) A ND1 <-> 14 LYS ( 18-) A NZ 0.21 2.79 INTRA 18 ILE ( 22-) A O <-> 21 GLY ( 25-) A N 0.16 2.54 INTRA 95 SER ( 99-) A N <-> 96 LEU ( 100-) A N 0.09 2.51 INTRA BL 36 TYR ( 40-) A CD1 <-> 255 PHE ( 260-) A C 0.09 3.11 INTRA 191 GLY ( 196-) A N <-> 202 VAL ( 207-) A O 0.08 2.62 INTRA BL 114 LEU ( 118-) A N <-> 141 ILE ( 146-) A O 0.05 2.65 INTRA BL 70 GLN ( 74-) A O <-> 72 LYS ( 76-) A N 0.04 2.66 INTRA 24 GLN ( 28-) A O <-> 249 ARG ( 254-) A NH2 0.04 2.66 INTRA BL 188 THR ( 193-) A N <-> 252 LYS ( 257-) A O 0.03 2.67 INTRA BL 99 GLN ( 103-) A NE2 <-> 238 ASP ( 243-) A OD1 0.03 2.67 INTRA BL 224 LEU ( 229-) A O <-> 236 MET ( 241-) A N 0.03 2.67 INTRA BL 97 ASP ( 101-) A OD1 <-> 222 ARG ( 227-) A NH2 0.03 2.67 INTRA 47 TYR ( 51-) A OH <-> 118 HIS ( 122-) A NE2 0.02 2.68 INTRA 121 THR ( 125-) A C <-> 123 TYR ( 128-) A N 0.02 2.88 INTRA BL 146 GLY ( 151-) A N <-> 213 VAL ( 218-) A O 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.
6 HIS ( 10-) A -5.94 96 LEU ( 100-) A -5.09
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: 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.
257 HOH ( 324 ) A O Metal-coordinating Histidine residue 90 fixed to 1 Metal-coordinating Histidine residue 92 fixed to 1
173 ASN ( 178-) A 248 ASN ( 253-) 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.
27 VAL ( 31-) A N 32 HIS ( 36-) A N 48 ASP ( 52-) A N 49 GLN ( 53-) A N 70 GLN ( 74-) A N 96 LEU ( 100-) A N 195 THR ( 200-) A N 199 LEU ( 204-) A N 214 SER ( 219-) A OG 239 ASN ( 244-) A ND2 240 TRP ( 245-) A N 248 ASN ( 253-) A N 255 PHE ( 260-) A N Only metal coordination for 92 HIS ( 96-) A NE2
157 ASP ( 162-) A H-bonding suggests Asn; but Alt-Rotamer 234 GLU ( 239-) A H-bonding suggests Gln
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 : -0.169 2nd generation packing quality : 0.738 Ramachandran plot appearance : -1.804 chi-1/chi-2 rotamer normality : -2.076 Backbone conformation : -0.822
Bond lengths : 0.544 (tight) Bond angles : 0.790 Omega angle restraints : 0.354 (tight) Side chain planarity : 0.676 Improper dihedral distribution : 1.106 B-factor distribution : 0.438 Inside/Outside distribution : 0.954
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 : 2.20
Structure Z-scores, positive is better than average:
1st generation packing quality : 0.4 2nd generation packing quality : 0.7 Ramachandran plot appearance : -0.5 chi-1/chi-2 rotamer normality : -0.7 Backbone conformation : -0.7
Bond lengths : 0.544 (tight) Bond angles : 0.790 Omega angle restraints : 0.354 (tight) Side chain planarity : 0.676 Improper dihedral distribution : 1.106 B-factor distribution : 0.438 Inside/Outside distribution : 0.954 ==============
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.