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.
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.
1 HIS ( 3-) A - CB 1 HIS ( 3-) A - CG 1 HIS ( 3-) A - ND1 1 HIS ( 3-) A - CD2 1 HIS ( 3-) A - CE1 1 HIS ( 3-) A - NE2 2 HIS ( 4-) A - ND1 2 HIS ( 4-) A - CD2 2 HIS ( 4-) A - CE1 2 HIS ( 4-) A - NE2 258 LYS ( 261-) A - CB 258 LYS ( 261-) A - CG 258 LYS ( 261-) A - CD 258 LYS ( 261-) A - CE 258 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.
1 HIS ( 3-) A - N 1 HIS ( 3-) A - CA 1 HIS ( 3-) A - C 1 HIS ( 3-) A - O 2 HIS ( 4-) A - C
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: Missing atoms
The atoms listed in the table below are missing from the entry. If many atoms
are missing, the other checks can become less sensitive. Be aware that it
often happens that groups at the termini of DNA or RNA are really missing,
so that the absence of these atoms normally is neither an error nor the
result of poor electron density. Some of the atoms listed here might also be
listed by other checks, most noticeably by the options in the previous
section that list missing atoms in several categories. The plausible atoms
with zero occupancy are not listed here, as they already got assigned a
non-zero occupancy, and thus are no longer 'missing'.
258 LYS ( 261-) A O
Obviously, the temperature at which the X-ray data was collected has some importance too:
Crystal temperature (K) :293.000
Warning: More than 2 percent of buried atoms has low B-factor
For protein structures determined at room temperature, no more than
about 1 percent of the B factors of buried atoms is below 5.0.
Percentage of buried atoms with B less than 5 : 2.16
Note: B-factor plot
The average atomic B-factor per residue is plotted as function of the residue
Chain identifier: A
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.
92 HIS ( 94-) A CG CD2 1.41 5.3 117 HIS ( 119-) A CG CD2 1.42 5.8
34 HIS ( 36-) A -C N CA 114.34 -4.1 92 HIS ( 94-) A CG ND1 CE1 110.92 5.3 92 HIS ( 94-) A CB CG CD2 134.72 4.3 94 HIS ( 96-) A CG ND1 CE1 110.70 5.1 94 HIS ( 96-) A CB CG CD2 134.90 4.5 105 HIS ( 107-) A CG ND1 CE1 109.61 4.0 117 HIS ( 119-) A CG ND1 CE1 110.65 5.0 117 HIS ( 119-) A CB CG CD2 136.38 5.6 205 THR ( 208-) A N CA C 99.13 -4.3
Improper dihedrals are a measure of the chirality/planarity of the structure at a specific atom. Values around -35 or +35 are expected for chiral atoms, and values around 0 for planar atoms. Planar side chains are left out of the calculations, these are better handled by the planarity checks.
Three numbers are given for each atom in the table. The first is the Z-score for the improper dihedral. The second number is the measured improper dihedral. The third number is the expected value for this atom type. A final column contains an extra warning if the chirality for an atom is opposite to the expected value.
34 HIS ( 36-) A CA -7.2 20.86 34.11 The average deviation= 1.280
204 VAL ( 207-) A 5.28 205 THR ( 208-) A 4.35 224 ARG ( 227-) A 4.34
Tau angle RMS Z-score : 1.606
Warning: Torsion angle evaluation shows unusual residues
The residues listed in the table below contain bad or abnormal
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.
81 PRO ( 83-) A -2.7 11 PRO ( 13-) A -2.6 98 LEU ( 100-) A -2.6 28 PRO ( 30-) A -2.3 58 LEU ( 60-) A -2.3 164 ILE ( 167-) A -2.2 233 GLU ( 236-) A -2.2 239 VAL ( 242-) A -2.1 51 GLN ( 53-) A -2.1 160 VAL ( 163-) A -2.1 48 SER ( 50-) A -2.1 148 GLY ( 151-) A -2.0 90 GLN ( 92-) 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.
27 SER ( 29-) A PRO omega poor 73 ASP ( 75-) A Poor phi/psi 74 LYS ( 76-) A Poor phi/psi 108 ASP ( 110-) A Poor phi/psi 109 LYS ( 111-) A Poor phi/psi 175 ASN ( 178-) A Poor phi/psi 198 PRO ( 201-) A PRO omega poor 200 LEU ( 203-) A Poor phi/psi 240 ASP ( 243-) A Poor phi/psi 249 LYS ( 252-) A Poor phi/psi chi-1/chi-2 correlation Z-score : -2.748
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 TRP ( 5-) A 0 5 TYR ( 7-) A 0 8 HIS ( 10-) A 0 17 ASP ( 19-) A 0 18 PHE ( 20-) A 0 20 ILE ( 22-) A 0 22 LYS ( 24-) A 0 25 ARG ( 27-) A 0 27 SER ( 29-) A 0 34 HIS ( 36-) A 0 35 THR ( 37-) A 0 36 ALA ( 38-) A 0 48 SER ( 50-) A 0 50 ASP ( 52-) A 0 51 GLN ( 53-) A 0 56 ARG ( 58-) A 0 60 ASN ( 62-) A 0 62 HIS ( 64-) A 0 63 SER ( 65-) A 0 64 PHE ( 66-) A 0 70 ASP ( 72-) A 0 71 SER ( 73-) A 0 74 LYS ( 76-) A 0 75 ALA ( 77-) A 0 78 LYS ( 80-) A 0And so on for a total of 115 lines.
Standard deviation of omega values : 1.795
Warning: Backbone oxygen evaluation
The residues listed in the table below have an unusual backbone oxygen
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!
232 GLY ( 235-) A 1.96 69
198 PRO ( 201-) A 102.5 envelop C-beta (108 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.
62 HIS ( 64-) A NE2 <-> 260 HOH ( 366 ) A O 0.37 2.33 INTRA 13 HIS ( 15-) A ND1 <-> 16 LYS ( 18-) A NZ 0.21 2.79 INTRA BL 244 PRO ( 247-) A O <-> 246 GLN ( 249-) A NE2 0.16 2.54 INTRA BL 65 ASN ( 67-) A ND2 <-> 260 HOH ( 371 ) A O 0.13 2.57 INTRA 6 GLY ( 8-) A O <-> 10 GLY ( 12-) A N 0.12 2.58 INTRA BL 35 THR ( 37-) A N <-> 36 ALA ( 38-) A N 0.12 2.48 INTRA B3 34 HIS ( 36-) A ND1 <-> 260 HOH ( 376 ) A O 0.11 2.59 INTRA 258 LYS ( 261-) A C <-> 260 HOH ( 407 ) A O 0.10 2.70 INTRA 158 VAL ( 161-) A CG1 <-> 222 LYS ( 225-) A CD 0.10 3.10 INTRA 51 GLN ( 53-) A N <-> 260 HOH ( 380 ) A O 0.08 2.62 INTRA 165 LYS ( 168-) A NZ <-> 225 LYS ( 228-) A O 0.07 2.63 INTRA 98 LEU ( 100-) A CD1 <-> 100 GLY ( 102-) A N 0.07 3.03 INTRA 252 GLN ( 255-) A NE2 <-> 254 LYS ( 257-) A NZ 0.04 2.81 INTRA 94 HIS ( 96-) A CD2 <-> 117 HIS ( 119-) A CE1 0.04 3.16 INTRA BL 37 LYS ( 39-) A O <-> 255 ALA ( 258-) A N 0.04 2.66 INTRA 216 SER ( 219-) A O <-> 219 GLN ( 222-) A N 0.03 2.67 INTRA BL 55 LEU ( 57-) A N <-> 67 GLU ( 69-) A O 0.03 2.67 INTRA BL 56 ARG ( 58-) A NE <-> 67 GLU ( 69-) A CD 0.03 3.07 INTRA 76 VAL ( 78-) A CG2 <-> 78 LYS ( 80-) A NZ 0.03 3.07 INTRA 2 HIS ( 4-) A ND1 <-> 3 TRP ( 5-) A O 0.03 2.57 INTRA 30 ASP ( 32-) A OD1 <-> 109 LYS ( 111-) A N 0.02 2.68 INTRA BL 148 GLY ( 151-) A N <-> 215 VAL ( 218-) A O 0.02 2.68 INTRA BL 249 LYS ( 252-) A CB <-> 250 ASN ( 253-) A N 0.02 2.68 INTRA B3 169 LYS ( 172-) A NZ <-> 260 HOH ( 354 ) A O 0.01 2.69 INTRA 28 PRO ( 30-) A O <-> 246 GLN ( 249-) A N 0.01 2.69 INTRA BL 216 SER ( 219-) A C <-> 218 GLU ( 221-) A N 0.01 2.89 INTRA BL 5 TYR ( 7-) A N <-> 6 GLY ( 8-) A N 0.01 2.59 INTRA BL 133 GLN ( 136-) A N <-> 134 GLN ( 137-) A N 0.01 2.59 INTRA B3
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.
8 HIS ( 10-) A -5.86 2 HIS ( 4-) A -5.24 98 LEU ( 100-) A -5.23 133 GLN ( 136-) A -5.13 34 HIS ( 36-) A -5.10
Chain identifier: A
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.
247 PRO ( 250-) A - 250 ASN ( 253-) A -1.59
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.
260 HOH ( 407 ) A O Metal-coordinating Histidine residue 92 fixed to 1 Metal-coordinating Histidine residue 94 fixed to 1 Metal-coordinating Histidine residue 117 fixed to 1
8 HIS ( 10-) 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.
29 VAL ( 31-) A N 43 LYS ( 45-) A N 74 LYS ( 76-) A N 98 LEU ( 100-) A N 127 ASP ( 130-) A N 185 SER ( 188-) A N 197 THR ( 200-) A N 201 LEU ( 204-) A N 241 ASN ( 244-) A ND2 242 TRP ( 245-) A N 245 ALA ( 248-) A N 257 PHE ( 260-) A N Only metal coordination for 94 HIS ( 96-) A NE2
12 GLU ( 14-) 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 : -0.446 2nd generation packing quality : 0.245 Ramachandran plot appearance : -1.960 chi-1/chi-2 rotamer normality : -2.748 Backbone conformation : -1.134
Bond lengths : 0.522 (tight) Bond angles : 0.814 Omega angle restraints : 0.326 (tight) Side chain planarity : 0.452 (tight) Improper dihedral distribution : 1.063 B-factor distribution : 1.152 Inside/Outside distribution : 0.961
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 : 0.2 2nd generation packing quality : 0.4 Ramachandran plot appearance : -0.6 chi-1/chi-2 rotamer normality : -1.3 Backbone conformation : -1.0
Bond lengths : 0.522 (tight) Bond angles : 0.814 Omega angle restraints : 0.326 (tight) Side chain planarity : 0.452 (tight) Improper dihedral distribution : 1.063 B-factor distribution : 1.152 Inside/Outside distribution : 0.961 ==============
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.