Full wwPDB EM Validation Report i ○ Dec 16, 2020 – 09:26 PM JST PDB ID : 6L3H EMDB ID : EMD-0822 Title : Cryo-EM structure of dimeric quinol dependent Nitric Oxide Reductase (qNOR) from the pathogen Neisseria meninigitidis Authors : Jamali, M.M.A.; Gopalasingam, C.C.; Johnson, R.M.; Tosha, T.; Muench, S.P.; Muramoto, K.; Antonyuk, S.V.; Shiro, Y.; Hasnain, S.S. Deposited on : 2019-10-11 Resolution : 3.06 Å(reported) Based on initial model : 6L1X This is a Full wwPDB EM Validation Report for a publicly released PDB entry. We welcome your comments at [email protected]A user guide is available at https://www.wwpdb.org/validation/2017/EMValidationReportHelp with specific help available everywhere you see the i ○ symbol. The following versions of software and data (see references i ○) were used in the production of this report: EMDB validation analysis : 0.0.0.dev61 Mogul : 1.8.5 (274361), CSD as541be (2020) MolProbity : 4.02b-467 buster-report : 1.1.7 (2018) Percentile statistics : 20191225.v01 (using entries in the PDB archive December 25th 2019) Ideal geometry (proteins) : Engh & Huber (2001) Ideal geometry (DNA, RNA) : Parkinson et al. (1996) Validation Pipeline (wwPDB-VP) : 2.15.1
Transcript
Full wwPDB EM Validation Report i○
Dec 16, 2020 – 09:26 PM JST
PDB ID : 6L3HEMDB ID : EMD-0822
Title : Cryo-EM structure of dimeric quinol dependent Nitric Oxide Reductase(qNOR) from the pathogen Neisseria meninigitidis
Page 2 Full wwPDB EM Validation Report EMD-0822, 6L3H
1 Overall quality at a glance i○
The following experimental techniques were used to determine the structure:ELECTRON MICROSCOPY
The reported resolution of this entry is 3.06 Å.
Percentile scores (ranging between 0-100) for global validation metrics of the entry are shown inthe following graphic. The table shows the number of entries on which the scores are based.
The table below summarises the geometric issues observed across the polymeric chains and their fitto the map. The red, orange, yellow and green segments of the bar indicate the fraction of residuesthat contain outliers for >=3, 2, 1 and 0 types of geometric quality criteria respectively. A greysegment represents the fraction of residues that are not modelled. The numeric value for eachfraction is indicated below the corresponding segment, with a dot representing fractions <=5%The upper red bar (where present) indicates the fraction of residues that have poor fit to the EMmap (all-atom inclusion < 40%). The numeric value is given above the bar.
Page 3 Full wwPDB EM Validation Report EMD-0822, 6L3H
2 Entry composition i○
There are 4 unique types of molecules in this entry. The entry contains 11966 atoms, of which 0are hydrogens and 0 are deuteriums.
In the tables below, the AltConf column contains the number of residues with at least one atomin alternate conformation and the Trace column contains the number of residues modelled with atmost 2 atoms.
• Molecule 1 is a protein called Nitric-oxide reductase.
Mol Chain Residues Atoms AltConf Trace
1 A 741 Total C N O S5895 3907 943 1020 25 0 0
1 B 741 Total C N O S5895 3907 943 1020 25 0 0
• Molecule 2 is PROTOPORPHYRIN IX CONTAINING FE (three-letter code: HEM) (for-mula: C34H32FeN4O4).
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• Molecule 3 is FE (III) ION (three-letter code: FE) (formula: Fe).
Mol Chain Residues Atoms AltConf
3 B 1 Total Fe1 1 0
3 A 1 Total Fe1 1 0
• Molecule 4 is CALCIUM ION (three-letter code: CA) (formula: Ca).
Mol Chain Residues Atoms AltConf
4 B 1 Total Ca1 1 0
4 A 1 Total Ca1 1 0
Page 5 Full wwPDB EM Validation Report EMD-0822, 6L3H
3 Residue-property plots i○
These plots are drawn for all protein, RNA, DNA and oligosaccharide chains in the entry. Thefirst graphic for a chain summarises the proportions of the various outlier classes displayed in thesecond graphic. The second graphic shows the sequence view annotated by issues in geometry andatom inclusion in map density. Residues are color-coded according to the number of geometricquality criteria for which they contain at least one outlier: green = 0, yellow = 1, orange = 2and red = 3 or more. A red diamond above a residue indicates a poor fit to the EM map forthis residue (all-atom inclusion < 40%). Stretches of 2 or more consecutive residues without anyoutlier are shown as a green connector. Residues present in the sample, but not in the model, areshown in grey.
Page 6 Full wwPDB EM Validation Report EMD-0822, 6L3H
4 Experimental information i○
Property Value SourceEM reconstruction method SINGLE PARTICLE DepositorImposed symmetry POINT, C2 DepositorNumber of particles used 233556 DepositorResolution determination method FSC 0.143 CUT-OFF DepositorCTF correction method PHASE FLIPPING AND AMPLITUDE
Page 7 Full wwPDB EM Validation Report EMD-0822, 6L3H
5 Model quality i○
5.1 Standard geometry i○
Bond lengths and bond angles in the following residue types are not validated in this section:HEM, CA, FE
The Z score for a bond length (or angle) is the number of standard deviations the observed valueis removed from the expected value. A bond length (or angle) with |Z| > 5 is considered anoutlier worth inspection. RMSZ is the root-mean-square of all Z scores of the bond lengths (orangles).
Mol Chain Bond lengths Bond anglesRMSZ #|Z| >5 RMSZ #|Z| >5
1 A 0.38 0/6084 0.46 0/82941 B 0.38 0/6084 0.46 0/8294All All 0.38 0/12168 0.46 0/16588
Chiral center outliers are detected by calculating the chiral volume of a chiral center and verifying ifthe center is modelled as a planar moiety or with the opposite hand.A planarity outlier is detectedby checking planarity of atoms in a peptide group, atoms in a mainchain group or atoms of asidechain that are expected to be planar.
Mol Chain #Chirality outliers #Planarity outliers1 A 0 11 B 0 1All All 0 2
There are no bond length outliers.
There are no bond angle outliers.
There are no chirality outliers.
All (2) planarity outliers are listed below:
Mol Chain Res Type Group1 A 471 PHE Peptide1 B 471 PHE Peptide
5.2 Too-close contacts i○
In the following table, the Non-H and H(model) columns list the number of non-hydrogen atomsand hydrogen atoms in the chain respectively. The H(added) column lists the number of hydrogenatoms added and optimized by MolProbity. The Clashes column lists the number of clashes withinthe asymmetric unit, whereas Symm-Clashes lists symmetry-related clashes.
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Mol Chain Non-H H(model) H(added) Clashes Symm-Clashes1 A 5895 0 5805 43 01 B 5895 0 5805 43 02 A 86 0 60 4 02 B 86 0 60 4 03 A 1 0 0 0 03 B 1 0 0 0 04 A 1 0 0 0 04 B 1 0 0 0 0All All 11966 0 11730 94 0
The all-atom clashscore is defined as the number of clashes found per 1000 atoms (includinghydrogen atoms). The all-atom clashscore for this structure is 4.
All (94) close contacts within the same asymmetric unit are listed below, sorted by their clashmagnitude.
In the following table, the Percentiles column shows the percent Ramachandran outliers of thechain as a percentile score with respect to all PDB entries followed by that with respect to all EMentries.
The Analysed column shows the number of residues for which the backbone conformation wasanalysed, and the total number of residues.
All All 1474/1502 (98%) 1374 (93%) 100 (7%) 0 100 100
There are no Ramachandran outliers to report.
5.3.2 Protein sidechains i○
In the following table, the Percentiles column shows the percent sidechain outliers of the chainas a percentile score with respect to all PDB entries followed by that with respect to all EMentries.
The Analysed column shows the number of residues for which the sidechain conformation wasanalysed, and the total number of residues.
Mol Chain Analysed Rotameric Outliers Percentiles
1 A 607/615 (99%) 605 (100%) 2 (0%) 92 96
1 B 607/615 (99%) 605 (100%) 2 (0%) 92 96
All All 1214/1230 (99%) 1210 (100%) 4 (0%) 92 96
All (4) residues with a non-rotameric sidechain are listed below:
Mol Chain Res Type1 A 655 ARG1 A 722 TRP1 B 655 ARG1 B 722 TRP
Sometimes sidechains can be flipped to improve hydrogen bonding and reduce clashes. All (1) suchsidechains are listed below:
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5.4 Non-standard residues in protein, DNA, RNA chains i○
There are no non-standard protein/DNA/RNA residues in this entry.
5.5 Carbohydrates i○
There are no monosaccharides in this entry.
5.6 Ligand geometry i○
Of 8 ligands modelled in this entry, 4 are monoatomic - leaving 4 for Mogul analysis.
In the following table, the Counts columns list the number of bonds (or angles) for which Mogulstatistics could be retrieved, the number of bonds (or angles) that are observed in the model andthe number of bonds (or angles) that are defined in the Chemical Component Dictionary. TheLink column lists molecule types, if any, to which the group is linked. The Z score for a bondlength (or angle) is the number of standard deviations the observed value is removed from theexpected value. A bond length (or angle) with |Z| > 2 is considered an outlier worth inspection.RMSZ is the root-mean-square of all Z scores of the bond lengths (or angles).
Mol Type Chain Res Link Bond lengths Bond anglesCounts RMSZ #|Z| > 2 Counts RMSZ #|Z| > 2
2 HEM B 801 1,4 27,50,50 1.91 4 (14%) 17,82,82 1.56 4 (23%)2 HEM A 802 1,4 27,50,50 2.01 4 (14%) 17,82,82 1.43 3 (17%)2 HEM B 802 1,4 27,50,50 2.01 4 (14%) 17,82,82 1.43 3 (17%)2 HEM A 801 1,4 27,50,50 1.90 4 (14%) 17,82,82 1.55 4 (23%)
In the following table, the Chirals column lists the number of chiral outliers, the number of chiralcenters analysed, the number of these observed in the model and the number defined in theChemical Component Dictionary. Similar counts are reported in the Torsion and Rings columns.’-’ means no outliers of that kind were identified.
Mol Type Chain Res Link Chirals Torsions Rings2 HEM B 801 1,4 - 0/6/54/54 -2 HEM A 802 1,4 - 0/6/54/54 -2 HEM B 802 1,4 - 0/6/54/54 -2 HEM A 801 1,4 - 0/6/54/54 -
All (16) bond length outliers are listed below:
Mol Chain Res Type Atoms Z Observed(Å) Ideal(Å)2 B 802 HEM C3B-C2B -5.50 1.32 1.402 A 802 HEM C3B-C2B -5.50 1.32 1.40
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Continued from previous page...Mol Chain Res Type Atoms Z Observed(Å) Ideal(Å)2 B 802 HEM C3C-C2C -5.45 1.32 1.402 A 802 HEM C3C-C2C -5.38 1.32 1.402 B 801 HEM C3C-C2C -5.35 1.33 1.402 A 801 HEM C3C-C2C -5.26 1.33 1.402 B 801 HEM C3B-C2B -4.62 1.34 1.402 A 801 HEM C3B-C2B -4.62 1.34 1.402 B 801 HEM C3B-CAB 3.40 1.54 1.472 A 802 HEM C3B-CAB 3.39 1.54 1.472 A 801 HEM C3B-CAB 3.39 1.54 1.472 B 802 HEM C3B-CAB 3.35 1.54 1.472 A 802 HEM C3C-CAC 3.28 1.54 1.472 B 802 HEM C3C-CAC 3.27 1.54 1.472 A 801 HEM C3C-CAC 3.26 1.54 1.472 B 801 HEM C3C-CAC 3.25 1.54 1.47
All (14) bond angle outliers are listed below:
Mol Chain Res Type Atoms Z Observed(o) Ideal(o)2 A 801 HEM CAA-CBA-CGA -2.77 108.03 112.672 B 801 HEM CAA-CBA-CGA -2.76 108.04 112.672 A 802 HEM CBD-CAD-C3D -2.54 107.80 112.482 B 802 HEM CBD-CAD-C3D -2.52 107.83 112.482 B 801 HEM CMA-C3A-C4A -2.45 124.70 128.462 A 801 HEM CMA-C3A-C4A -2.42 124.74 128.462 B 801 HEM CMB-C2B-C3B 2.40 129.17 124.682 A 801 HEM CMB-C2B-C3B 2.38 129.13 124.682 B 801 HEM C1D-C2D-C3D 2.17 108.51 107.002 A 801 HEM C1D-C2D-C3D 2.10 108.46 107.002 A 802 HEM CBA-CAA-C2A -2.10 108.62 112.492 B 802 HEM CBA-CAA-C2A -2.09 108.63 112.492 B 802 HEM CMA-C3A-C4A -2.09 125.25 128.462 A 802 HEM CMA-C3A-C4A -2.08 125.27 128.46
There are no chirality outliers.
There are no torsion outliers.
There are no ring outliers.
4 monomers are involved in 8 short contacts:
Mol Chain Res Type Clashes Symm-Clashes2 B 801 HEM 2 02 A 802 HEM 2 0
Continued on next page...
Page 14 Full wwPDB EM Validation Report EMD-0822, 6L3H
Continued from previous page...Mol Chain Res Type Clashes Symm-Clashes2 B 802 HEM 2 02 A 801 HEM 2 0
The following is a two-dimensional graphical depiction of Mogul quality analysis of bond lengths,bond angles, torsion angles, and ring geometry for all instances of the Ligand of Interest. Inaddition, ligands with molecular weight > 250 and outliers as shown on the validation Tables willalso be included. For torsion angles, if less then 5% of the Mogul distribution of torsion angles iswithin 10 degrees of the torsion angle in question, then that torsion angle is considered an outlier.Any bond that is central to one or more torsion angles identified as an outlier by Mogul will behighlighted in the graph. For rings, the root-mean-square deviation (RMSD) between the ringin question and similar rings identified by Mogul is calculated over all ring torsion angles. If theaverage RMSD is greater than 60 degrees and the minimal RMSD between the ring in question andany Mogul-identified rings is also greater than 60 degrees, then that ring is considered an outlier.The outliers are highlighted in purple. The color gray indicates Mogul did not find sufficientequivalents in the CSD to analyse the geometry.
Ligand HEM B 801
Bond lengths Bond angles
Torsions Rings
Page 15 Full wwPDB EM Validation Report EMD-0822, 6L3H
Ligand HEM A 802
Bond lengths Bond angles
Torsions Rings
Page 16 Full wwPDB EM Validation Report EMD-0822, 6L3H
Ligand HEM B 802
Bond lengths Bond angles
Torsions Rings
Page 17 Full wwPDB EM Validation Report EMD-0822, 6L3H
Page 18 Full wwPDB EM Validation Report EMD-0822, 6L3H
6 Map visualisation i○
This section contains visualisations of the EMDB entry EMD-0822. These allow visual inspectionof the internal detail of the map and identification of artifacts.
No raw map or half-maps were deposited for this entry and therefore no images, graphs, etc.pertaining to the raw map can be shown.
6.1 Orthogonal projections i○
6.1.1 Primary map
X Y Z
The images above show the map projected in three orthogonal directions.
Page 19 Full wwPDB EM Validation Report EMD-0822, 6L3H
The images above show central slices of the map in three orthogonal directions.
6.3 Largest variance slices i○
6.3.1 Primary map
X Index: 94 Y Index: 86 Z Index: 102
The images above show the largest variance slices of the map in three orthogonal directions.
6.4 Orthogonal surface views i○
6.4.1 Primary map
X Y Z
The images above show the 3D surface view of the map at the recommended contour level 0.0268.These images, in conjunction with the slice images, may facilitate assessment of whether an ap-propriate contour level has been provided.
Page 21 Full wwPDB EM Validation Report EMD-0822, 6L3H
7 Map analysis i○
This section contains the results of statistical analysis of the map.
7.1 Map-value distribution i○
The map-value distribution is plotted in 128 intervals along the x-axis. The y-axis is logarithmic.A spike in this graph at zero usually indicates that the volume has been masked.
Page 22 Full wwPDB EM Validation Report EMD-0822, 6L3H
7.2 Volume estimate i○
The volume at the recommended contour level is 77 nm3; this corresponds to an approximate massof 69 kDa.
The volume estimate graph shows how the enclosed volume varies with the contour level. Therecommended contour level is shown as a vertical line and the intersection between the line andthe curve gives the volume of the enclosed surface at the given level.
Page 24 Full wwPDB EM Validation Report EMD-0822, 6L3H
8 Fourier-Shell correlation i○
Fourier-Shell Correlation (FSC) is the most commonly used method to estimate the resolution ofsingle-particle and subtomogram-averaged maps. The shape of the curve depends on the imposedsymmetry, mask and whether or not the two 3D reconstructions used were processed from acommon reference. The reported resolution is shown as a black line. A curve is displayed for thehalf-bit criterion in addition to lines showing the 0.143 gold standard cut-off and 0.5 cut-off.
8.1 FSC i○
*Reported resolution corresponds to spatial frequency of 0.327 Å−1
Page 26 Full wwPDB EM Validation Report EMD-0822, 6L3H
9 Map-model fit i○
This section contains information regarding the fit between EMDB map EMD-0822 and PDBmodel 6L3H. Per-residue inclusion information can be found in section 3 on page 5.
9.1 Map-model overlay i○
X Y Z
The images above show the 3D surface view of the map at the recommended contour level 0.0268at 50% transparency in yellow overlaid with a ribbon representation of the model coloured in blue.These images allow for the visual assessment of the quality of fit between the atomic model andthe map.
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