Temperature value: Difference between revisions
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In [[X-ray crystallography|crystallography]], '''uncertainty''' in the positions of atoms increases with '''disorder''' in the protein crystal. Disorder may have two components, static and dynamic. [[Resolution]] represents the average uncertainty for all atoms. In contrast, the ''temperature value'' (also called ''temperature factor'' or ''B factor'') quantitates the uncertainty for each atom. At typical resolutions for protein crystals (where ''occupancy'' cannot be distinguished from ''B value''), a high temperature factor reflects a low [[Electron density map|empirical electron density]] for the atom, and ''vice versa''. Generally, a temperature value of less than 30 Å<sup>2</sup> signifies confidence in its position, while a temperature value of greater than 60 Å<sup>2</sup> signifies disorder<ref name="proxychem">[http://www.proxychem.com/macromolecular_crystallography.html Macromolecular Crystallography] at ProXyChem.com</ref>. | In [[X-ray crystallography|crystallography]], '''uncertainty''' in the positions of atoms increases with '''disorder''' in the protein crystal. Disorder may have two components, static and dynamic. [[Resolution]] represents the average uncertainty for all atoms. In contrast, the ''temperature value'' (also called ''temperature factor'' or ''B factor'') quantitates the uncertainty for each atom. At typical resolutions for protein crystals (where ''occupancy'' cannot be distinguished from ''B value''), a high temperature factor reflects a low [[Electron density map|empirical electron density]] for the atom, and ''vice versa''. This is illustrated in [http://www.bioinformatics.org/molvis/edm/edm2.htm Electron Density Maps]. Generally, a temperature value of less than 30 Å<sup>2</sup> signifies confidence in its position, while a temperature value of greater than 60 Å<sup>2</sup> signifies disorder<ref name="proxychem">[http://www.proxychem.com/macromolecular_crystallography.html Macromolecular Crystallography] at ProXyChem.com</ref>. | ||
The temperature value is recorded in the [[atomic coordinate file]]. In the [[PDB_file#PDB_Data_Format|PDB file format]], it is the last numeric value (columns 61-66) in each ATOM and HETATM record. [[#Coloring by Temperature|Coloring by temperature]] is a common way to visualize the uncertainty for each atom. | The temperature value is recorded in the [[atomic coordinate file]]. In the [[PDB_file#PDB_Data_Format|PDB file format]], it is the last numeric value (columns 61-66) in each ATOM and HETATM record. [[#Coloring by Temperature|Coloring by temperature]] is a common way to visualize the uncertainty for each atom. | ||
[[PDB files]] for models determined by [[cryo-EM]] often specify values in the temperature/B factor field. However, a 2017 analysis concluded that "the treatment of the atomic displacement (B) factors was meaningless in almost all analyzed cryo-EM models"<ref>PMID: 28867613</ref>. | |||
==Definition== | ==Definition== | ||
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Some regions of the molecule may have higher average disorder, and others lower average disorder. Typically, the ends of chains have higher average disorder, and hence their positions are less certain than are residues in the core of a tightly packed domain, where disorder is less. | Some regions of the molecule may have higher average disorder, and others lower average disorder. Typically, the ends of chains have higher average disorder, and hence their positions are less certain than are residues in the core of a tightly packed domain, where disorder is less. | ||
==B-factors and coordinate error== | |||
Disorder in a crystal is reflected in lower resolution of the diffraction data, as described above. Disorder in the coordinates based on that data can be modeled by introducing B-factors for each atom. They model atomic positions being displaced (by an average distance that is related to the B-factor) from an average position (given by the coordinates). There is no direct relationship between B-factor and coordinate error. If we obtain perfect information about the electron density at atomic resolution, we can infer the average position of an atom even if it has a high B-factor (this amounts to finding the center of a fuzzy dilute cloud as opposed to a compact dense cloud). However, there is an indirect relationship: Higher disorder in a crystal results in lower resolution of the diffraction data, resulting in high coordinate errors. At the same time, the average B-factor of the model will be high, reflecting the disorder of the crystal, so overall coordinate error and overall B-factor will be correlated. For individual atoms or regions of the structure with higher disorder, there is a higher chance for systematic errors in building the model, so this correlation of high B-factors and high coordinate errors extends to separate regions of the protein as well (see [http://www.ccp4.ac.uk/newsletters/newsletter33/murshudov.html], Figure 4 for an example of correlating B-factors with coordinate errors, given specific values for resolution, completeness and free R-factor, which influence coordinate errors as well.) | |||
==Coloring by Temperature== | ==Coloring by Temperature== | ||
<applet load=' | <applet load='' size='250' frame='true' align='right' | ||
scene='Temperature_value/1uwb/2' /> | scene='Temperature_value/1uwb/2' /> | ||
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Comparison of the temperature colors of two separate models is risky, since some methodological issues can bias the temperature factors. There are [[Temperature color schemes|two temperature coloring schemes: absolute and relative]]. In general, high [[Resolution|resolution]] models tend to have fewer red atoms than do models with modest resolution. For example, a <scene name='Temperature_value/1kzk/1'>small HIV protease at 1.1 Angstroms resolution</scene> ([[1kzk]]) has <scene name='Temperature_value/1kzk/2'>few red atoms</scene>, while the <scene name='Temperature_value/1uwb/2'>larger, less compact HIV reverse transcriptase at 3.2 Angstroms resolution</scene> ([[1uwb]]) has <scene name='Temperature_value/1uwb/1'>many red atoms</scene>. (These scenes are colored by ''relative'' temperature. Coloring by absolute temperature looks very similar in these particular cases. See [[Temperature color schemes#Examples|other examples]].) | Comparison of the temperature colors of two separate models is risky, since some methodological issues can bias the temperature factors. There are [[Temperature color schemes|two temperature coloring schemes: absolute and relative]]. In general, high [[Resolution|resolution]] models tend to have fewer red atoms than do models with modest resolution. For example, a <scene name='Temperature_value/1kzk/1'>small HIV protease at 1.1 Angstroms resolution</scene> ([[1kzk]]) has <scene name='Temperature_value/1kzk/2'>few red atoms</scene>, while the <scene name='Temperature_value/1uwb/2'>larger, less compact HIV reverse transcriptase at 3.2 Angstroms resolution</scene> ([[1uwb]]) has <scene name='Temperature_value/1uwb/1'>many red atoms</scene>. (These scenes are colored by ''relative'' temperature. Coloring by absolute temperature looks very similar in these particular cases. See [[Temperature color schemes#Examples|other examples]].) | ||
See also [[Temperature value vs. resolution]]. | |||
In [[1uwb]], many of the surface sidechains are missing (due to disorder). Missing sidechains are labeled '''S-''' in ''FirstGlance in Jmol'' (linked under the molecule on all [[PDB code]]-titled pages in Proteopedia). | In [[1uwb]], many of the surface sidechains are missing (due to disorder). Missing sidechains are labeled '''S-''' in ''FirstGlance in Jmol'' (linked under the molecule on all [[PDB code]]-titled pages in Proteopedia). | ||
Most [[molecular modeling and visualization software]] packages have an option to color by temperature, including [[FirstGlance in Jmol]], which is linked on every page in Proteopedia that is titled with a [[PDB code]]. In [[FirstGlance in Jmol]] (version | Most [[molecular modeling and visualization software]] packages have an option to color by temperature, including [[FirstGlance in Jmol]], which is linked on every page in Proteopedia that is titled with a [[PDB code]]. In [[FirstGlance in Jmol]] (version 2), this option will be found under the ''Views'' tab, as ''Local Uncertainty''. | ||
==Missing Residues and Atoms== | |||
==Missing Residues== | ===Missing Residues=== | ||
Often the very ends of chains, or surface loops, may be so disordered as to prevent assigning any atomic positions at all, leading to '''missing residues'''. [[FirstGlance in Jmol]] (linked beneath the molecule on every [[PDB code]]-titled page in Proteopedia) has | Often the very ends of chains, or surface loops, may be so disordered as to prevent assigning any atomic positions at all, leading to '''missing residues'''. That is, these residues were present in the crystallized protein, but have no coordinates in the atomic model because their electron densities were too indistinct. | ||
[[FirstGlance in Jmol]] (linked beneath the molecule on every [[PDB code]]-titled page in Proteopedia) lists missing residues and marks their positions with eye-catching "empty baskets". See snapshots at [[Missing residues and incomplete sidechains]]. It is much easier to overlook missing residues, especially at the ends of chains, in other molecular viewers such as PyMOL, ChimeraX, MolStar, and iCn3D (see [[Missing residues and incomplete sidechains|comparision snapshots]]). | |||
The sequence listing for a PDB code offered by [http://pdbe.org/ PDB-Europe] makes it easy to see missing residues: they are highlighted with a gray background. FirstGlance in Jmol has links to these listings under ''Sequences''. | |||
Alternatively, at the [[PDB]], the ''Sequence'' tab provides a graphic representation of the sequence that indicates gaps in two ways. First, the thin black line underneath the sequence is broken; second, touching a residue above breaks in the line reports "no identifier from ATOM record (no structural data available)". However, it is easy to overlook breaks in the line. | |||
===Incomplete Sidechains=== | |||
In addition to entire residues missing from the atomic model, side chains atoms may be missing (due to disorder), even when the main chain atoms are present. FirstGlance in Jmol puts the label '''S-''' on every residue with missing side chain atoms making this obvious. PyMOL, ChimeraX, MolStar, and iCn3D leave their users unaware of these missing atoms (see [[Missing_residues_and_incomplete_sidechains#Incomplete_Sidechains|comparision snapshots]]), which are especially important when looking at charge distribution or calculating electrostatic potential maps (see [[Missing_residues_and_incomplete_sidechains#Do_incomplete_sidechains_matter.3F|examples]]). Solutions are to use the AlphaFold model (where no atoms are missing) or to add back the missing atoms, use the free [https://www.protein-science.com/ Protein Repair & Analysis Server]. | |||
In the [[PDB file format]], missing residues are listed in REMARK 465, while missing atoms are listed in REMARK 470. | |||
==Data Format== | ==Data Format== | ||
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==See Also== | ==See Also== | ||
*[[Temperature color schemes]] | |||
*[[Temperature value vs. resolution]] | |||
*[[Resolution]] | *[[Resolution]] | ||
*[[Quality assessment for molecular models]] | *[[Quality assessment for molecular models]] | ||
*[[NMR Ensembles of Models]] | *[[NMR Ensembles of Models]] | ||
*[[Anisotropic refinement]] | *[[Anisotropic refinement]] | ||
*[[Intrinsically Disordered Protein]] | |||
==References Cited== | |||
<references /> | |||
==Content Donors== | ==Content Donors== | ||
The section [[#B-factors and coordinate error]] was written by '''[[User:Karsten Theis]]''' and moved to this article by Eric Martz. | |||
Large portions of the initial version of this page were adapted from the [http://proteinexplorer.org/igloss.htm Glossary of ProteinExplorer.Org] by the principal author, [[User:Eric Martz|Eric Martz]]. | Large portions of the initial version of this page were adapted from the [http://proteinexplorer.org/igloss.htm Glossary of ProteinExplorer.Org] by the principal author, [[User:Eric Martz|Eric Martz]]. | ||
[[Category: BioMolViz]] | |||
[[Category: Structural Model Skepticism]] | |||
