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Twilight: A tool for ligand density validation 

Visualizing Ligand Molecules in Twilight Electron Density

Christian X.  Weichenbergera*,  Edwin  Pozharskib* and  Bernhard  Ruppc

aCenter for Biomedicine, European Academy of Bozen/Bolzano (EURAC), Viale Druso 1, Bozen/Bolzano, I-39100, Italy,  bUniversity of Maryland, Baltimore, MD, USA, and  ck.k. Hofkristallamt, 991 Audrey Place, Vista, CA, 92084, USA

Correspondence email:  Christian.Weichenberger@eurac.edu; epozh001@umaryland.edu


We present a software script for facilitating analysis and visual inspection of ligand molecules in context of the electron density maps calculated from experimental data associated with protein structures determined by X-ray crystallography.

Original Abstract (2013)

Three-dimensional models of protein structures determined by X-ray crystallography are based on the interpretation of experimentally derived electron density maps. The real-space correlation coefficient (RSCC) provides an easily comprehensible, objective measure for residue-based fit of atom coordinates with electron density. Among protein structure models, protein-ligand complexes are of special interest, given their contribution to understanding the molecular underpinnings of biological activity and to drug design. For consumers of such models it is not trivial to determine the degree to which ligand structure modelling is biased by subjective electron density interpretation. We present a standalone script, Twilight, for analysis, visualization, and annotation of a pre-filtered set of 2,815 protein/ligand complexes deposited with the PDB as of January 15, 2012, with ligand RSCC values that are below a threshold of 0.6. The script runs on various platforms and is available for download at https://www.ruppweb.org/twilight/.

Download this Acta Crystallographica F paper and its companion in Acta Crystallographica D describing the actual findings from the IUCr web site (preferred) or request a link by email. Please read also the editorial Expectation bias and information content Z. Dauter, M. S. Weiss, H. Einspahr and E. N. Baker, Acta Crystallogr. D69, 141 (2013).


The TWILIGHT GUI tool itself has been discontinued - but you can download sortable Excel tables of the small molecule LIGAND and PEPTIDE collections.


 Twilight (I)


Twilight reloaded: The peptide experience

Techniques, tools and best practices for ligand electron-density analysis and results from their application to deposited crystal structures. E. Pozharski, C. X. Weichenberger and B. Rupp, Acta Crystallogr D69, 150-167 (2013)

Visualizing Ligand Molecules in Twilight Electron Density. C. X. Weichenberger, E. Pozharski and B. Rupp, Acta Crystallogr. F69(2), 195-200(2013)

Twilight reloaded: The Peptide Experience. C. X. Weichenberger, E. Pozharski, B. Rupp. Acta Crystallogr. D73, 211-222 (2017). Extension of peptide ligands with a composite score. Tables only, not compatible with Twilight GUI.

2018-2020 Twilight reloaded (peptide) files, cumulative: pepligands-2020-01-15.tsv


Download program

Change Log

A new Twilight GUI, necessary due to SSL requirements and changes in the fetch destination for PDB data files, needs to be downloaded from here. The old 2012 version is still available here. Here is the readme file for installing Twilight and the update info.

Data files ->

Change Log

The Twilight program distribution comes packaged with a data file current to the respective revision year (i.e., the current 2018 Twilight program distribution includes data only up to 2018, and you need to also download the latest complete data file. Note that there are changes (See Change Log) to the archiving and possible differences on entry availibility due to retirement of the Uppsala EDS. 
Year Release date Complete data (.bz2) Yearly data (.bz2)
2019 Feb 09, 2020 ligands-2020-01-16.tsv newligands-2019.tsv
2018 Feb 07, 2019 ligands-2019-01-16.tsv newligands-2018.tsv
2017 Feb 17, 2018 ligands-2018-01-11.tsv newligands-2017.tsv
2016 Jan 11, 2017 ligands-2017-01-11.tsv newligands-2016.tsv
2015 Jan 13, 2016 ligands-2016-01-13.tsv newligands-2015.tsv
2014 Jan 15, 2015 ligands-2015-01-15.tsv newligands-2014.tsv
2013 Jan 15, 2014 ligands-2014-01-15.tsv newligands-2013.tsv
2012 Jan 16, 2013 ligands-2013-01-16.tsv newligands-2012.tsv
2012 First release ligands-2012-01-15.tsv N/A
The complete data file contains data up to the specified release date, whereas the yearly data file only contains PDB entries that have become available between the previous update's release date and the current release date.

NOTE: The data files are .bz2 compressed files- please unzip them before starting TWILIGHT

Here is the readme file for installing Twilight

My texbook

A useful resource if you want to learn more about protein crystallography, ligand structures, and structure validation

For now

You can still use the web tutorial for free!