ATOLL

Visualization tool for transmembrane protein structures. Visit the atoll.drugdesign.unistra.fr website to run ATOLL using a GUI.

Description

ATOLL (Aligned Transmembrane dOmain Layout fLattening) is a program for visualizing 3D structures of the transmembrane helix bundle of membrane proteins. ATOLL projects portions of helices onto the plane of the membrane, as if looking at the protein from above the membrane. It allows the comparison of different conformations of the same protein or of different proteins. ATOLL principle and examples of applications are described in an application note submitted to Bioinformatics.

Requirements

ATOLL is written in Python 3 and requires the NumPy, SciPy, Matplotlib, PyYAML, Biopython and MDAnalysis modules. It is strongly recommended to create an Anaconda environment using the conda3.yaml file provided. The program is compatible with Windows, Mac OS and Linux operating systems.

How ATOLL works

Input files

To launch ATOLL, several files must be provided:

• structure(s) to be analysed
• reference structure
• sequence alignment file
• annotation file

Structure file(s)

The structure(s) that will be analysed by ATOLL are provided in one or several structure files. Each structure file contain one or more structures. It is possible to compare different proteins represented each by multiple structures (e.g., comparison of the molecular dynamics (MD) trajectories of two proteins). The file can contain, in addition to the proteins to be studied, other types of molecule (solvent, lipids, etc...). However, it is recommended to remove all non-essential objects from the file, to avoid increasing input reading time, especially in the case of molecular dynamics (MD) trajectories.

Warning: If a protein is an oligomer (several chains), it must be described as a single chain with an incremental numbering of the residues. You can use the "sanitize" routine to process your structure.

python sanitize bin/atoll.py --structure <structure_file_to_process> --output <the_processed_structure_file>

The supported file formats and corresponding extensions are :

• Static
  • Protein Data Bank (PDB): .pdb
  • SYBYL (MOL2): .mol2
• Multiple
  • Topology
    • AMBER TOP: .prmtop, .top, parm7
    • CHARMM PSF: .psf
    • Protein Data Bank (PDB): .topdb
  • Coordinate
    • AMBER CRD: .inpcrd
    • AMBER RST: .inprst
    • AMBER TRJ: .trj
    • AMBER NetCDF: .ncdf, .nc
    • CHARMM DCD: .dcd
    • GROMACS XTC: .xtc
    • GROMACS TRR: .trr

Static format refers to structure file usually containing a single structure (PDB or MOL2), while multiple format refers to MD trajectories (CRD, RST, NC, DCD, ...).

For molecular dynamics trajectories, the topology file and coordinate file(s) of each protein must be placed in a dedicated directory. ATOLL scans the directory to find the topology and coordinate files from their extension.

Depending on the static or multiple nature of the file, ATOLL uses different representations (see publication).

Important : If the structure files contain proteins which differ in their sequence and / or in the numbering of residues, a sequence alignment file must be provided too (see 'Sequence alignment file' below).

Reference file

The reference structure defines the correct orientation of the protein in the membrane, which is XY plane.

There are three ways of preparing the reference structure in the appropriate coordinate frame: using with your favorite software for protein display and edition; using ATOLL webserver (https://atoll.drugdesign.unistra.fr); or downloading a structure from the structural database of orientations of proteins in membrane OPM. Note that if there are several structures in the reference file, ATOLL only takes into account the first one, the following ones being ignored. The supported file formats are those of static structures.

The reference structure is also used to define the projected TMs. If the reference structure is different from the input proteins, its sequence must be given in the alignment file (see 'Sequence alignment file' below).

All the input structures must be superimposed onto the TMs of the reference structure.

Sequence alignment file

A multiple sequence alignement is requiered if the protein sequence is not strictly conserved between the reference and all the entries, i.e., if there are differences in the sequence length and composition (e.g., missing residues), or if residue numbering is inconsistent.

The stockholm file format (.sto or .stk) is the only supported format. Format developed by E Sonnhammers (https://sonnhammer.sbc.su.se/Stockholm.html)

ATOLL requieres the following features and tags:

• #GF SG : number of sequences in alignment
• #GS RE reference : definition of the reference

Example: 4 sequences aligned, the third one (P2X7_RAT/1-60) is the reference

# STOCKHOLM 1.0
#=GF SQ 4
#=GS P2X7_RAT/1-60 RE reference
P2X4_ZEBRAFISH/1-40       --------------------GSSKKVGTLNRFTQALVIAYVIGYVFVYNKGYQDTDTVL-S
amP2X_GULFCOASTTICK/1-43  -----------------GSREFDQKIGVLNRLIQLLILGYIIGYVIIYQKGYQQFSTFN-A
P2X7_RAT/1-60             MPACCSWNDVFQYETNKVTRIQSVNYGTIKWILHMTVFSY-VSFALMSDKLYQRKEPLI-S
P2X3_HUMAN/1-57           ----GSRADFFTYETPKVIVVKSWTIGIINRVVQLLIISYFVGWVFLHEKAYQVRDTAIES

Annotation file

The annotation file is mandantory. It defines ATOLL parameters. Data is formatted as a table in CSV (comma separator) or TSV (tab separator) format, and comprising 6 fields:

• "Entry": unique label for each entry.
• "Sequence name": associated label in the sequence alignment file.
• "Group": define a group for several entries.
• "Type": specifies whether it is a static or multiple entry.
• "Path": path of the entry. Must be a file if the entry is static and a directory if the entry is multiple.
• "Color": entry color when generating the images. For a full list of colors see the following link

Each static structure and each directory with multiple structures is considered as an entry.

Running ATOLL

The program is launched via a terminal. Here is an example from tutorial 02:

python atoll ../../bin/atoll.py -ref 4lde_clean.pdb -seq sequences.sto -out results -rn position -inf info.tsv -ra 16-31+51-67+86-103+132-148+177-194+255-269+289-302 -rh 10-38+46-74+82-113+126-147+175-203+246-276+284-302 --overwrite

In detail, the different options are :

• -ref: path of the reference structure.
• -seq: path of the sequence alignment file.
• -inf: path of the annotation file.
• -out: path of the directory where outputs will be stored.
• -ra: residues used during structural alignment.
• -rh: definition of the transmembrane helices to be projected.
• -rn: how the residues provided in -ra and -rh are interpreted. The allowed values are "position" which corresponds to the position of the residues in the sequence alignment file or in the reference structure, and "resid" which corresponds to the residue number.
• --overwrite: If the output directory exists, it will be overwritten.

NB: The residue synthax in -ra and -rh allows to define a range with the character '-' and to separate the ranges with the character '+'.

FUNDING

This work was supported by the Agence Nationale de Recherches sur le Sida [AO 2016-2 CSS2 17252 to E.K, PhD grant to C.J.]; and the Laboratoire d’Excellence Medalis de Agence Nationale de la Recherche [ANR_10-LABX-0034 to EK].