ARCIMBOLDO_SHREDDER Spherical tutorial

Aims of the tutorial

This tutorial shows how to use the spherical mode in ARCIMBOLDO-SHREDDER¹. This mode generates, starting from a distant homolog template that is annotated in terms of secondary and tertiary structure, a set of compact search models used as a library in ARCIMBOLDO BORGES, profiting from specific options in PHASER and SHELXE for model improvement. This example uses the latest ARCIMBOLDO_SHREDDER as of July 2018.

Experimental details and data

Test data for our tutorial have been obtained from the protein Slt (PDB ID: 5OHU). It is a lytic transglycosylase from Pseudomonas aeruginosa. This structure was solved² with data extending to 2.1 Å. The asymmetric unit contains one monomer of 642 residues.

Data are summarized in the following table:

Figure 1. Slt Structure colored after B-value.

Step by Step tutorial

1. Selection of a starting template

A BLAST³ search of the LTG sequence against the PDB finds a model, 1QSA, with 33% of sequence identity over a range of 603 amino acids.

Figure 2. LTG alignment against 1QSA

Figure 3. LTG final structure (blue) superposed with 1QSA template (orange).

A classical molecular replacement approach with the 1QSA model fails due to the high deviation between the model and the final structure, which presents an overall similar core but many local differences. In fact, the r.m.s.d. between both structures is 4.6 Å over a core of 582 Ca atoms (Figure 3)

Required Input

All required files can be downloaded here.

The description of the configuration file follows:

[CONNECTION]:
distribute_computing: local_grid
setup_bor_path: /path/to/setup.bor

[LOCAL]
path_local_phaser: /path/to/local_phaser
path_local_shelxe: /path/to/local_shelxe

[GENERAL]:
working_directory: /path/to/working_directory
mtz_path: %(working_directory)s/5ohu.mtz
hkl_path: %(working_directory)s/5ohu.hkl

[ARCIMBOLDO-SHREDDER]
name_job: 5ohu_spheres
molecular_weight: 70153
f_label: F
sigf_label: SIGF
number_of_component: 1
model_file: /absolute/path/to/1qsacut.pdb
SHRED_METHOD: spherical
rmsd_shredder: 1.2
alixe: True

Execution

You can run the program from the ccp4i interface or from a terminal, redirecting the output to a log file. This tutorial describes use through a command file, as interfaces are self-explanatory.

1. Interactively

ARCIMBOLDO_SHREDDER 5ohu.bor

2. In background

nohup ARCIMBOLDO_SHREDDER 5ohu.bor >& log &

Output and Results

In the directory where you launched ARCIMBOLDO_SHREDDER, you will find a directory called models containing a library of pdbs. Around each Calpha in the template, models are cut in a spatial way, producing a set of non-redundant, overlapping, compact models. In the default mode, they are also annotated in different chains in order to decompose them and perform gyre and gimble⁶ refinement, aiming to give more degrees of freedom and obtain a more accurate model.

The directory called ARCIMBOLDO_BORGES contains the output of the ARCIMBOLDO_BORGES run using the library in models.The html output of the library search is found here. The first section echoes all parameters used for the run, so that defaults are listed along with values for the parameters set through the .bor file. This allows to reproduce the run even if defaults may change in future versions.The next section displays a graph and a table summarizing the rotation clustering step as in ARCIMBOLDO_BORGES⁷. A sortable table follows, summarizing the results for all PHASER and SHELXE steps, including top and average figures of merit for each rotation cluster that has been evaluated.

The last section of the html contains links to access the best current solution: the pdb of the traced structure and the map in .phs format. In this case, a map coming from a combination of 61 solutions, using as a reference the solution produced in rotation cluster 7 by the model obtained using as center the residue 589, produces a complete interpretable map after density modification and autotracing with SHELXE⁸. Figure 4 shows a detail of the map before density modification (orange map) and autotracing and after (blue map). A correlation coefficient of 36.76% with 475 residues traced is obtained.

Figure 4. A detail of the electron density map before autotracing (orange) and after (blue).