Computational protocol: Use of noncrystallographic symmetry for automated model building at medium to low resolution

Similar protocols

Protocol publication

[…] In order to carry out computational tests, we used high-resolution structures from the PDB as well as a number of structures that had been submitted to the ARP/wARP model-building web service and made available for testing purposes. A good representative example is the 1.6 Å resolution structure of the B subunit of a mutated shiga-like toxin (PDB entry 1c48; Ling et al., 1998). The molecule is arranged as a homopentamer, with each subunit composed of 69 residues. This structure was predominantly used for the basic development of the method. The full test set used for subsequent examination of the effectiveness of the method consisted of 11 multimeric structures that were determined at resolutions ranging from 2.4 to 3.2 Å with asymmetric unit contents of between 300 and 2300 residues in 2–10 NCS-related subunits. The structures were characterized by varying secondary-structural content, so that there were predominantly helical, stranded and mixed α–β models. [...] We initially tested the ability of the PNS Extender module to automatically identify and apply NCS relations to the appropriate parts of the model: the ‘exclusion’ test. A single model, the mutated shiga-like toxin B subunit (PDB entry 1c48), was used for this purpose. We artificially fragmented the structure by cutting out parts of the model in order to mimic real cases, in which intermediate models may contain a large number of unconnected fragments. To generate cases with various degrees of fragmentation, we built ten differently fragmented structures. Starting from the complete structure, we successively deleted 5% of residues from each model, with 95% of the structure left in the first test case, 90% in the second case through to 50% in the tenth case. The models were fragmented by cutting out blocks of residues (15–30 amino acids; see Fig. 6b for the seventh case with 65% of the model left) from different parts of the structure.Subsequently, we evaluated the extent of the improvement observed when all of the test structures described in §2.1 were built using the automated model-building protocol of ARP/wARP including the PNS Extender module. Each protocol was executed with five cycles of model update and refinement after each of the ten model-building cycles. For these tests, we used ARP/wARP v.7.2, REFMAC v.5.5.0109 and CCP4 v.6.1.13 (Winn et al., 2011). […]

Pipeline specifications

Software tools ARP/wARP, CCP4
Application Protein structure analysis