Computational protocol: Molecular replacement then and now

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Protocol publication

[…] Analysis of the distribution in the PDB of the major software used to solve structures using MR from 1970 to date shows that over 90% of the structures to date have been solved using either X-PLOR/CNS (Brünger, 1992b ), AMoRe (Navaza, 1994), MOLREP (Vagin & Teplyakov, 2010) or Phaser (McCoy et al., 2007). Although X-PLOR/CNS and AMoRe have been widely used in the past, Phaser and MOLREP are probably the current cutting-edge choice for routine molecular-replacement efforts and some examples of these packages are briefly described below. Before that though, it is probably worth briefly mentioning the earlier programs, which are in many ways the precursors of what we have today. [...] MOLREP and Phaser are probably the current cutting-edge choice for routine molecular-replacement efforts, mostly owing to their full automation and ease of use. The first paper describing MOLREP was published in 1997 (Vagin & Teplyakov, 1997): in the view of the author, MOLREP can be seen as a more sophisticated and more versatile version of AMoRe, of which it retains many of the functionalities, with some useful improvements. It can be run as a fully automated package, including an automatic choice of all parameters and a soft resolution cutoff (instead of the normal low-resolution cutoff, it uses a special coefficient that allows the removal of structure factors in this resolution range without introducing a series-termination effect). In addition to the standard rotation and translation searches, it allows phased rotation and translation functions, and a locked cross-rotation function that can use as input the peak-list output of the self-rotation function. The self-rotation calculation also outputs a PostScript representation of the results, which is visually extremely clear. Another interesting feature is an original full-symmetry translation function combined with a packing function. Information from the model already placed in the cell is incorporated in both the translation and the packing functions and, if the initial number of monomers is known, it is possible to locate all of the monomers in one simple run. In addition, it automatically chooses from symmetry-related models the solution closest to the molecule(s) that have already been placed, thus avoiding the need for the researcher to run symmetry-based coordinate transformation to reposition the new solution. Phaser (McCoy et al., 2007) is, in the view of the author, probably the most efficient molecular-replacement package available to date. It is included in both the PHENIX (Adams et al., 2010) and the CCP4 software suites. It is actually a general macromolecular phasing tool, since it provides both molecular-replacement and experimental phasing methods. The phasing algorithms in Phaser were developed using maximum-likelihood and multivariate statistics and have proven to be significantly better than traditional methods. In the molecular-replacement mode, rotation and translation functions are followed by a packing function, which is used to identify the solutions with a minimal number of Cα clashes within a given distance (and thus likely to be the most correct): although the process of actually counting the clashes can be slow compared with computing a collision function, as is performed in MOLREP, the packing analysis provides a very powerful constraint on the translation function and in some cases provides a way to identify potentially correct solutions within a large set of incorrect solutions. When space-group ambiguity is present, Phaser allows all potential space groups to be scanned (or just those input by the user). Phaser can use single or multiple initial models (to search, for example, for hetero-assemblies).Both MOLREP and Phaser are included in MrBUMP (Keegan & Winn, 2007) and BALBES (Long et al., 2008), two automated molecular-replacement pipelines available from the CCP4 suite that starting from a target sequence and experimental structure factors will search for homologous structures in the PDB, create search models from the template structures, perform molecular replacement and test the solutions with several rounds of restrained refinement (Keegan et al., 2011). […]

Pipeline specifications

Software tools CNS, Molrep, PHENIX, CCP4
Application Protein structure analysis
Organisms Dipturus trachyderma