IPCAS: a direct-method-based pipeline from phasing to model building and refinement for macromolecular structure determination

Abstract

A new version (2.0) of the pipeline IPCAS (Iterative Protein Crystal structure Automatic Solution) has been released, in which the program OASIS performs direct-method single-wavelength anomalous diffraction/single isomorphous replacement phasing and direct-method-aided partial-structure extension. IPCAS incorporates the widely used packages CCP4 and PHENIX for locating heavy atoms, density modification, molecular replacement, model building and refinement. Important extensions to the previous version of IPCAS include a resolution screening method for non-crystallographic symmetry searching, an alternate model-building protocol for avoiding premature convergence and direct-method image processing for electron microscopy maps, including singleparticle cryo-EM maps. Moreover, a new graphical user interface is provided for controlling and real-time monitoring of the whole dual-space iterative process, which works as a plugin to CCP4i. Applications of the new IPCAS to difficult cases have yielded promising results, including 'direct-method phasing and fragment extension' from weak anomalous diffraction signal data and 'directmethod-aided partial-structure extension' from low-homology models.

Solving protein structures by combining structure prediction, molecular replacement and direct-methods-aided model completion

“…Specifically, we tested different combinations of predicted models by Alpha-Fold (Jumper et al, 2021) from CASP14 (Kryshtafovych, Schwede et al, 2021) in three cases: full-length model, multiple single-domain models and individual single-domain model. In this strategy, we performed MR using Phaser (McCoy et al, 2007) in CCP4 (Winn et al, 2011), followed by phases extension using OASIS (Zhang et al, 2010), density modification using DM (Cowtan, 1994) or Parrot (Cowtan, 2010), and alternative model building using Phenix.AutoBuild (Terwilliger et al, 2008) and Buccaneer (Cowtan, 2006) within the framework of IPCAS 2.0 (Ding et al, 2020). Our results demonstrate that our approach effectively corrects model bias introduced by the predicted models and improves the final structures.…”

“…The incorrect placement of the D2 model directly resulted in a high phase error (74 ) of the MR structure in case 26. However, due to the unique NCS search function in IPCAS, the accurate NCS matrix was successfully obtained in the first cycle of the IPCAS iteration (Ding et al, 2020), and the missing part of model was completed in the third cycle, reducing the phase error to approximately 25 . The remaining three cases have a more ideal starting structure, so the correction in IPCAS also works well.…”

Crystal Structure of Mycobacterium tuberculosis Elongation Factor G1

“…The initial phases had a figure of merit (FOM) of 0.444 in the resolution range 47.95-3.30 Å. The intrinsic phase ambiguity was broken by iterative direct-methods SAD phasing rooted in the pipeline Iterative Protein Crystal structure Automatic Solution (IPCAS) (Ding et al, 2020). The iteration control was set as OASIS-DM-AutoBuild/Buccaneer with 15 cycles.…”

Direct phasing algorithm for protein crystals with high solvent content using low-resolution diffraction data

“…Until now, current IPA-based phasing algorithms have not been adopted for routine application in protein structure determination. Nevertheless, dualspace constraints have many applications in substructure determination (Weeks & Miller, 1999;Schneider & Sheldrick, 2002), phase extension (Ding et al, 2020) and density modification (Main, 1990;Rossmann, 1995). With regard to density constraints, solvent flattening (Wang, 1985), histogram matching (Zhang & Main, 1990) and NCS averaging (Bricogne, 1974) are the most common methods.…”