Dataset Information

Ab initio structure solution of a proteolytic fragment using ARCIMBOLDO.

ABSTRACT: Crystal structure determination requires solving the phase problem. This can be accomplished using ab initio direct methods for small molecules and macromolecules at resolutions higher than 1.2?Å, whereas macromolecular structure determination at lower resolution requires either molecular replacement using a homologous structure or experimental phases using a derivative such as covalent labeling (for example selenomethionine or mercury derivatization) or heavy-atom soaking (for example iodide ions). Here, a case is presented in which crystals were obtained from a 30.8?kDa protein sample and yielded a 1.6?Å resolution data set with a unit cell that could accommodate approximately 8?kDa of protein. Thus, it was unclear what had been crystallized. Molecular replacement with pieces of homologous proteins and attempts at iodide ion soaking failed to yield a solution. The crystals could not be reproduced. Sequence-independent molecular replacement using the structures available in the Protein Data Bank also failed to yield a solution. Ultimately, ab initio structure solution proved successful using the program ARCIMBOLDO, which identified two ?-helical elements and yielded interpretable maps. The structure was the C-terminal dimerization domain of the intended target from Mycobacterium smegmatis. This structure is presented as a user-friendly test case in which an unknown protein fragment could be determined using ARCIMBOLDO.

SUBMITTER: Abendroth J

PROVIDER: S-EPMC6130419 | biostudies-literature | 2018 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Ab initio structure solution of a proteolytic fragment using ARCIMBOLDO.

Abendroth Jan J Sankaran Banumathi B Myler Peter J PJ Lorimer Donald D DD Edwards Thomas E TE

Acta crystallographica. Section F, Structural biology communications 20180829 Pt 9

Crystal structure determination requires solving the phase problem. This can be accomplished using ab initio direct methods for small molecules and macromolecules at resolutions higher than 1.2 Å, whereas macromolecular structure determination at lower resolution requires either molecular replacement using a homologous structure or experimental phases using a derivative such as covalent labeling (for example selenomethionine or mercury derivatization) or heavy-atom soaking (for example iodide io ...[more]

PMID: 30198884

Similar Datasets

Project description:The phase problem remains a major barrier to overcome in protein structure solution by X-ray crystallography. In recent years, new molecular-replacement approaches using ab initio models and ideal secondary-structure components have greatly contributed to the solution of novel structures in the absence of clear homologues in the PDB or experimental phasing information. This has been particularly successful for highly α-helical structures, and especially coiled-coils, in which the relatively rigid α-helices provide very useful molecular-replacement fragments. This has been seen within the program AMPLE, which uses clustered and truncated ensembles of numerous ab initio models in structure solution, and is already accomplished for α-helical and coiled-coil structures. Here, an expansion in the scope of coiled-coil structure solution by AMPLE is reported, which has been achieved through general improvements in the pipeline, the removal of tNCS correction in molecular replacement and two improved methods for ab initio modelling. Of the latter improvements, enforcing the modelling of elongated helices overcame the bias towards globular folds and provided a rapid method (equivalent to the time requirements of the existing modelling procedures in AMPLE) for enhanced solution. Further, the modelling of two-, three- and four-helical oligomeric coiled-coils, and the use of full/partial oligomers in molecular replacement, provided additional success in difficult and lower resolution cases. Together, these approaches have enabled the solution of a number of parallel/antiparallel dimeric, trimeric and tetrameric coiled-coils at resolutions as low as 3.3 Å, and have thus overcome previous limitations in AMPLE and provided a new functionality in coiled-coil structure solution at lower resolutions. These new approaches have been incorporated into a new release of AMPLE in which automated elongated monomer and oligomer modelling may be activated by selecting `coiled-coil' mode.

Dataset Information

Ab initio structure solution of a proteolytic fragment using ARCIMBOLDO.

Publications

Ab initio structure solution of a proteolytic fragment using ARCIMBOLDO.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets