Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Protein Sci
2016 Mar 01;253:720-33. doi: 10.1002/pro.2861.
Show Gene links
Show Anatomy links
Protein purification and crystallization artifacts: The tale usually not told.
Niedzialkowska E
,
Gasiorowska O
,
Handing KB
,
Majorek KA
,
Porebski PJ
,
Shabalin IG
,
Zasadzinska E
,
Cymborowski M
,
Minor W
.
???displayArticle.abstract???
The misidentification of a protein sample, or contamination of a sample with the wrong protein, may be a potential reason for the non-reproducibility of experiments. This problem may occur in the process of heterologous overexpression and purification of recombinant proteins, as well as purification of proteins from natural sources. If the contaminated or misidentified sample is used for crystallization, in many cases the problem may not be detected until structures are determined. In the case of functional studies, the problem may not be detected for years. Here several procedures that can be successfully used for the identification of crystallized protein contaminants, including: (i) a lattice parameter search against known structures, (ii) sequence or fold identification from partially built models, and (iii) molecular replacement with common contaminants as search templates have been presented. A list of common contaminant structures to be used as alternative search models was provided. These methods were used to identify four cases of purification and crystallization artifacts. This report provides troubleshooting pointers for researchers facing difficulties in phasing or model building.
Almo,
Protein production from the structural genomics perspective: achievements and future needs.
2013, Pubmed
Almo,
Protein production from the structural genomics perspective: achievements and future needs.
2013,
Pubmed
Berman,
The Protein Data Bank.
2000,
Pubmed
Bibby,
AMPLE: a cluster-and-truncate approach to solve the crystal structures of small proteins using rapidly computed ab initio models.
2012,
Pubmed
Bolanos-Garcia,
Structural analysis and classification of native proteins from E. coli commonly co-purified by immobilised metal affinity chromatography.
2006,
Pubmed
Chen,
MolProbity: all-atom structure validation for macromolecular crystallography.
2010,
Pubmed
Chu,
Comparison of sequence and structure-based datasets for nonredundant structural data mining.
2005,
Pubmed
Collins,
Policy: NIH plans to enhance reproducibility.
2014,
Pubmed
Cowtan,
The Buccaneer software for automated model building. 1. Tracing protein chains.
2006,
Pubmed
Dong,
In situ proteolysis for protein crystallization and structure determination.
2007,
Pubmed
Emsley,
Features and development of Coot.
2010,
Pubmed
Emsley,
Coot: model-building tools for molecular graphics.
2004,
Pubmed
Eschenfeldt,
A family of LIC vectors for high-throughput cloning and purification of proteins.
2009,
Pubmed
Grosse-Kunstleve,
Numerically stable algorithms for the computation of reduced unit cells.
2004,
Pubmed
Gräslund,
Protein production and purification.
2008,
Pubmed
Hassell,
Crystallization of protein-ligand complexes.
2007,
Pubmed
Holm,
Dali server: conservation mapping in 3D.
2010,
Pubmed
Judge,
Protein purification by bulk crystallization: the recovery of ovalbumin.
1995,
Pubmed
Kato,
Copurification of small heat shock protein with alpha B crystallin from human skeletal muscle.
1992,
Pubmed
Keegan,
MrBUMP: an automated pipeline for molecular replacement.
2008,
Pubmed
Kim,
High-throughput protein purification and quality assessment for crystallization.
2011,
Pubmed
Krissinel,
Secondary-structure matching (SSM), a new tool for fast protein structure alignment in three dimensions.
2004,
Pubmed
Langer,
Automated macromolecular model building for X-ray crystallography using ARP/wARP version 7.
2008,
Pubmed
Liu,
Lysozyme contamination facilitates crystallization of a heterotrimeric cortactin-Arg-lysozyme complex.
2012,
Pubmed
Lohkamp,
A mixture of fortunes: the curious determination of the structure of Escherichia coli BL21 Gab protein.
2008,
Pubmed
Long,
BALBES: a molecular-replacement pipeline.
2008,
Pubmed
Majorek,
Double trouble-Buffer selection and His-tag presence may be responsible for nonreproducibility of biomedical experiments.
2014,
Pubmed
Majorek,
Structural, functional, and inhibition studies of a Gcn5-related N-acetyltransferase (GNAT) superfamily protein PA4794: a new C-terminal lysine protein acetyltransferase from pseudomonas aeruginosa.
2013,
Pubmed
Marchler-Bauer,
CDD: NCBI's conserved domain database.
2015,
Pubmed
Murshudov,
Refinement of macromolecular structures by the maximum-likelihood method.
1997,
Pubmed
Otwinowski,
Processing of X-ray diffraction data collected in oscillation mode.
1997,
Pubmed
Porebski,
Fitmunk: improving protein structures by accurate, automatic modeling of side-chain conformations.
2016,
Pubmed
Prinz,
Believe it or not: how much can we rely on published data on potential drug targets?
2011,
Pubmed
Rychlewski,
Comparison of sequence profiles. Strategies for structural predictions using sequence information.
2000,
Pubmed
Sauder,
High throughput protein production and crystallization at NYSGXRC.
2008,
Pubmed
Sheldrick,
A short history of SHELX.
2008,
Pubmed
Smyth,
Crystal structures of fusion proteins with large-affinity tags.
2003,
Pubmed
Terwilliger,
SOLVE and RESOLVE: automated structure solution, density modification and model building.
2004,
Pubmed
Vagin,
Molecular replacement with MOLREP.
2010,
Pubmed
Winn,
Overview of the CCP4 suite and current developments.
2011,
Pubmed
Ye,
FATCAT: a web server for flexible structure comparison and structure similarity searching.
2004,
Pubmed