| Newsletter No. 364
|| November 6, 2017
| TIPS, TRICKS, METHODS - A Convenient Phasing Vehicle
Zbigniew Dauter (NCI): Selenourea: A Convenient Phasing Vehicle
Majority of novel X-ray crystal structures of proteins are currently solved using the anomalous diffraction signal provided by selenium after incorporation of selenomethionine instead of natural methionine by genetic engineering methods. However, selenium can be also inserted into protein crystals in the form of selenourea (SeC(NH2)2), by adding selenourea into mother liquor or cryo-solution or in the form of powder into a drop with native crystals, in analogy to the classic procedure of heavy-atom derivatization. Selenourea is able to bind to reactive groups at the surface of macromolecules primarily through hydrogen bonds, where the selenium atom may serve as acceptor and amide groups as H-bond donors. Selenourea has different chemical properties than other heavy-atom reagents and halide ions and provides a convenient way of phasing crystal structures of macromolecules. A de novo protein crystal structure at low resolution of 2.9 Å with total 1125 residues distributed in seven chains in asymmetric unit at was recently successfully solved with this method. For details, see “Selenourea: a convenient phasing vehicle for macromolecular X-ray crystal structures,” Sci. Rep. 6, 37123 (2016) by Zhipu Luo.
Xinhua Ji: It was recently reported that microseeding was used to produce untwinned crystals of an O-demethylase using twinned crystals as seeds (Acta Crystal. F72:897-902, 2016). Microseeding separates nucleation events from crystal growth events so that it can yield different crystal forms. Hence, it is promising that this technique can be used to produce untwinned crystals from twinned seeds.
ARCHIVE: Introduction, Pre-crystallization, Crystallization, Post-crystallization, Derivatization, Cryoprotection, Diffraction, Symmetry, Structure Solution, Structure Refinement, Structure Analysis & Presentation, Biophysical Methods.
DISCUSSION - Maps, Models, and Structures
Alexander Wlodawer, Mi Li, Zbigniew Dauter: High-Resolution Cryo-EM Maps and Models: A Crystallographer's Perspective (PMID: 28867613)
The appearance of ten high-resolution cryoelectron microscopy (cryo-EM) maps of proteins, ribosomes, and viruses was compared with the experimentally phased crystallographic electron density maps of four proteins. We found that maps calculated at a similar resolution by the two techniques are quite comparable in their appearance, although cryo-EM maps, even when sharpened, seem to be a little less detailed. An analysis of models fitted to the cryo-EM maps indicated the presence of significant problems in almost all of them, including incorrect geometry, clashes between atoms, and discrepancies between the map density and the fitted models. In particular, the treatment of the atomic displacement (B) factors was meaningless in almost all analyzed cryo-EM models. Stricter cryo-EM structure deposition standards and their better enforcement are needed.
Mariusz Jaskolski: Pathological Behavior of Atomic Occupancy Factors
ARCHIVE: Test-set-and-R-free, Twinning, Low Resolution Crystallography, PHASER, HKL2000, Parallel Expression, NCS, Missing Atoms, Trends in Crystallography, Absorption Correction, Data for Refinement and Publication.
| LECTURES AND TUTORIALS - CRYSTALLOGRAPHY
DR. ZBIGNIEW DAUTER'S LECTURES AT THE NIH (2005)
Part 1: "How to read international tables?"
Part 2: "Data collection strategy" and "Twinning"
"Phasing methods - a general introduction to all methods"
Part 3: "SAD phasing, Quick halide soaking, and Radiation damage
with possible use of it for phasing"
RIGAKU WEBINAR SERIES (2009 - PRESENT)
LOW RESOLUTION PHASING AND REFINEMENT (2011)
CRYSTALLOGRAPHY: SEEING THINGS IN A DIFFERENT LIGHT (2013)
CRYSTALLOGRAPHY: FOR ASPIRING CRYSTALLOGRAPHERS (2013)
STRUCTURE FACTOR TUTORIAL (2014)
DATA COLLECTION FOR STRUCTURE DETERMENATION (2014)
ACHESYM: AN ALGORITHM AND SERVER FOR STANDARDIZED PLACEMENT OF MACROMOLECULAR MODELS IN THE UNIT CELL (2014)
A GLIMPSE OF STRUCTURAL BIOLOGY THROUGH X-RAY CRYSTALLOGRAPHY (2014)
CRYSTAL CLEAR (2014)
NATIVE SAD IS MATURING (2015)
LITERATURE ON CRYSTALLOGRAPHY THEORY AND METHODS (2017)
PROTEIN CRYSTALLOGRAPHY Methods and Protocols (2017)
| LINKS - New Server: RestraintLib
Databases: BMCD, DisProt, ExPASy, HAD, HIC-Up, Metal Sites, NDB, PDB, PDBe,
Protein Geometry, Scattering
Programs: CCP4, COOT, DSSR, HKL, PHENIX, PyMOL, SOLVE, XDS
Servers: ACHESYM, Anisotropy, CheckMyMetal, Crystal, C6, Dali, DSSR, ESPript,
Grade, PDBePISA, Phyre, MolProbity, Protein, Robetta Fragment, HHpred,
Facilities: APS SER-CAT, APS SAXS Capabilities
|Copyright © NIH X-Ray Diffraction Group Maintained by Dr. Xinhua Ji|
|on the NIH-NCI-CCR-MCL server (http://mcl1.ncifcrf.gov)|