scholarly journals MR-REX: molecular replacement by cooperative conformational search and occupancy optimization on low-accuracy protein models

2018 ◽  
Vol 74 (7) ◽  
pp. 606-620 ◽  
Author(s):  
Jouko J. Virtanen ◽  
Yang Zhang
Keyword(s):  
Protein Structures ◽  
Target Position ◽  
Protein Crystal ◽  
Conformational Search ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
Homologous Proteins ◽  
X Ray ◽  
Replica Exchange Monte Carlo ◽  
Protein Models

Molecular replacement (MR) has commonly been employed to derive the phase information in protein crystal X-ray diffraction, but its success rate decreases rapidly when the search model is dissimilar to the target. MR-REX has been developed to perform an MR search by replica-exchange Monte Carlo simulations, which enables cooperative rotation and translation searches and simultaneous clash and occupancy optimization. MR-REX was tested on a set of 1303 protein structures of different accuracies and successfully placed 699 structures at positions that have an r.m.s.d. of below 2 Å to the target position, which is 10% higher than was obtained by Phaser. However, cases studies show that many of the models for which Phaser failed and MR-REX succeeded can be solved by Phaser by pruning them and using nondefault parameters. The factors effecting success and the parts of the methodology which lead to success are studied. The results demonstrate a new avenue for molecular replacement which outperforms (and has results that are complementary to) the state-of-the-art MR methods, in particular for distantly homologous proteins.

scholarly journals Room-temperature ultrahigh-resolution time-of-flight neutron and X-ray diffraction studies of H/D-exchanged crambin

2012 ◽  
Vol 68 (2) ◽  
pp. 119-123 ◽  
Author(s):  
Julian C.-H. Chen ◽  
Zoë Fisher ◽  
Andrey Y. Kovalevsky ◽  
Marat Mustyakimov ◽  
B. Leif Hanson ◽  
...  
Keyword(s):  
Neutron Diffraction ◽  
Room Temperature ◽  
Protein Structures ◽  
Atomic Displacement ◽  
Protein Crystal ◽  
X Ray Diffraction ◽  
Resolution Time ◽  
X Ray ◽  
Two Temperatures ◽  
Atomic Displacement Parameters

The room-temperature (RT) X-ray structure of H/D-exchanged crambin is reported at 0.85 Å resolution. As one of the very few proteins refined with anisotropic atomic displacement parameters at two temperatures, the dynamics of atoms in the RT and 100 K structures are compared. Neutron diffraction data from an H/D-exchanged crambin crystal collected at the Protein Crystallography Station (PCS) showed diffraction beyond 1.1 Å resolution. This is the highest resolution neutron diffraction reported to date for a protein crystal and will reveal important details of the anisotropic motions of H and D atoms in protein structures.

scholarly journals Nanovolume optimization of protein crystal growth using the microcapillary protein crystallization system

2010 ◽  
Vol 43 (5) ◽  
pp. 1078-1083 ◽  
Author(s):  
Cory J. Gerdts ◽  
Glenn L. Stahl ◽  
Alberto Napuli ◽  
Bart Staker ◽  
Jan Abendroth ◽  
...  
Keyword(s):  
Protein Crystallization ◽  
Protein Structures ◽  
Protein Crystal ◽  
X Ray Diffraction ◽  
Vapor Diffusion ◽  
X Ray ◽  
Gradient Optimization ◽  
Crystallization System ◽  
Initial Crystal ◽  
Novel Protein

The Microcapillary Protein Crystallization System (MPCS) is a microfluidic, plug-based crystallization technology that generates X-ray diffraction-ready protein crystals in nanolitre volumes. In this study, 28 out of 29 (93%) proteins crystallized by traditional vapor diffusion experiments were successfully crystallized by chemical gradient optimization experiments using the MPCS technology. In total, 90 out of 120 (75%) protein/precipitant combinations leading to initial crystal hits from vapor diffusion experiments were successfully crystallized using MPCS technology. Many of the resulting crystals produced high-quality X-ray diffraction data, and six novel protein structures that were derived from crystals harvested from MPCS CrystalCards are reported.

scholarly journals Comparison of protein structures determined by NMR in solution and by X-ray diffraction in single crystals

1992 ◽  
Vol 25 (3) ◽  
pp. 325-377 ◽  
Author(s):  
Martin Billeter
Keyword(s):  
Single Crystals ◽  
Protein Structures ◽  
Three Dimensional ◽  
Data Bank ◽  
Detailed Knowledge ◽  
X Ray Diffraction ◽  
Homologous Proteins ◽  
X Ray ◽  
Structural Details ◽  
Almost All

Following the first determinations of protein structures in the late 1950s and the early 1960s (see for example Kendrewet al.1960; Perutz, 1964), the three-dimensional structures of several hundred proteins have been elucidated by X-ray diffraction on single crystals. By the end of 1991, approximately 150 entries of proteins with substantially different sequences and a well resolved structure (Hobohmet al.1992) were deposited in the Protein Data Bank (Bernsteinet al.1977; Abolaet al.1987). In addition, many structures of homologous proteins or of mutants have been described, bringing the total number of entries to about 600. While it was soon accepted that almost all of these structures do indeed give a correct picture of the fold of the active protein in spite of the non-physiological environment of single crystals, it is not clear to what extent structural details are reliably described by these structures. In particular the surface of a protein may be modified due to the dense packing of protein molecules in the crystal lattice. A detailed knowledge of the protein surface is, however, essential for the understanding of the function of the protein.

scholarly journals Optimization in S-SAD phasing - difference between solved and unsolved structure

2014 ◽  
Vol 70 (a1) ◽  
pp. C613-C613
Author(s):  
Jan Stránský ◽  
Tomáš Kovaľ ◽  
Lars Østergaard ◽  
Jarmila Dušková ◽  
Tereza Skálová ◽  
...  
Keyword(s):  
Data Processing ◽  
De Novo ◽  
Protein Structures ◽  
X Ray Diffraction ◽  
X Ray ◽  
Structure Solution ◽  
Sad Phasing ◽  
Optimal Values ◽  
Grant Agency ◽  
Intensity Reading

Development of X-ray diffraction technologies have made de novo phasing of protein structures by single-wavelength anomalous dispersion by sulphur (S-SAD) more common. As anomalous differences in the sulphur atomic factors are in the order of errors of measurement, careful intensity reading and data processing are crucial. S-SAD was used for de novo phasing of a small 12 kDa protein with 4 sulphur atoms per molecule at 2.3 Å, where the data did not enable a straightforward structure solution. Data processing was performed using XDS [1] and scaling using XSCALE. The sulphur substructure was determined by SHELXD [2] and phases were obtained from SHELXE [2]. Both algorithms strongly depend on input parameters and default values did not lead to the correct phases. Therefore a systematic search of optimal values of several parameters was used to find a solution. This method helped to confirm sulphur substructure and to differentiate the handedness of the solutions. Moreover, a script for comfortable conversion of SHELX outputs to MTZ format was developed, using programmes included in the CCP4 package [3]. The previously unsolvable protein structure was successfully resolved with the described procedure. This work was supported by the Grant Agency of the Czech Technical University in Prague, (SGS13/219/OHK4/3T/14), the Czech Science Foundation (P302/11/0855), project BIOCEV CZ.1.05/1.1.00/02.0109 from the ERDF.

Crystallization and preliminary X-ray diffraction studies of human catalase

1999 ◽  
Vol 55 (9) ◽  
pp. 1614-1615 ◽  
Author(s):  
R. A. P. Nagem ◽  
E. A. L. Martins ◽  
V. M. Gonçalves ◽  
R. Aparício ◽  
I. Polikarpov
Keyword(s):  
Search Model ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
Beef Liver ◽  
X Ray ◽  
Diffusion Technique ◽  
Cell Dimensions ◽  
Synchrotron Radiation Diffraction ◽  
Replacement Solution ◽  
Unit Cell Dimensions

The enzyme catalase (H2O2–H2O2 oxidoreductase; E.C. 11.1.6) was purified from haemolysate of human placenta and crystallized using the vapour-diffusion technique. Synchrotron-radiation diffraction data have been collected to 1.76 Å resolution. The enzyme crystallized in the space group P212121, with unit-cell dimensions a = 83.6, b = 139.4, c = 227.5 Å. A molecular-replacement solution of the structure has been obtained using beef liver catalase (PDB code 4blc) as a search model.

scholarly journals Hydrogen atoms in protein structures: high-resolution X-ray diffraction structure of the DFPase

2013 ◽  
Vol 6 (1) ◽  
pp. 308 ◽  
Author(s):  
Mikael Elias ◽  
Dorothee Liebschner ◽  
Jurgen Koepke ◽  
Claude Lecomte ◽  
Benoit Guillot ◽  
...  
Keyword(s):  
High Resolution ◽  
Protein Structures ◽  
X Ray Diffraction ◽  
Hydrogen Atoms ◽  
X Ray ◽  
Diffraction Structure

scholarly journals X-ray crystal structure of a malonate-semialdehyde dehydrogenase fromPseudomonassp. strain AAC

2017 ◽  
Vol 73 (1) ◽  
pp. 24-28 ◽  
Author(s):  
Matthew Wilding ◽  
Colin Scott ◽  
Thomas S. Peat ◽  
Janet Newman
Keyword(s):  
Crystal Structure ◽  
Search Model ◽  
Molecular Replacement ◽  
X Rays ◽  
X Ray Diffraction ◽  
Acetyl Coa ◽  
Space Group ◽  
X Ray ◽  
Semialdehyde Dehydrogenase

The NAD-dependent malonate-semialdehyde dehydrogenase KES23460 fromPseudomonassp. strain AAC makes up half of a bicistronic operon responsible for β-alanine catabolism to produce acetyl-CoA. The KES23460 protein has been heterologously expressed, purified and used to generate crystals suitable for X-ray diffraction studies. The crystals belonged to space groupP212121and diffracted X-rays to beyond 3 Å resolution using the microfocus beamline of the Australian Synchrotron. The structure was solved using molecular replacement, with a monomer from PDB entry 4zz7 as the search model.

scholarly journals Purification, crystallization and preliminary X-ray diffraction analysis of a hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyltransferase (HCT) fromCoffea canephorainvolved in chlorogenic acid biosynthesis

2012 ◽  
Vol 68 (7) ◽  
pp. 824-828 ◽  
Author(s):  
Laura A. Lallemand ◽  
James G. McCarthy ◽  
Sean McSweeney ◽  
Andrew A. McCarthy
Keyword(s):  
Structural Data ◽  
Coffea Canephora ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Key Enzyme ◽  
Substrate Specifities ◽  
Drop Technique ◽  
Better Than

Chlorogenic acids (CGAs) are a group of soluble phenolic compounds that are produced by a variety of plants, includingCoffea canephora(robusta coffee). The last step in CGA biosynthesis is generally catalysed by a specific hydroxycinnamoyl-CoA quinate hydroxycinnamoyltransferase (HQT), but it can also be catalysed by the more widely distributed hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyltransferase (HCT). Here, the cloning and overexpression of HCT fromC. canephorainEscherichia colias well as its purification and crystallization are presented. Crystals were obtained by the sitting-drop technique at 293 K and X-ray diffraction data were collected on the microfocus beamline ID23-2 at the ESRF. The HCT crystals diffracted to better than 3.0 Å resolution, belonged to space groupP42212 with unit-cell parametersa=b= 116.1,c= 158.9 Å and contained two molecules in the asymmetric unit. The structure was solved by molecular replacement and is currently under refinement. Such structural data are needed to decipher the molecular basis of the substrate specifities of this key enzyme, which belongs to the large plant acyl-CoA-dependent BAHD acyltransferase superfamily.

scholarly journals Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of AerF fromMicrocystis aeruginosa, a putative reductase participating in aeruginosin biosynthesis

2015 ◽  
Vol 71 (4) ◽  
pp. 466-470 ◽  
Author(s):  
Ruyi Ding ◽  
Cui Xu ◽  
Xu Chen ◽  
Mengyun Bao ◽  
Xiaoting Qiu
Keyword(s):  
Diffraction Analysis ◽  
Biosynthetic Pathway ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
Antithrombotic Activity ◽  
X Ray ◽  
Cell Parameters ◽  
Maximum Resolution ◽  
Replacement Method ◽  
Molecular Replacement Method

The 2-carboxy-6-hydroxyoctahydroindole moiety is an essential residue for the antithrombotic activity of aeruginosins, which are a class of cyanobacteria-derived bioactive linear tetrapeptides. The biosynthetic pathway of the 2-carboxy-6-hydroxyoctahydroindole moiety has not yet been resolved. AerF was indicated to be involved in the biosynthesis of the 2-carboxy-6-hydroxyoctahydroindole moiety. This study reports the cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of AerF fromMicrocystis aeruginosawith a C-terminal His6tag. The crystal diffracted to a maximum resolution of 1.38 Å and belonged to the tetragonal space groupP4322, with unit-cell parametersa=b= 101.581,c= 116.094 Å. The calculated Matthews coefficient and solvent content of the crystal were 2.47 Å3 Da−1and 50.32%, respectively. The initial model of the structure was obtained by the molecular-replacement method and refinement of the structure is in progress.

scholarly journals MSMEG_6292, a Mycobacterium smegmatis RNA polymerase secondary channel-binding protein: purification, crystallization and X-ray diffraction analysis

2018 ◽  
Vol 74 (9) ◽  
pp. 543-548
Author(s):  
Abyson Joseph ◽  
Valakunja Nagaraja ◽  
Ramanathan Natesh
Keyword(s):  
Rna Polymerase ◽  
Mycobacterium Smegmatis ◽  
Transcriptional Activity ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
Secondary Channel ◽  
X Ray ◽  
Cell Parameters ◽  
Crystallization Method ◽  
Better Than

The transcriptional activity of RNA polymerase (RNAP) is controlled by a diverse set of regulatory factors. A subset of these regulators modulate the activity of RNAP through its secondary channel. Gre factors reactivate stalled elongation complexes by enhancing the intrinsic cleavage activity of RNAP. In the present study, the protein MSMEG_6292, a Gre-factor homologue from Mycobacterium smegmatis, was expressed heterologously in Escherichia coli and purified using standard chromatographic techniques. The hanging-drop vapour-diffusion crystallization method yielded diffraction-quality crystals. The crystals belonged to the trigonal space group P3121 (or its enantiomorph P3221), with unit-cell parameters a = b = 83.15, c = 107.07 Å, α = β = 90, γ = 120°. The crystals diffracted to better than 3.0 Å resolution. Molecular-replacement attempts did not yield any phasing models; hence, platinum derivatization was carried out with K2PtCl4 and derivative data were collected to 3.4 Å resolution.

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