Publications by authors named "Alexei A Vagin"

12 Publications

  • Page 1 of 1

Crystal structure of D-serine dehydratase from Escherichia coli.

Biochim Biophys Acta 2012 Mar 27;1824(3):422-32. Epub 2011 Nov 27.

The Institute of Crystallography, Russian Academy of Sciences, Moscow, Russia.

D-Serine dehydratase from Escherichia coli is a member of the β-family (fold-type II) of the pyridoxal 5'-phosphate-dependent enzymes, catalyzing the conversion of D-serine to pyruvate and ammonia. The crystal structure of monomeric D-serine dehydratase has been solved to 1.97Å-resolution for an orthorhombic data set by molecular replacement. In addition, the structure was refined in a monoclinic data set to 1.55Å resolution. The structure of DSD reveals a larger pyridoxal 5'-phosphate-binding domain and a smaller domain. The active site of DSD is very similar to those of the other members of the β-family. Lys118 forms the Schiff base to PLP, the cofactor phosphate group is liganded to a tetraglycine cluster Gly279-Gly283, and the 3-hydroxyl group of PLP is liganded to Asn170 and N1 to Thr424, respectively. In the closed conformation the movement of the small domain blocks the entrance to active site of DSD. The domain movement plays an important role in the formation of the substrate recognition site and the catalysis of the enzyme. Modeling of D-serine into the active site of DSD suggests that the hydroxyl group of D-serine is coordinated to the carboxyl group of Asp238. The carboxyl oxygen of D-serine is coordinated to the hydroxyl group of Ser167 and the amide group of Leu171 (O1), whereas the O2 of the carboxyl group of D-serine is hydrogen-bonded to the hydroxyl group of Ser167 and the amide group of Thr168. A catalytic mechanism very similar to that proposed for L-serine dehydratase is discussed.
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http://dx.doi.org/10.1016/j.bbapap.2011.10.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7380123PMC
March 2012

Structural flexibility of the macrophage dengue virus receptor CLEC5A: implications for ligand binding and signaling.

J Biol Chem 2011 Jul 12;286(27):24208-18. Epub 2011 May 12.

Henry Wellcome Building for Molecular Physiology, University of Oxford, Roosevelt Drive, Oxford OX3 7BN, United Kingdom.

The human C-type lectin-like molecule CLEC5A is a critical macrophage receptor for dengue virus. The binding of dengue virus to CLEC5A triggers signaling through the associated adapter molecule DAP12, stimulating proinflammatory cytokine release. We have crystallized an informative ensemble of CLEC5A structural conformers at 1.9-Å resolution and demonstrate how an on-off extension to a β-sheet acts as a binary switch regulating the flexibility of the molecule. This structural information together with molecular dynamics simulations suggests a mechanism whereby extracellular events may be transmitted through the membrane and influence DAP12 signaling. We demonstrate that CLEC5A is homodimeric at the cell surface and binds to dengue virus serotypes 1-4. We used blotting experiments, surface analyses, glycan microarray, and docking studies to investigate the ligand binding potential of CLEC5A with particular respect to dengue virus. This study provides a rational foundation for understanding the dengue virus-macrophage interaction and the role of CLEC5A in dengue virus-induced lethal disease.
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http://dx.doi.org/10.1074/jbc.M111.226142DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3129202PMC
July 2011

REFMAC5 for the refinement of macromolecular crystal structures.

Acta Crystallogr D Biol Crystallogr 2011 Apr 18;67(Pt 4):355-67. Epub 2011 Mar 18.

Structural Biology Laboratory, Department of Chemistry, University of York, Heslington, York, England.

This paper describes various components of the macromolecular crystallographic refinement program REFMAC5, which is distributed as part of the CCP4 suite. REFMAC5 utilizes different likelihood functions depending on the diffraction data employed (amplitudes or intensities), the presence of twinning and the availability of SAD/SIRAS experimental diffraction data. To ensure chemical and structural integrity of the refined model, REFMAC5 offers several classes of restraints and choices of model parameterization. Reliable models at resolutions at least as low as 4 Å can be achieved thanks to low-resolution refinement tools such as secondary-structure restraints, restraints to known homologous structures, automatic global and local NCS restraints, `jelly-body' restraints and the use of novel long-range restraints on atomic displacement parameters (ADPs) based on the Kullback-Leibler divergence. REFMAC5 additionally offers TLS parameterization and, when high-resolution data are available, fast refinement of anisotropic ADPs. Refinement in the presence of twinning is performed in a fully automated fashion. REFMAC5 is a flexible and highly optimized refinement package that is ideally suited for refinement across the entire resolution spectrum encountered in macromolecular crystallography.
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http://dx.doi.org/10.1107/S0907444911001314DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3069751PMC
April 2011

Evaluating the solution from MrBUMP and BALBES.

Acta Crystallogr D Biol Crystallogr 2011 Apr 18;67(Pt 4):313-23. Epub 2011 Mar 18.

STFC Rutherford Appleton Laboratory, Chilton, England.

Molecular replacement is one of the key methods used to solve the problem of determining the phases of structure factors in protein structure solution from X-ray image diffraction data. Its success rate has been steadily improving with the development of improved software methods and the increasing number of structures available in the PDB for use as search models. Despite this, in cases where there is low sequence identity between the target-structure sequence and that of its set of possible homologues it can be a difficult and time-consuming chore to isolate and prepare the best search model for molecular replacement. MrBUMP and BALBES are two recent developments from CCP4 that have been designed to automate and speed up the process of determining and preparing the best search models and putting them through molecular replacement. Their intention is to provide the user with a broad set of results using many search models and to highlight the best of these for further processing. An overview of both programs is presented along with a description of how best to use them, citing case studies and the results of large-scale testing of the software.
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http://dx.doi.org/10.1107/S0907444911007530DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3069746PMC
April 2011

BALBES: a molecular-replacement pipeline.

Acta Crystallogr D Biol Crystallogr 2008 Jan 5;64(Pt 1):125-32. Epub 2007 Dec 5.

York Structural Biology Laboratory, Chemistry Department, University of York, Heslington, York, England.

The number of macromolecular structures solved and deposited in the Protein Data Bank (PDB) is higher than 40 000. Using this information in macromolecular crystallography (MX) should in principle increase the efficiency of MX structure solution. This paper describes a molecular-replacement pipeline, BALBES, that makes extensive use of this repository. It uses a reorganized database taken from the PDB with multimeric as well as domain organization. A system manager written in Python controls the workflow of the process. Testing the current version of the pipeline using entries from the PDB has shown that this approach has huge potential and that around 75% of structures can be solved automatically without user intervention.
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http://dx.doi.org/10.1107/S0907444907050172DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2394813PMC
January 2008

Model preparation in MOLREP and examples of model improvement using X-ray data.

Acta Crystallogr D Biol Crystallogr 2008 Jan 5;64(Pt 1):33-9. Epub 2007 Dec 5.

Structural Biology Laboratory, Department of Chemistry, University of York, Heslington, York YO10 5YW, England.

The success of molecular replacement is critically dependent on the quality of the search model. Several model-preparation procedures are integrated in the molecular-replacement program MOLREP. These include model modification on the basis of amino-acid sequence alignment and model correction based on analysis of the solvent-accessibility of the atoms. The packing function used in MOLREP for the translational search is explained in the context of model preparation. In difficult cases, bioinformatics-based modifications are not sufficient for successful molecular replacement. An approach implemented in MOLREP for solving cases with translational noncrystallographic symmetry is an example of model preparation in which analysis of X-ray data plays an essential role. In addition, two examples are presented in which the X-ray data were used to refine partial models for subsequent use in molecular replacement.
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http://dx.doi.org/10.1107/S0907444907049839DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2394799PMC
January 2008

Structural framework for DNA translocation via the viral portal protein.

EMBO J 2007 Apr 15;26(7):1984-94. Epub 2007 Mar 15.

York Structural Biology Laboratory, Department of Chemistry, University of York, York, UK.

Tailed bacteriophages and herpesviruses load their capsids with DNA through a tunnel formed by the portal protein assembly. Here we describe the X-ray structure of the bacteriophage SPP1 portal protein in its isolated 13-subunit form and the pseudoatomic structure of a 12-subunit assembly. The first defines the DNA-interacting segments (tunnel loops) that pack tightly against each other forming the most constricted part of the tunnel; the second shows that the functional dodecameric state must induce variability in the loop positions. Structural observations together with geometrical constraints dictate that in the portal-DNA complex, the loops form an undulating belt that fits and tightly embraces the helical DNA, suggesting that DNA translocation is accompanied by a 'mexican wave' of positional and conformational changes propagating sequentially along this belt.
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http://dx.doi.org/10.1038/sj.emboj.7601643DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1847669PMC
April 2007

Intensity statistics in twinned crystals with examples from the PDB.

Acta Crystallogr D Biol Crystallogr 2006 Jan 14;62(Pt 1):83-95. Epub 2005 Dec 14.

Structural Biology Laboratory, Department of Chemistry, University of York, Heslington, York YO10 5YW, England.

Entries deposited in the Protein Data Bank as of February 2004 for which both model and X-ray data were available were analysed to identify cases of twinning using such simple statistics as the R factor between potential twin-related reflections. Careful consideration of all identified twins showed that in many cases twinning was ignored during structure solution and refinement. Manual analysis of the models showed that twinning often occurs in association with rotational pseudosymmetry parallel to the twinning operator. The coexistence of these two phenomena complicates the detection and diagnostics of twinning using currently available twinning tests. It was concluded that a twinning-detection step should be incorporated in every stage of structure analysis from data acquisition to refinement and validation.
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http://dx.doi.org/10.1107/S0907444905036759DOI Listing
January 2006

The structure of the oligopeptide-binding protein, AppA, from Bacillus subtilis in complex with a nonapeptide.

J Mol Biol 2005 Jan;345(4):879-92

Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5YW, UK.

Besides their role as a source of amino acids for Bacillus subtilis, exogenous peptides play important roles in the signalling pathways leading to the development of competence and sporulation. B.subtilis has three peptide transport systems all belonging to the ATP-binding cassette family, a dipeptide permease (Dpp) and two oligopeptide permeases (Opp and App) with overlapping specificity. These comprise a membrane-spanning channel through which the peptide passes, a pair of ATPases which couple ATP hydrolysis to peptide translocation and a lipid-modified, membrane-anchored extracellular "binding-protein" that serves as the receptor for the system. Here, we present the crystal structure of a soluble form of the peptide-binding protein AppA, which has been solved to 1.6 A spacing by anomalous scattering and molecular replacement methods. The structure reveals a protein made of two distinct lobes with a topology similar to those of DppA from Escherichia coli and OppA from Salmonella typhimurium. Examination of the interlobe region reveals an enlarged pocket, containing electron density defining a nonapeptide ligand. The main-chain of the peptide is well defined and makes a series of polar contacts with the protein including salt-bridges at both its termini. The side-chain density is ambiguous in places, consistent with the interpretation that a population of peptides is bound, whose average electron density resembles the amino acid sequence N-VDSKNTSSW-C.
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http://dx.doi.org/10.1016/j.jmb.2004.10.089DOI Listing
January 2005

REFMAC5 dictionary: organization of prior chemical knowledge and guidelines for its use.

Acta Crystallogr D Biol Crystallogr 2004 Dec 26;60(Pt 12 Pt 1):2184-95. Epub 2004 Nov 26.

Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5YW, England.

One of the most important aspects of macromolecular structure refinement is the use of prior chemical knowledge. Bond lengths, bond angles and other chemical properties are used in restrained refinement as subsidiary conditions. This contribution describes the organization and some aspects of the use of the flexible and human/machine-readable dictionary of prior chemical knowledge used by the maximum-likelihood macromolecular-refinement program REFMAC5. The dictionary stores information about monomers which represent the constitutive building blocks of biological macromolecules (amino acids, nucleic acids and saccharides) and about numerous organic/inorganic compounds commonly found in macromolecular crystallography. It also describes the modifications the building blocks undergo as a result of chemical reactions and the links required for polymer formation. More than 2000 monomer entries, 100 modification entries and 200 link entries are currently available. Algorithms and tools for updating and adding new entries to the dictionary have also been developed and are presented here. In many cases, the REFMAC5 dictionary allows entirely automatic generation of restraints within REFMAC5 refinement runs.
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http://dx.doi.org/10.1107/S0907444904023510DOI Listing
December 2004

Crystallization and preliminary X-ray diffraction studies of a fungal hydrolase from Ophiostoma novo-ulmi.

Acta Crystallogr D Biol Crystallogr 2004 Oct 23;60(Pt 10):1879-82. Epub 2004 Sep 23.

Henry Wellcome Building for Biocatalysis, School of Biological and Chemical Sciences, University of Exeter, Exeter EX4 4QD, England.

Dutch elm disease fungus Ophiostoma novo-ulmi contains a hydrolase activity which catalyses the resolution of racemic ethyl naproxen to the corresponding acid. The recombinant enzyme has been crystallized by the vapour-diffusion method in two crystal forms. The crystals of the first form belong to space group P2(1)2(1)2, with unit-cell parameters a = 115.9, b = 174.4, c = 62.1 A. The enzyme also crystallizes in space group P2(1)2(1)2, with unit-cell parameters a = 72.9, b = 212.7, c = 61.7 A. Synchrotron data have been collected for both crystal forms to 2.6 and 2.3 A, respectively. A molecular-replacement solution has been found using a remote starting model of a bacterial esterase (23% sequence identity) for both crystal forms. Multicrystal averaging has resulted in interpretable electron-density maps.
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http://dx.doi.org/10.1107/S0907444904018153DOI Listing
October 2004

The crystal structure of a complex of Campylobacter jejuni dUTPase with substrate analogue sheds light on the mechanism and suggests the "basic module" for dimeric d(C/U)TPases.

J Mol Biol 2004 Oct;342(5):1583-97

Structural Biology Laboratory, Department of Chemistry, University of York, Heslington, York YO10 5YW, UK.

The crystal structure of the dUTPase from the important gastric pathogen Campylobacter jejuni has been solved at 1.65 A spacing. This essential bacterial enzyme is the second representative of the new family of dimeric dUTPases to be structurally characterised. Members of this family have a novel all-alpha fold and are unrelated to the all-beta dUTPases of the majority of organisms including eukaryotes such as humans, bacteria such as Escherichia coli, archaea like Methanococcus jannaschii and animal viruses. Therefore, dimeric dUTPases can be considered as candidate drug targets. The X-ray structure of the C.jejuni dUTPase in complex with the non-hydrolysable substrate analogue dUpNHp allows us to define the positions of three catalytically significant phosphate-binding magnesium ions and provides a starting point for a detailed understanding of the mechanism of dUTP/dUDP hydrolysis by dimeric dUTPases. Indeed, a water molecule present in the structure is ideally situated to act as the attacking nucleophile during hydrolysis. A comparison of the dUTPases from C.jejuni and Trypanosoma cruzi reveals a common fold with certain distinct features, both in the rigid and mobile domains as defined in the T.cruzi structure. Homologues of the C.jejuni dUTPase have been identified in several other bacteria and bacteriophages, including the dCTPase of phage T4. Sequence comparisons of these proteins define a new superfamily of d(C/U)TPases that includes three distinct enzyme families: (1) dUTPases in trypanosomatides, C.jejuni and several other Gram-negative bacteria, (2) predicted dUTPases in various Gram-positive bacteria and their phages, and (3) dCTP/dUTPases in enterobacterial T4-like phages. All these enzymes share a basic module that consists of two alpha-helices from the rigid domain, two helices from the mobile domain and connecting loops. These results in concert with a number of conserved residues responsible for interdomain cross-talk provide valuable insight towards rational drug design.
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http://dx.doi.org/10.1016/j.jmb.2004.07.050DOI Listing
October 2004