Crystal structure of Arabidopsis thaliana peroxiredoxin A C119S mutant
This article reports the crystal structure of a PrxA C119S mutant refined to 2.6   Å resolution. The protein exists as a decamer both in the crystal structure and in solution. The structure is in the reduced state suitable for the approach of peroxide, though conformational changes are needed for the resolving process. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - September 19, 2018 Category: Biochemistry Authors: Yang, Y. Cai, W. Wang, J. Pan, W. Liu, L. Wang, M. Zhang, M. Tags: peroxiredoxins peroxiredoxin A Arabidopsis thaliana redox regulation chloroplast thioredoxin systems oxidative and photo-oxidative stress research communications Source Type: research

Structure of glyoxysomal malate dehydrogenase (MDH3) from Saccharomyces cerevisiae
Malate dehydrogenase (MDH), a carbohydrate and energy metabolism enzyme in eukaryotes, catalyzes the interconversion of malate to oxaloacetate (OAA) in conjunction with that of nicotinamide adenine dinucleotide (NAD+) to NADH. Three isozymes of MDH have been reported in Saccharomyces cerevisiae: MDH1, MDH2 and MDH3. MDH1 is a mitochondrial enzyme and a member of the tricarboxylic acid cycle, whereas MDH2 is a cytosolic enzyme that functions in the glyoxylate cycle. MDH3 is a glyoxysomal enzyme that is involved in the reoxidation of NADH, which is produced during fatty-acid β -oxidation. The affinity of MDH3 for OAA is...
Source: Acta Crystallographica Section F - September 19, 2018 Category: Biochemistry Authors: Moriyama, S. Nishio, K. Mizushima, T. Tags: malate dehydrogenase glyoxysome fatty-acid β -oxidation X-ray crystallography MDH3 Saccharomyces cerevisiae research communications Source Type: research

Conformational changes on substrate binding revealed by structures of Methylobacterium extorquens malate dehydrogenase
Three high-resolution X-ray crystal structures of malate dehydrogenase (MDH; EC 1.1.1.37) from the methylotroph Methylobacterium extorquens AM1 are presented. By comparing the structures of apo MDH, a binary complex of MDH and NAD+, and a ternary complex of MDH and oxaloacetate with ADP-ribose occupying the pyridine nucleotide-binding site, conformational changes associated with the formation of the catalytic complex were characterized. While the substrate-binding site is accessible in the enzyme resting state or NAD+-bound forms, the substrate-bound form exhibits a closed conformation. This conformational change involves ...
Source: Acta Crystallographica Section F - September 19, 2018 Category: Biochemistry Authors: Gonz á lez, J.M. Marti-Arbona, R. Chen, J.C.-H. Broom-Peltz, B. Unkefer, C.J. Tags: malate dehydrogenase methylotrophs biofuels Methylobacterium extorquens research communications Source Type: research

Investigation into the binding of dyes within protein crystals
It was found that the crystals of at least a dozen different proteins could be thoroughly stained to an intense color with a panel of dyes. Many, if not most, of the stained protein crystals retained the dyes almost indefinitely when placed in large volumes of dye-free mother liquor. Dialysis experiments showed that most of the dyes that were retained in crystals also bound to the protein when free in solution; less frequently, some dyes bound only in the crystal. The experiments indicated a strong association of the dyes with the proteins. Four protein crystals were investigated by X-ray diffraction to ascertain the mode ...
Source: Acta Crystallographica Section F - September 3, 2018 Category: Biochemistry Authors: McPherson, A. Larson, S.B. Tags: dyes stains disordered binding detergents difference Fourier complexes colored crystals staining protein crystals research communications Source Type: research

X-ray structure of alisporivir in complex with cyclophilin A at 1.5   Å resolution
This study provides detailed molecular insights into the CypA – ALV interaction. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - September 3, 2018 Category: Biochemistry Authors: Dujardin, M. Bouckaert, J. Rucktooa, P. Hanoulle, X. Tags: cyclophilins peptidylprolyl isomerase alisporivir inhibitors isomerases non-immunosuppressive CsA derivative HIV HCV Duchenne muscular dystrophy research communications Source Type: research

TssA from Aeromonas hydrophila: expression, purification and crystallographic studies
TssA is a core subunit of the type VI secretion system, which is a major player in interspecies competition in Gram-negative bacteria. Previous studies on enteroaggregative Escherichia coli TssA suggested that it is comprised of three putative domains: a conserved N-terminal domain, a middle domain and a ring-forming C-terminal domain. X-ray studies of the latter two domains have identified their respective structures. Here, the results of the expression and purification of full-length and domain constructs of TssA from Aeromonas hydrophila are reported, resulting in diffraction-quality crystals for the middle domain (Nt2)...
Source: Acta Crystallographica Section F - September 3, 2018 Category: Biochemistry Authors: Dix, S.R. Sun, R. Harris, M.J. Batters, S.L. Sedelnikova, S.E. Baker, P.J. Thomas, M.S. Rice, D.W. Tags: type VI secretion system TssA subunit Aeromonas hydrophila research communications Source Type: research

Mosquito-larvicidal binary toxin receptor protein (Cqm1): crystallization and X-ray crystallographic analysis
Cqm1 from Culex quinquefasciatus has been identified as the receptor for Lysinibacillus sphaericus binary toxin (BinAB). It is an amylomaltase that is presented on the epithelial membrane in the larval midgut through a glycosyl-phosphatidylinositol anchor. The active core of this protein (residues 23 – 560) was overexpressed in Escherichia coli, purified and successfully crystallized by the sitting-drop vapor-diffusion method using d-arabinose and CaCl2 as additives, as identified using high-throughput differential scanning fluorimetry analysis. X-ray diffraction data were collected to a resolution of 2.8   Å...
Source: Acta Crystallographica Section F - September 3, 2018 Category: Biochemistry Authors: Sharma, M. Lakshmi, A. Gupta, G.D. Kumar, V. Tags: Cqm1 Culex quinquefasciatus binary toxin receptor protein Lysinibacillus sphaericus amylomaltase mosquito-larvicidal binary toxin crystallization research communications Source Type: research

Staphylococcus aureus lipase: purification, kinetic characterization, crystallization and crystallographic study
Staphylococcus aureus lipase (SAL), a triacylglycerol esterase, is an important virulence factor in S. aureus and may be a therapeutic target for infectious diseases caused by S. aureus. For the purposes of anti-SAL drug development using structure-based drug design, X-ray crystallographic analysis of SAL overexpressed in Escherichia coli was performed. The recombinant protein was purified using a three-step protocol involving immobilized metal-affinity chromatography, cation-exchange chromatography and anion-exchange chromatography flowthrough methods, yielding 40   mg of protein per litre of bacterial culture. Crystals...
Source: Acta Crystallographica Section F - September 3, 2018 Category: Biochemistry Authors: Tanaka, M. Kamitani, S. Kitadokoro, K. Tags: lipase Staphylococcus aureus virulence factors toxins crystallization research communications Source Type: research

Serendipitous crystallization and structure determination of bacterioferritin from Achromobacter
Bacterioferritins (Bfrs) are ferritin-like molecules with a hollow spherical 24-mer complex design that are unique to bacterial and archaeal species. They play a critical role in storing iron(III) within the complex at concentrations much higher than the feasible solubility limits of iron(III), thus maintaining iron homeostasis within cells. Here, the crystal structure of bacterioferritin from Achromobacter (Ach Bfr) that crystallized serendipitously during a crystallization attempt of an unrelated mycobacterial protein is reported at 1.95   Å resolution. Notably, Fe atoms were bound to the structure along with a p...
Source: Acta Crystallographica Section F - August 29, 2018 Category: Biochemistry Authors: Dwivedy, A. Jha, B. Singh, K.H. Ahmad, M. Ashraf, A. Kumar, D. Biswal, B.K. Tags: bacterioferritin Achromobacter dinuclear center iron storage heme research communications Source Type: research

MBP-binding DARPins facilitate the crystallization of an MBP fusion protein
The production of high-quality crystals is the main bottleneck in determining the structures of proteins using X-ray crystallography. In addition to being recognized as a very effective solubility-enhancing fusion partner, Escherichia coli maltose-binding protein (MBP) has also been successfully employed as a `fixed-arm' crystallization chaperone in more than 100 cases. Here, it is reported that designed ankyrin-repeat proteins (DARPins) that bind with high affinity to MBP can promote the crystallization of an MBP fusion protein when the fusion protein alone fails to produce diffraction-quality crystals. As a proof of prin...
Source: Acta Crystallographica Section F - August 29, 2018 Category: Biochemistry Authors: Gumpena, R. Lountos, G.T. Waugh, D.S. Tags: crystallization chaperones designed ankyrin-repeat proteins dual-specificity phosphatase 1 maltose-binding protein surface-entropy-reduction mutagenesis research communications Source Type: research

MSMEG_6292, a Mycobacterium smegmatis RNA polymerase secondary channel-binding protein: purification, crystallization and X-ray diffraction analysis
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 th...
Source: Acta Crystallographica Section F - August 29, 2018 Category: Biochemistry Authors: Joseph, A. Nagaraja, V. Natesh, R. Tags: transcription regulation RNA polymerase Gre-factor homologue mycobacteria X-ray diffraction Mycobacterium smegmatis MSMEG_6292 research communications Source Type: research

TssA from Burkholderia cenocepacia: expression, purification, crystallization and crystallographic analysis
TssA is a core component of the type VI secretion system, and phylogenetic analysis of TssA subunits from different species has suggested that these proteins fall into three distinct clades. Whilst representatives of two clades, TssA1 and TssA2, have been the subjects of investigation, no members of the third clade (TssA3) have been studied. Constructs of TssA from Burkholderia cenocepacia, a representative of clade 3, were expressed, purified and subjected to crystallization trials. Data were collected from crystals of constructs of the N-terminal and C-terminal domains. Analysis of the data from the crystals of these con...
Source: Acta Crystallographica Section F - August 29, 2018 Category: Biochemistry Authors: Owen, H.J. Sun, R. Ahmad, A. Sedelnikova, S.E. Baker, P.J. Thomas, M.S. Rice, D.W. Tags: type VI secretion system TssA Burkholderia cenocepacia research communications Source Type: research

Ab initio structure solution of a proteolytic fragment using ARCIMBOLDO
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  ...
Source: Acta Crystallographica Section F - August 29, 2018 Category: Biochemistry Authors: Abendroth, J. Sankaran, B. Myler, P.J. Lorimer, D.D. Edwards, T.E. Tags: proteolytic fragments ARCIMBOLDO ab initio structure determination structural genomics infectious diseases Mycobacterium smegmatis research communications Source Type: research

Structure of the Fc fragment of the NIST reference antibody RM8671
In this report, the structure of the Fc fragment of the NIST IgG1 mAb (reference material 8671) is described at 2.1   Å resolution in space group P212121, with approximate unit-cell parameters a   = 50, b = 80, c = 138   Å . Prior Fc structures with a wide variety of modifications are also surveyed, focusing on those in the same crystal form. To facilitate the analysis of conformations, a reference frame and a two-parameter metric are proposed, considering the CH2 domains as mobile with respect to a fixed dimeric CH3 core. Over several human Fc structures, a significant variation in Fc elbow conformati...
Source: Acta Crystallographica Section F - August 29, 2018 Category: Biochemistry Authors: Gallagher, D.T. Galvin, C.V. Karageorgos, I. Tags: antibodies NIST reference antibody RM8671 conformation crystal structure Fc human immune X-ray research communications Source Type: research

Structure of the GH9 glucosidase/glucosaminidase from Vibrio cholerae
Glycoside hydrolase family 9 (GH9) of carbohydrate-processing enzymes primarily consists of inverting endoglucanases. A subgroup of GH9 enzymes are believed to act as exo-glucosidases or exo-glucosaminidases, with many being found in organisms of the family Vibrionaceae, where they are proposed to function within the chitin-catabolism pathway. Here, it is shown that the GH9 enzyme from the pathogen Vibrio cholerae (hereafter referred to as VC0615) is active on both chitosan-derived and β -glucoside substrates. The structure of VC0615 at 3.17   Å resolution is reported from a crystal form with poor diffraction ...
Source: Acta Crystallographica Section F - August 6, 2018 Category: Biochemistry Authors: Wu, L. Davies, G.J. Tags: glycoside hydrolase enzymes carbohydrates VC0615 glucosidase glucosaminidases Vibrio hydrolases research communications Source Type: research

Crystal structure of the effector-binding domain of Synechococcus elongatus CmpR in complex with ribulose 1,5-bisphosphate
The CO2-concentrating mechanism (CCM) has evolved to improve the efficiency of photosynthesis in autotrophic cyanobacteria. CmpR, a LysR-type transcriptional regulator (LTTR) from Synechococcus elongatus PCC 7942, was found to regulate CCM-related genes under low-CO2 conditions. Here, the dimeric structure of the effector-binding domain of CmpR (CmpR-EBD) in complex with the co-activator ribulose 1,5-bisphosphate (RuBP) is reported at 2.15   Å resolution. One RuBP molecule binds to the inter-domain cleft between the two subunits of the CmpR-EBD dimer. Structural comparison combined with sequence analyses demonstrat...
Source: Acta Crystallographica Section F - August 1, 2018 Category: Biochemistry Authors: Mahounga, D.M. Sun, H. Jiang, Y.-L. Tags: crystal structure LysR-type transcription factors cyanobacteria Synechococcus elongatus PCC 7942 CmpR-EBD complex with ribulose 1,5-bisphosphate CO2-concentrating mechanism research communications Source Type: research

Structural studies of the unusual metal-ion site of the GH124 endoglucanase from Ruminiclostridium thermocellum
The recent discovery of `lytic' polysaccharide monooxygenases, copper-dependent enzymes for biomass degradation, has provided new impetus for the analysis of unusual metal-ion sites in carbohydrate-active enzymes. In this context, the CAZY family GH124 endoglucanase from Ruminiclostridium thermocellum contains an unusual metal-ion site, which was originally modelled as a Ca2+ site but features aspartic acid, asparagine and two histidine imidazoles as coordinating residues, which are more consistent with a transition-metal binding environment. It was sought to analyse whether the GH124 metal-ion site might accommodate other...
Source: Acta Crystallographica Section F - August 1, 2018 Category: Biochemistry Authors: Urresti, S. Cartmell, A. Liu, F. Walton, P.H. Davies, G.J. Tags: endoglucanase Ruminiclostridium thermocellum metal ion oligosaccharides enzymes carbohydrates bio-inorganic 2-oxohistidine research communications Source Type: research

Carbohydrate structure hits the groove
(Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - August 1, 2018 Category: Biochemistry Authors: Agirre, J. van Raaij, M.J. Tags: special issues carbohydrates glycoproteins – carbohydrate complexes research communications Source Type: research

Structure of a Talaromyces pinophilus GH62 arabinofuranosidase in complex with AraDNJ at 1.25   Å resolution
The enzymatic hydrolysis of complex plant biomass is a major societal goal of the 21st century in order to deliver renewable energy from nonpetroleum and nonfood sources. One of the major problems in many industrial processes, including the production of second-generation biofuels from lignocellulose, is the presence of `hemicelluloses' such as xylans which block access to the cellulosic biomass. Xylans, with a polymeric β -1,4-xylose backbone, are frequently decorated with acetyl, glucuronyl and arabinofuranosyl `side-chain' substituents, all of which need to be removed for complete degradation of the xylan. As such,...
Source: Acta Crystallographica Section F - July 26, 2018 Category: Biochemistry Authors: Moroz, O.V. Sobala, L.F. Blagova, E. Coyle, T. Peng, W. M ø rkeberg Krogh, K.B.R. Stubbs, K.A. Wilson, K.S. Davies, G.J. Tags: biofuels glycosidases enzymes enzyme inhibitors Talaromyces pinophilus arabinofuranosidase research communications Source Type: research

Crystal structure of highly glycosylated human leukocyte elastase in complex with an S2 ′ site binding inhibitor
Glycosylated human leukocyte elastase (HLE) was crystallized and structurally analysed in complex with a 1,3-thiazolidine-2,4-dione derivative that had been identified as an HLE inhibitor in preliminary studies. In contrast to previously described HLE structures with small-molecule inhibitors, in this structure the inhibitor does not bind to the S1 and S2 substrate-recognition sites; rather, this is the first HLE structure with a synthetic inhibitor in which the S2 ′ site is blocked that normally binds the second side chain at the C-terminal side of the scissile peptide bond in a substrate protein. The inhibitor also...
Source: Acta Crystallographica Section F - July 26, 2018 Category: Biochemistry Authors: Hochscherf, J. Pietsch, M. Tieu, W. Kuan, K. Abell, A.D. G ü tschow, M. Niefind, K. Tags: human leukocyte elastase human neutrophil elastase hydrolases N-glycosylation S2 ′ site research communications Source Type: research

Conformations of the type-1 lacto-N-biose I unit in protein complex structures
The lacto-N-biose I (Gal β 1 – 3GlcNAc; LNB) disaccharide is present as a core unit of type-1 blood group antigens of animal glycoconjugates and milk oligosaccharides. Type-1 antigens often serve as cell-surface receptors for infection by pathogens. LNB in human milk oligosaccharides functions as a prebiotic for bifidobacteria and plays a key role in the symbiotic relationship of commensal gut microbes in infants. Protein Data Bank (PDB) entries exhibiting the LNB unit were investigated using the GlycoMapsDB web tool. There are currently 159 β -LNB and nine α -LNB moieties represented in ligands in th...
Source: Acta Crystallographica Section F - July 26, 2018 Category: Biochemistry Authors: Fushinobu, S. Tags: human milk oligosaccharides Lewis blood group antigens glycosphingolipids LS-tetrasaccharides type-1 ABO(H) blood group antigens research communications Source Type: research

Making glycoproteins a little bit sweeter with PDB-REDO
Glycosylation is one of the most common forms of protein post-translational modification, but is also the most complex. Dealing with glycoproteins in structure model building, refinement, validation and PDB deposition is more error-prone than dealing with nonglycosylated proteins owing to limitations of the experimental data and available software tools. Also, experimentalists are typically less experienced in dealing with carbohydrate residues than with amino-acid residues. The results of the reannotation and re-refinement by PDB-REDO of 8114 glycoprotein structure models from the Protein Data Bank are analyzed. The posit...
Source: Acta Crystallographica Section F - July 26, 2018 Category: Biochemistry Authors: van Beusekom, B. L ü tteke, T. Joosten, R.P. Tags: glycoproteins PDB-REDO pdb-care validation carbohydrates research communications Source Type: research

Spin ballet for sweet encounters: saturation-transfer difference NMR and X-ray crystallography complement each other in the elucidation of protein – glycan interactions
Biomolecular NMR spectroscopy has limitations in the determination of protein structures: an inherent size limit and the requirement for expensive and potentially difficult isotope labelling pose considerable hurdles. Therefore, structural analysis of larger proteins is almost exclusively performed by crystallography. However, the diversity of biological NMR applications outperforms that of any other structural biology technique. For the characterization of transient complexes formed by proteins and small ligands, notably oligosaccharides, one NMR technique has recently proven to be particularly powerful: saturation-transf...
Source: Acta Crystallographica Section F - July 26, 2018 Category: Biochemistry Authors: Blaum, B.S. Neu, U. Peters, T. Stehle, T. Tags: saturation-transfer difference NMR STD-NMR polyomavirus carbohydrates lectins structural biology topical reviews Source Type: research

A perspective on structural and mechanistic aspects of protein O-fucosylation
Protein O-fucosylation is an important post-translational modification (PTM) found in cysteine-rich repeats in proteins. Protein O-fucosyltransferases 1 and 2 (PoFUT1 and PoFUT2) are the enzymes responsible for this PTM and selectively glycosylate specific residues in epidermal growth factor-like (EGF) repeats and thrombospondin type I repeats (TSRs), respectively. Within the past six years, crystal structures of both enzymes have been reported, revealing important information on how they recognize protein substrates and achieve catalysis. Here, the structural information available today is summarized and how PoFUT1 and Po...
Source: Acta Crystallographica Section F - July 26, 2018 Category: Biochemistry Authors: Lira-Navarrete, E. Hurtado-Guerrero, R. Tags: O-fucosylation protein O-fucosyltransferases epidermal growth factor-like repeats EGF repeats thrombospondin type I repeats TSRs enzyme mechanisms GDP-fucose topical reviews Source Type: research

Crystal structures and kinetic analyses of N-acetylmannosamine-6-phosphate 2-epimerases from Fusobacterium nucleatum and Vibrio cholerae
Sialic acids are nine-carbon sugars that are found abundantly on the cell surfaces of mammals as glycoprotein or glycolipid complexes. Several Gram-negative and Gram-positive bacteria have the ability to scavenge and catabolize sialic acids to use as a carbon source. This gives them an advantage in colonizing sialic acid-rich environments. The genes of the sialic acid catabolic pathway are generally present as the operon nanAKE. The third gene in the operon encodes the enzyme N-acetylmannosamine-6-phosphate 2-epimerase (NanE), which catalyzes the conversion of N-acetylmannosamine 6-phosphate to N-acetylglucosamine 6-phosph...
Source: Acta Crystallographica Section F - June 28, 2018 Category: Biochemistry Authors: Manjunath, L. Guntupalli, S.R. Currie, M.J. North, R.A. Dobson, R.C.J. Nayak, V. Subramanian, R. Tags: sialic acid catabolism N-acetylmannosamine-6-phosphate 2-epimerase Fusobacterium nucleatum Vibrio cholerae research communications Source Type: research

Cinder: keeping crystallographers app-y
The process of producing suitable crystals for X-ray diffraction analysis most often involves the setting up of hundreds (or thousands) of individual crystallization trials, each of which must be repeatedly examined for crystals or hints of crystallinity. Currently, the only real way to address this bottleneck is to use an automated imager to capture images of the trials. However, the images still need to be assessed for crystals or other outcomes. Ideally, there would exist some rapid and reliable machine-analysis tool to translate the images into a quantitative result. However, as yet no such tool exists in wide usage, d...
Source: Acta Crystallographica Section F - June 26, 2018 Category: Biochemistry Authors: Rosa, N. Ristic, M. Marshall, B. Newman, J. Tags: crystallization images scoring machine learning Cinder mobile apps research communications Source Type: research

Crystallization and X-ray analysis of all of the players in the autoregulation of the ataRT toxin – antitoxin system
The ataRT operon from enteropathogenic Escherichia coli encodes a toxin – antitoxin (TA) module with a recently discovered novel toxin activity. This new type II TA module targets translation initiation for cell-growth arrest. Virtually nothing is known regarding the molecular mechanisms of neutralization, toxin catalytic action or translation autoregulation. Here, the production, biochemical analysis and crystallization of the intrinsically disordered antitoxin AtaR, the toxin AtaT, the AtaR – AtaT complex and the complex of AtaR – AtaT with a double-stranded DNA fragment of the operator region of the pr...
Source: Acta Crystallographica Section F - June 26, 2018 Category: Biochemistry Authors: Jur ė nas, D. Van Melderen, L. Garcia-Pino, A. Tags: toxin – antitoxin acetyltransferase protein DNA complexes AtaR AtaT Escherichia coli research communications Source Type: research

Crystal structure of Escherichia coli purine nucleoside phosphorylase complexed with acyclovir
Escherichia coli purine nucleoside phosphorylase (PNP), which catalyzes the reversible phosphorolysis of purine ribonucleosides, belongs to the family I hexameric PNPs. Owing to their key role in the purine salvage pathway, PNPs are attractive targets for drug design against some pathogens. Acyclovir (ACV) is an acyclic derivative of the PNP substrate guanosine and is used as an antiviral drug for the treatment of some human viral infections. The crystalline complex of E. coli PNP with acyclovir was prepared by co-crystallization in microgravity using counter-diffusion through a gel layer in a capillary. The structure of t...
Source: Acta Crystallographica Section F - June 26, 2018 Category: Biochemistry Authors: Timofeev, V.I. Zhukhlistova, N.E. Abramchik, Y.A. Muravieva, T.I. Esipov, R.S. Kuranova, I.P. Tags: purine nucleoside phosphorylase crystal structure acyclovir inhibitors Escherichia coli structure-based drug design tumour-directed gene therapy research communications Source Type: research

Crystal structure of human anterior gradient protein 3
Oxidative protein folding in the endoplasmic reticulum is catalyzed by the protein disulfide isomerase family of proteins. Of the 20 recognized human family members, the structures of eight have been deposited in the PDB along with domains from six more. Three members of this family, ERp18, anterior gradient protein 2 (AGR2) and anterior gradient protein 3 (AGR3), are single-domain proteins which share sequence similarity. While ERp18 has a canonical active-site motif and is involved in native disulfide-bond formation, AGR2 and AGR3 lack elements of the active-site motif found in other family members and may both interact ...
Source: Acta Crystallographica Section F - June 26, 2018 Category: Biochemistry Authors: Nguyen, V.D. Biterova, E. Salin, M. Wierenga, R.K. Ruddock, L.W. Tags: thioredoxin fold protein disulfide isomerase endoplasmic reticulum isomerases anterior gradient protein 3 research communications Source Type: research

A novel bacterial class V dye-decolourizing peroxidase from the extremophile Deinococcus radiodurans: cloning, expression optimization, purification, crystallization, initial characterization and X-ray diffraction analysis
Deinococcus radiodurans is a bacterium with extreme resistance to desiccation and radiation. The resistance mechanism is unknown, but an efficient reactive oxygen species (ROS) scavenging system and DNA-repair and DNA-protection mechanisms are believed to play important roles. Here, the cloning and small- and medium-scale expression tests of a novel dye-decolourizing peroxidase from D. radiodurans (DrDyP) using three different Escherichia coli strains and three different temperatures in order to identify the optimum conditions for the expression of recombinant DrDyP are presented. The best expression conditions were used f...
Source: Acta Crystallographica Section F - June 26, 2018 Category: Biochemistry Authors: Frade, K.S.T. Fernandes, A.C.P. Silveira, C.M. Fraz â o, C. Moe, E. Tags: peroxidases oxidative stress radiation resistance dye-decolourizing peroxidases Deinococcus radiodurans research communications Source Type: research

Structural polymorphism of the Escherichia coli poly- α -l-glutamate synthetase RimK
Bacterial RimK is an enzyme that catalyzes the polyglutamylation of the C-terminus of ribosomal protein S6 and the synthesis of poly- α -l-glutamate peptides using l-glutamic acid. In the present study, the crystal structure of the Escherichia coli RimK protein complexed with the ATP analogue AMP-PNP was determined at 2.05   Å resolution. Two different conformations of RimK, closed and open forms, were observed in the crystals. The structural polymorphism revealed in this study provided important information to understand the mechanism by which RimK catalyzes the synthesis of poly- α -l-glutamate peptid...
Source: Acta Crystallographica Section F - June 26, 2018 Category: Biochemistry Authors: Arimura, Y. Kono, T. Kino, K. Kurumizaka, H. Tags: RimK poly- α -l-glutamate synthetase structural polymorphism Escherichia coli research communications Source Type: research

Structure and analysis of nucleoside diphosphate kinase from Borrelia burgdorferi prepared in a transition-state complex with ADP and vanadate moieties
Nucleoside diphosphate kinases (NDKs) are implicated in a wide variety of cellular functions owing to their enzymatic conversion of NDP to NTP. NDK from Borrelia burgdorferi (BbNDK) was selected for functional and structural analysis to determine whether its activity is required for infection and to assess its potential for therapeutic inhibition. The Seattle Structural Genomics Center for Infectious Diseases (SSGCID) expressed recombinant BbNDK protein. The protein was crystallized and structures were solved of both the apoenzyme and a liganded form with ADP and vanadate ligands. This provided two structures and allowed t...
Source: Acta Crystallographica Section F - May 31, 2018 Category: Biochemistry Authors: Dumais, M. Davies, D.R. Lin, T. Staker, B.L. Myler, P.J. Van Voorhis, W.C. Tags: nucleoside diphosphate kinase structural genomics Seattle Structural Genomics Center for Infectious Disease bacterial metabolism protein crystallization protein structure Borrelia burgdorferi Lyme disease research communications Source Type: research

PsEst3, a new psychrophilic esterase from the Arctic bacterium Paenibacillus sp. R4: crystallization and X-ray crystallographic analysis
Esterases are very useful biocatalysts in industry: they hydrolyze esters and split them into a carboxylic acid and an alcohol. The psychrophilic esterase PsEst3 was obtained from Paenibacillus sp. R4, which was isolated from the active layer of the permafrost in Council, Alaska. PsEst3 was successfully overexpressed using a psychrophilic chaperonin co-expression system and was purified by nickel-affinity and size-exclusion chromatography. Recombinant PsEst3 was crystallized at 290   K using the hanging-drop vapour-diffusion method. X-ray diffraction data were collected to 2.1   Å resolution. The crystal was dete...
Source: Acta Crystallographica Section F - May 31, 2018 Category: Biochemistry Authors: Kim, H. Park, A.K. Lee, J.H. Shin, S.C. Park, H. Kim, H.-W. Tags: crystal structure esterase Paenibacillus sp. R4 psychrophilic Artic bacteria research communications Source Type: research

PicW2 from Picea wilsonii: preparation, purification, crystallization and X-ray diffraction analysis
Low temperature is a major limiting factor for plant growth and development. Dehydrin proteins are generally induced in response to low-temperature stress. In previous research, a full-length dehydrin gene, PicW2, was isolated from Picea wilsonii and its expression was associated with hardiness to cold. In order to gain insight into the mechanism of low-temperature tolerance by studying its three-dimensional crystal structure, prokaryotically expressed PicW2 dehydrin protein was purified using chitosan-affinity chromatography and gel filtration, and crystallized using the vapour-diffusion method. The crystal grew in a cond...
Source: Acta Crystallographica Section F - May 24, 2018 Category: Biochemistry Authors: Zhang, B. Guo, G. Lu, F. Song, Y. Liu, Y. Xu, J. Gao, W. Tags: PicW2 dehydrin Picea wilsonii crystallization low-temperature tolerance research communications Source Type: research

Crystal structure of Escherichia coli purine nucleoside phosphorylase in complex with 7-deazahypoxanthine
Purine nucleoside phosphorylases (EC 2.4.2.1; PNPs) reversibly catalyze the phosphorolytic cleavage of glycosidic bonds in purine nucleosides to generate ribose 1-phosphate and a free purine base, and are key enzymes in the salvage pathway of purine biosynthesis. They also catalyze the transfer of pentosyl groups between purine bases (the transglycosylation reaction) and are widely used for the synthesis of biologically important analogues of natural nucleosides, including a number of anticancer and antiviral drugs. Potent inhibitors of PNPs are used in chemotherapeutic applications. The detailed study of the binding of pu...
Source: Acta Crystallographica Section F - May 23, 2018 Category: Biochemistry Authors: Timofeev, V.I. Zhukhlistova, N.E. Abramchik, Y.A. Fateev, I.I. Kostromina, M.A. Muravieva, T.I. Esipov, R.S. Kuranova, I.P. Tags: purine nucleoside phosphorylase Escherichia coli 7DHX inhibitor complex transferases 7-deazahypoxanthine research communications Source Type: research

Crystallographic analysis of the Staphylococcus epidermidis lipase involved in esterification in aqueous solution
This study reports the crystallization and crystallographic analysis of recombinant GehC (rGehC; Lys303 – Lys688) with a molecular weight of 43   kDa. rGehC was crystallized at 293   K using PEG 10   000 as a precipitant, and a 99.9% complete native data set was collected from a cooled crystal at 77   K to a resolution of 1.9   Å with an overall Rmerge value of 7.3%. The crystals were orthorhombic and belonged to space group P212121, with unit-cell parameters a = 42.07, b = 59.31, c   = 171.30   Å , α = β = γ = 90 ° . Solvent-content calculations suggest that there is...
Source: Acta Crystallographica Section F - May 21, 2018 Category: Biochemistry Authors: Liu, C.-H. Chen, Y.-T. Hou, M.-H. Hu, N.-J. Chen, C.-S. Shaw, J.-F. Tags: Staphylococcus epidermidis lipase SeLip catalytic mechanism aqueous media research communications Source Type: research

Perfect merohedral twinning combined with noncrystallographic symmetry potentially causes the failure of molecular replacement with low-homology search models for the flavin-dependent halogenase HalX from Xanthomonas campestris
Flavin-dependent halogenases can be used as biocatalysts because they regioselectively halogenate their substrates under mild reaction conditions. New halogenases with novel substrate specificities will add to the toolbox of enzymes available to organic chemists. HalX, the product of the xcc-b100_4193 gene, is a putative flavin-dependent halogenase from Xanthomonas campestris. The enzyme was recombinantly expressed and crystallized in order to aid in identifying its hitherto unknown substrate. Native data collected to a resolution of 2.5   Å showed indications of merohedral twinning in a hexagonal lattice. Attempts...
Source: Acta Crystallographica Section F - May 18, 2018 Category: Biochemistry Authors: Buss, M. Geerds, C. Patschkowski, T. Niehaus, K. Niemann, H.H. Tags: FAD-dependent halogenase hemihedral twinning perfect twin rotational pseudosymmetry merohedral twinning research communications Source Type: research

T4 lysozyme-facilitated crystallization of the human molybdenum cofactor-dependent enzyme mARC
In this study, the use of a fusion-protein approach with T4 lysozyme (T4L) to determine the structure of this hitherto noncrystallizable enzyme by X-ray crystallography is described. A set of four different hmARC-T4L fusion proteins were designed. Two of them contained either an N-terminal or a C-terminal T4L moiety fused to hmARC, while the other two contained T4L as an internal fusion partner tethered to the hmARC enzyme between two predicted secondary-structure elements. One of these internal fusion constructs could be expressed and crystallized successfully. The hmARC-T4L crystals diffracted to 1.7   Å resoluti...
Source: Acta Crystallographica Section F - May 17, 2018 Category: Biochemistry Authors: Kubitza, C. Ginsel, C. Bittner, F. Havemeyer, A. Clement, B. Scheidig, A.J. Tags: T4 lysozyme fusion protein crystallization strategy carrier-driven crystallization research communications Source Type: research

Crystal structure of an inactive variant of the quorum-sensing master regulator HapR from the protease-deficient non-O1, non-O139 Vibrio cholerae strain V2
In this study, the structure of a DNA-binding-deficient variant of HapR (HapRV2) derived from the protease-deficient V. cholerae serotype O37 strain V2 is reported. The structure reveals no structural differences compared with wild-type HapR. However, structural alignment of HapRV2 with the TetR-family member QacR in complex with its operator DNA suggests that the aspartate residue located between the regulatory and DNA-binding domains may clash with and electrostatically repel the phosphate backbone of DNA to prevent binding. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - May 17, 2018 Category: Biochemistry Authors: Cruite, J. Succo, P. Raychaudhuri, S. Kull, F.J. Tags: HapR quorum sensing Vibrio cholerae TetR transcriptional regulator research communications Source Type: research

Carbonic anhydrase II microcrystals suitable for XFEL studies
In this study, the growth of carbonic anhydrase II microcrystals (40 – 80   µ m in length) suitable for the collection of XFEL diffraction data at the Pohang Accelerator Laboratory is demonstrated. The crystals diffracted to 1.7   Å resolution and were indexed in space group P21, with unit-cell parameters a = 42.2, b = 41.2, c = 72.0   Å , β   =   104.2 ° . These preliminary results provide the necessary framework for time-resolved experiments to study carbonic anhydrase catalysis at XFEL beamlines. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - May 17, 2018 Category: Biochemistry Authors: Lomelino, C.L. Kim, J.K. Lee, C. Lim, S.W. Andring, J.T. Mahon, B.P. Chung, M. Kim, C.U. McKenna, R. Tags: X-ray free-electron lasers XFELs serial femtosecond crystallography time-resolved crystallography microcrystals carbonic anhydrase II research communications Source Type: research

The CD163 long-range scavenger receptor cysteine-rich repeat: expression, purification and X-ray crystallographic characterization
In this study, the CD163 long-range SRCR repeat was expressed in Drosophila Schneider 2 cells. The recombinant protein was homogeneous after purification by metal-affinity, cation-exchange and size-exclusion chromatography. Single crystals were obtained using 20% PEG 4000, 0.15   M potassium sodium tartrate tetrahydrate pH 8.5 and diffracted to 3.30   Å resolution. As the first view of a long-range SRCR repeat, this work lays the structural basis for a deep understanding of SRs and their multiple functions. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 25, 2018 Category: Biochemistry Authors: Li, R. Ma, H. Jiang, L. Qiao, S. Zhi, Y. Huang, M. Deng, R. Zhang, G. Tags: scavenger receptor long-range scavenger receptor cysteine-rich repeat SRCR repeat CD163 X-ray crystallography research communications Source Type: research

Structure of the tandem PX-PH domains of Bem3 from Saccharomyces cerevisiae
The structure of the tandem lipid-binding PX and pleckstrin-homology (PH) domains of the Cdc42 GTPase-activating protein Bem3 from Saccharomyces cerevisiae (strain S288c) has been determined to a resolution of 2.2   Å (Rwork = 21.1%, Rfree = 23.4%). It shows that the domains adopt a relative orientation that enables them to simultaneously bind to a membrane and suggests possible cooperativity in membrane binding. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 24, 2018 Category: Biochemistry Authors: Ali, I. Eu, S. Koch, D. Bleimling, N. Goody, R.S. M ü ller, M.P. Tags: PX domain PH domain phox pleckstrin homology phosphatidylinositol phosphates PIP Saccharomyces cerevisiae Bem3 research communications Source Type: research

Improved protein-crystal identification by using 2,2,2-trichloroethanol as a fluorescence enhancer
The identification of initial lead conditions for successful protein crystallization is crucial for structural studies using X-ray crystallography. In order to reduce the number of false-negative conditions, an emerging number of fluorescence-based methods have been developed which allow more efficient identification of protein crystals and help to distinguish them from salt crystals. Detection of the native tryptophan fluorescence of protein crystals is one of the most widely used methods. However, this method can fail owing to the properties of the crystallized protein or the chemical composition of the crystallization t...
Source: Acta Crystallographica Section F - April 24, 2018 Category: Biochemistry Authors: Pichlo, C. Toelzer, C. Chojnacki, K. Ö cal, S. Uthoff, M. Ruegenberg, S. Hermanns, T. Schacherl, M. Denzel, M.S. Hofmann, K. Niefind, K. Baumann, U. Tags: protein-crystal identification tryptophan fluorescence 2,2,2-trichloroethanol PPEP-1 research communications Source Type: research

In situ proteolysis of an N-terminal His tag with thrombin improves the diffraction quality of human aldo-keto reductase 1C3 crystals
Human aldo-keto reductase 1C3 (AKR1C3) stereospecifically reduces steroids and prostaglandins and is involved in the biotransformation of xenobiotics. Its role in various cancers makes it a potential therapeutic target for the development of inhibitors. Recombinant AKR1C3 with a thrombin-cleavable N-terminal His6 tag was expressed from a pET-28(+) vector for structural studies of enzyme – inhibitor complexes. A modified in situ proteolysis approach was applied to specifically remove the His tag by thrombin cleavage during crystallization screening trials. This improved the morphology and diffraction quality of the cr...
Source: Acta Crystallographica Section F - April 24, 2018 Category: Biochemistry Authors: Plav š a, J.J. Ř ez á č ov á , P. Kugler, M. Pachl, P. Brynda, J. Voburka, Z. Ć eli ć , A. Petri, E.T. Š kerlov á , J. Tags: aldo-keto reductase 1C3 in situ proteolysis His tags diffraction-quality improvement pET-28( ) 17 β -hydroxysteroid dehydrogenase 5 research communications Source Type: research

Crystal structure of chorismate mutase from Burkholderia thailandensis
Burkholderia thailandensis is often used as a model for more virulent members of this genus of proteobacteria that are highly antibiotic-resistant and are potential agents of biological warfare that are infective by inhalation. As part of ongoing efforts to identify potential targets for the development of rational therapeutics, the structures of enzymes that are absent in humans, including that of chorismate mutase from B. thailandensis, have been determined by the Seattle Structural Genomics Center for Infectious Disease. The high-resolution structure of chorismate mutase from B. thailandensis was determined in the monoc...
Source: Acta Crystallographica Section F - April 16, 2018 Category: Biochemistry Authors: Asojo, O.A. Dranow, D.M. Serbzhinskiy, D. Subramanian, S. Staker, B. Edwards, T.E. Myler, P.J. Tags: structural genomics Seattle Structural Genomics Center for Infectious Disease Burkholderia thailandensis chorismate mutase isomerases research communications Source Type: research

Crystal structures of human CK2 α 2 in new crystal forms arising from a subtle difference in salt concentration
In this study, new crystal forms were exploited and one provided a crystal structure of CK2 α 2 at 1.89   Å resolution. This result, together with the structure of CK2 α 1, will assist in the development of highly selective inhibitors for both isozymes. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 16, 2018 Category: Biochemistry Authors: Tsuyuguchi, M. Nakaniwa, T. Kinoshita, T. Tags: CK2 kinase catalytic subunit crystal structure human CK2 α polymorphism male contraceptives research communications Source Type: research

The putative siderophore-interacting protein from Vibrio anguillarum: protein production, analysis, crystallization and X-ray crystallographic studies
Siderophore-interacting proteins (SIPs) play an important role in iron acquisition in many bacteria. SIPs release iron from the internalized ferric siderophore complex by reducing ferric iron to ferrous iron, but how the iron is reduced is not well understood. Here, a sip gene was identified in the genome of Vibrio anguillarum 775. To further understand the catalytic mechanism of the protein, the SIP was overexpressed in Escherichia coli Rosetta (DE3) cells, purified and crystallized for X-ray diffraction analysis. The crystal diffracted to 1.113   Å resolution and belonged to space group P21, with unit-cell parame...
Source: Acta Crystallographica Section F - April 16, 2018 Category: Biochemistry Authors: Han, Y. Zang, K. Liu, C. Li, Y. Ma, Q. Tags: Vibrio anguillarum siderophore-interacting proteins iron release siderophores research communications Source Type: research

Crystal structure of the mouse innate immunity factor bacterial permeability-increasing family member A1
Bacterial permeability-increasing family member A1 (BPIFA1) is an innate immunity factor and one of the most abundantly secreted proteins in the upper airways. BPIFA1 is multifunctional, with antimicrobial, surfactant and lipopolysaccharide-binding activities, as well as established roles in lung hydration. Here, the 2.5   Å resolution crystal structure of BPIFA1 from Mus musculus (mBPIFA1) is presented and compared with those of human BPIFA1 (hBPIFA1) and structural homologs. Structural distinctions between mBPIFA1 and hBPIFA1 suggest potential differences in biological function, including the regulation of a key ...
Source: Acta Crystallographica Section F - April 16, 2018 Category: Biochemistry Authors: Little, M.S. Redinbo, M.R. Tags: bacterial permeability-increasing family member A1 BPIFA1 Mus musculus biological chemistry structural biology lung biology innate immunity proteins surfactant proteins pulmonary proteins protein surfactants antimicrobial proteins re Source Type: research

A cryoprotectant induces conformational change in glyceraldehyde-3-phosphate dehydrogenase
In this study, the three-dimensional crystal structure of GAPDH treated with trehalose is reported at 2.0   Å resolution. Trehalose was used as a cryoprotectant for the GAPDH crystals. The structure of trehalose-bound ecGAPDH was compared with the structures of both NAD+-free and NAD+-bound ecGAPDH. At the S-loop, the bound trehalose in the GAPDH structure induces a 2.4 ° rotation compared with the NAD+-free ecGAPDH structure and a 3.1 ° rotation compared with the NAD+-bound ecGAPDH structure. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - April 16, 2018 Category: Biochemistry Authors: Kim, Y.J. Tags: crystal structure glyceraldehyde-3-phosphate dehydrogenase GAPDH cryoprotectants trehalose research communications Source Type: research

Crystallization of the rice immune receptor RGA5A_S with the rice blast fungus effector AVR1-CO39 prepared via mixture and tandem strategies
RGA5 is a component of the Pia resistance-protein pair (RGA4/RGA5) from Oryza sativa L. japonica. It acts as an immune receptor that directly recognizes the effector AVR1-CO39 from Magnaporthe oryzae via a C-terminal non-LRR domain (RGA5A_S). The interaction between RGA5A_S and AVR1-CO39 relieves the repression of RGA4, leading to effector-independent cell death. To determine the structure of the complex of RGA5A_S and AVR1-CO39 and to understand the details of this interaction, the complex was prepared by fusing the proteins together, by mixing them in vitro or by co-expressing them in one host cell. Samples purified via ...
Source: Acta Crystallographica Section F - March 28, 2018 Category: Biochemistry Authors: Guo, L. Zhang, Y. Ma, M. Liu, Q. Zhang, Y. Peng, Y. Liu, J. Tags: RGA5A_S AVR1-CO39 rice Magnaporthe oryzae resistance protein Avr effector crystal complex research communications Source Type: research