Purification, crystallization and initial crystallographic analysis of the α-catenin homologue HMP-1 from Caenorhabditis elegans
Adherens junctions transmit mechanical force between cells. In these junctions, β-catenin binds to cadherins and to the N-terminal domain of α-catenin, which in turn binds to actin filaments via its C-terminal domain. The middle (M) domain of α-catenin plays an important role in responding to mechanical tension. The nematode Caenorhabditis elegans contains α- and β-catenin homologues called HMP-1 and HMP-2, respectively, but HMP-1 behaves differently from its mammalian homologue. Thus, structural and biochemical studies of HMP-1 have been initiated to understand the mechanism of HMP-1 and the ev...
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: Kang, H.Bang, I.Weis, W.I.Choi, H.-J. Tags: α -catenin HMP-1 adherens junction Caenorhabditis elegans research communications Source Type: research

Structural investigation of the thymidine phosphorylase from Salmonella typhimurium in the unliganded state and its complexes with thymidine and uridine
Highly specific thymidine phosphorylases catalyze the phosphorolytic cleavage of thymidine, with the help of a phosphate ion, resulting in thymine and 2-deoxy-α-d-ribose 1-phosphate. Thymidine phosphorylases do not catalyze the phosphorolysis of uridine, in contrast to nonspecific pyrimidine nucleoside phosphorylases and uridine phosphorylases. Understanding the mechanism of substrate specificity on the basis of the nucleoside is essential to support rational drug-discovery investigations of new antitumour and anti-infective drugs which are metabolized by thymidine phosphorylases. For this reason, X-ray structures of...
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: Balaev, V.V.Lashkov, A.A.Gabdulkhakov, A.G.Dontsova, M.V.Seregina, T.A.Mironov, A.S.Betzel, C.Mikhailov, A.M. Tags: X-ray analysis protein crystallography nucleoside phosphorylases nucleosides substrate specificity Salmonella typhimurium thymidine phosphorylase research communications Source Type: research

Neutron diffraction analysis of Pseudomonas aeruginosa peptidyl-tRNA hydrolase 1
Perdeuterated peptidyl-tRNA hydrolase 1 from Pseudomonas aeruginosa was crystallized for structural analysis using neutron diffraction. Crystals of perdeuterated protein were grown to 0.15 mm3 in size using batch crystallization in 22.5% polyethylene glycol 4000, 100 mM Tris pH 7.5, 10%(v/v) isopropyl alcohol with a 20-molar excess of trilysine as an additive. Neutron diffraction data were collected from a crystal at room temperature using the MaNDi single-crystal diffractometer at Oak Ridge National Laboratory. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: McFeeters, H.Vandavasi, V.G.Weiss, K.L.Coates, L.McFeeters, R.L. Tags: peptidyl-tRNA hydrolase 1 neutron diffraction perdeuteration Pseudomonas aeruginosa antibiotic resistance research communications Source Type: research

Crystal structure of FhuD at 1.6 Å resolution: a ferrichrome-binding protein from the animal and human pathogen Staphylococcus pseudintermedius
Staphylococcus pseudintermedius is a leading cause of disease in dogs, and zoonosis causes human infections. Methicillin-resistant S. pseudintermedius strains are emerging, resembling the global health threat of S. aureus. Therefore, it is increasingly important to characterize potential targets for intervention against S. pseudintermedius. Here, FhuD, an S. pseudintermedius surface lipoprotein implicated in iron uptake, was characterized. It was found that FhuD bound ferrichrome in an iron-dependent manner, which increased the thermostability of FhuD by>15°C. The crystal structure of ferrichrome-free FhuD was deter...
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: Abate, F.Cozzi, R.Maritan, M.Lo Surdo, P.Maione, D.Malito, E.Bottomley, M.J. Tags: metal transport staphylococcal disease siderophore iron class III solute-binding protein research communications Source Type: research

Purification, crystallization and X-ray crystallographic analysis of a putative exopolyphosphatase from Zymomonas mobilis
In this study, a putative exopolyphosphatase from Zymomonas mobilis (ZmPPX) was crystallized. Crystals of the wild-type enzyme diffracted to 3.3 Å resolution and could not be optimized further. The truncation of 29 amino acids from the N-terminus resulted in crystals that diffracted to 1.8 Å resolution. The crystals belonged to space group C2, with unit-cell parameters a = 122.0, b = 47.1, c = 89.5 Å, α = γ = 90, β = 124.5°. An active-site mutant that crystallized in the same space group and with similar unit-cell parameters diffracted to 1.56 Å resolution. One molecule ...
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: Zhang, A.Guo, E.Qian, L.Tang, N.-Y.Watt, R.M.Bartlam, M. Tags: exopolyphosphatase PPX/GppA family crystallization Zymomonas mobilis research communications Source Type: research

Crystal structure of recombinant tyrosinase-binding protein MtaL at 1.35   Å resolution
In this study, the crystal structure of recombinant MtaL is reported at 1.35   Å resolution. Comparison of its structure with that of the truncated and cleaved MtaL present in the complex with tyrosinase directly isolated from mushroom shows that the general β -trefoil fold is conserved. However, differences are detected in the loop regions, particularly in the β 2 – β 3 loop, which is intact and not cleaved in the recombinant MtaL. Furthermore, the N-terminal tail is rotated inwards, covering the tyrosinase-binding interface. Thus, MtaL must undergo conformational changes in order to bind matu...
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: Lai, X. Soler-Lopez, M. Ismaya, W.T. Wichers, H.J. Dijkstra, B.W. Tags: tyrosinase lectin tyrosinase-binding protein MtaL carbohydrate binding research communications Source Type: research

Purification, crystallization and initial crystallographic analysis of the α -catenin homologue HMP-1 from Caenorhabditis elegans
Adherens junctions transmit mechanical force between cells. In these junctions, β -catenin binds to cadherins and to the N-terminal domain of α -catenin, which in turn binds to actin filaments via its C-terminal domain. The middle (M) domain of α -catenin plays an important role in responding to mechanical tension. The nematode Caenorhabditis elegans contains α - and β -catenin homologues called HMP-1 and HMP-2, respectively, but HMP-1 behaves differently from its mammalian homologue. Thus, structural and biochemical studies of HMP-1 have been initiated to understand the mechanism of HMP-1 and t...
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: Kang, H. Bang, I. Weis, W.I. Choi, H.-J. Tags: α -catenin HMP-1 adherens junction Caenorhabditis elegans research communications Source Type: research

Crystal structure of FhuD at 1.6   Å resolution: a ferrichrome-binding protein from the animal and human pathogen Staphylococcus pseudintermedius
Staphylococcus pseudintermedius is a leading cause of disease in dogs, and zoonosis causes human infections. Methicillin-resistant S. pseudintermedius strains are emerging, resembling the global health threat of S. aureus. Therefore, it is increasingly important to characterize potential targets for intervention against S. pseudintermedius. Here, FhuD, an S. pseudintermedius surface lipoprotein implicated in iron uptake, was characterized. It was found that FhuD bound ferrichrome in an iron-dependent manner, which increased the thermostability of FhuD by>15 ° C. The crystal structure of ferrichrome-free FhuD was det...
Source: Acta Crystallographica Section F - February 19, 2016 Category: Biochemistry Authors: Abate, F. Cozzi, R. Maritan, M. Lo Surdo, P. Maione, D. Malito, E. Bottomley, M.J. Tags: metal transport staphylococcal disease siderophore iron class III solute-binding protein research communications Source Type: research

Cloning, expression, purification, crystallization and X-ray crystallographic analysis of recombinant human C1ORF123 protein
C1ORF123 is a human hypothetical protein found in open reading frame 123 of chromosome 1. The protein belongs to the DUF866 protein family comprising eukaryote-conserved proteins with unknown function. Recent proteomic and bioinformatic analyses identified the presence of C1ORF123 in brain, frontal cortex and synapses, as well as its involvement in endocrine function and polycystic ovary syndrome (PCOS), indicating the importance of its biological role. In order to provide a better understanding of the biological function of the human C1ORF123 protein, the characterization and analysis of recombinant C1ORF123 (rC1ORF123), ...
Source: Acta Crystallographica Section F - February 16, 2016 Category: Biochemistry Authors: Rahaman, S.N.A.Mat Yusop, J.Mohamed-Hussein, Z.-A.Ho, K.L.Teh, A.-H.Waterman, J.Ng, C.L. Tags: C1ORF123 hypothetical protein DUF866 polycystic ovary syndrome bioinformatic analysis research communications Source Type: research

Structure of a truncated form of leucine zipper II of JIP3 reveals an unexpected antiparallel coiled-coil arrangement
This study highlights that N-terminal truncation of LZII can change its coiled-coil orientation without affecting its overall stability. Further, a conserved buried asparagine residue was pinpointed as a possible structural determinant for this dramatic structural rearrangement. Thus, LZII of JIP3/4 is a versatile structural motif, modifications of which can impact partner recognition and thus biological function. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - February 16, 2016 Category: Biochemistry Authors: Llinas, P.Chenon, M.Nguyen, T.Q.Moreira, C.de Régibus, A.Coquard, A.Ramos, M.J.Guérois, R.Fernandes, P.A.Ménétrey, J. Tags: JIP3 JIP4 leucine zipper II crystal structure molecular dynamics antiparallel coiled coil buried asparagine research communications Source Type: research

The use of noncrystallographic symmetry averaging to solve structures from data affected by perfect hemihedral twinning
Hemihedral twinning is a crystal-growth anomaly in which a specimen is composed of two crystal domains that coincide with each other in three dimensions. However, the orientations of the crystal lattices in the two domains differ in a specific way. In diffraction data collected from hemihedrally twinned crystals, each observed intensity contains contributions from both of the domains. With perfect hemihedral twinning, the two domains have the same volumes and the observed intensities do not contain sufficient information to detwin the data. Here, the use of molecular replacement and of noncrystallographic symmetry (NCS) av...
Source: Acta Crystallographica Section F - February 16, 2016 Category: Biochemistry Authors: Sabin, C.Plevka, P. Tags: hemihedral perfect twinning noncrystallographic symmetry averaging molecular replacement virus structure detwinning twin domain merohedral twinning mask envelope research communications Source Type: research

Crystal structure of 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase from the ESKAPE pathogen Acinetobacter baumannii
The enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase catalyzes the sixth step of the seven-step shikimate pathway. Chorismate, the product of the pathway, is a precursor for the biosynthesis of aromatic amino acids, siderophores and metabolites such as folate, ubiquinone and vitamin K. The shikimate pathway is present in bacteria, fungi, algae, plants and apicomplexan parasites, but is absent in humans. The EPSP synthase enzyme produces 5-enolpyruvylshikimate 3-phosphate and phosphate from phosphoenolpyruvate and shikimate 3-phosphate via a transferase reaction, and is the target of the herbicide glyphosate. The A...
Source: Acta Crystallographica Section F - February 16, 2016 Category: Biochemistry Authors: Sutton, K.A.Breen, J.Russo, T.A.Schultz, L.W.Umland, T.C. Tags: shikimate pathway Acinetobacter baumannii essential genes antibiotic targets multidrug resistance EPSP synthase research communications Source Type: research

Crystal structure of maize serine racemase with pyridoxal 5′-phosphate
Serine racemase (SR) is a pyridoxal 5′-phosphate (PLP)-dependent enzyme that is responsible for d-serine biosynthesis in vivo. The first X-ray crystal structure of maize SR was determined to 2.1 Å resolution and PLP binding was confirmed in solution by UV–Vis absorption spectrometry. Maize SR belongs to the type II PLP-dependent enzymes and differs from the SR of a vancomycin-resistant bacterium. The PLP is bound to each monomer by forming a Schiff base with Lys67. Structural comparison with rat and fission yeast SRs reveals a similar arrangement of active-site residues but a different orientation of th...
Source: Acta Crystallographica Section F - February 16, 2016 Category: Biochemistry Authors: Zou, L.Song, Y.Wang, C.Sun, J.Wang, L.Cheng, B.Fan, J. Tags: structural comparison serine racemase maize crystal structure cofactor pyridoxal 5 ′ -phosphate research communications Source Type: research

Crystal structure of maize serine racemase with pyridoxal 5 ′ -phosphate
Serine racemase (SR) is a pyridoxal 5 ′ -phosphate (PLP)-dependent enzyme that is responsible for d-serine biosynthesis in vivo. The first X-ray crystal structure of maize SR was determined to 2.1   Å resolution and PLP binding was confirmed in solution by UV – Vis absorption spectrometry. Maize SR belongs to the type II PLP-dependent enzymes and differs from the SR of a vancomycin-resistant bacterium. The PLP is bound to each monomer by forming a Schiff base with Lys67. Structural comparison with rat and fission yeast SRs reveals a similar arrangement of active-site residues but a different orientation...
Source: Acta Crystallographica Section F - February 16, 2016 Category: Biochemistry Authors: Zou, L. Song, Y. Wang, C. Sun, J. Wang, L. Cheng, B. Fan, J. Tags: structural comparison serine racemase maize crystal structure cofactor pyridoxal 5 ′ -phosphate research communications Source Type: research

Crystal structure of human GDF11
Members of the TGF-β family of proteins are believed to play critical roles in cellular signaling processes such as those involved in muscle differentiation. The extent to which individual family members have been characterized and linked to biological function varies greatly. The role of myostatin, also known as growth differentiation factor 8 (GDF8), as an inhibitor of muscle differentiation is well understood through genetic linkages. In contrast, the role of growth differentiation factor 11 (GDF11) is much less well understood. In humans, the mature forms of GDF11 and myostatin are over 94% identical. In order to ...
Source: Acta Crystallographica Section F - February 15, 2016 Category: Biochemistry Authors: Padyana, A.K.Vaidialingam, B.Hayes, D.B.Gupta, P.Franti, M.Farrow, N.A. Tags: growth differentiation factor 11 GDF11 TGF- β family research communications Source Type: research

Improving diffraction resolution using a new dehydration method
The production of high-quality crystals is one of the major obstacles in determining the three-dimensional structure of macromolecules by X-ray crystallography. It is fairly common that a visually well formed crystal diffracts poorly to a resolution that is too low to be suitable for structure determination. Dehydration has proven to be an effective post-crystallization treatment for improving crystal diffraction quality. Several dehydration methods have been developed, but no single one of them is suitable for all crystals. Here, a new convenient and effective dehydration method is reported that makes use of a dehydrating...
Source: Acta Crystallographica Section F - January 28, 2016 Category: Biochemistry Authors: Huang, Q.Szebenyi, D.M.E. Tags: protein crystal dehydration diffraction resolution Cas5a LptA research communications Source Type: research

Protein stability: a crystallographer's perspective
Protein stability is a topic of major interest for the biotechnology, pharmaceutical and food industries, in addition to being a daily consideration for academic researchers studying proteins. An understanding of protein stability is essential for optimizing the expression, purification, formulation, storage and structural studies of proteins. In this review, discussion will focus on factors affecting protein stability, on a somewhat practical level, particularly from the view of a protein crystallographer. The differences between protein conformational stability and protein compositional stability will be discussed, along...
Source: Acta Crystallographica Section F - January 28, 2016 Category: Biochemistry Authors: Deller, M.C.Kong, L.Rupp, B. Tags: protein stability protein crystallization protein disorder crystallizability research communications Source Type: research

Crystallization and X-ray diffraction of LGN in complex with the actin-binding protein afadin
Asymmetric stem-cell divisions are fundamental for morphogenesis and tissue homeostasis. They rely on the coordination between cortical polarity and the orientation of the mitotic spindle, which is orchestrated by microtubule pulling motors recruited at the cortex by NuMA–LGN–Gαi complexes. LGN has emerged as a central component of the spindle-orientation pathway that is conserved throughout species. Its domain structure consists of an N-terminal TPR domain associating with NuMA, followed by four GoLoco motifs binding to Gαi subunits. The LGNTPR region is also involved in interactions with other mem...
Source: Acta Crystallographica Section F - January 26, 2016 Category: Biochemistry Authors: Carminati, M.Cecatiello, V.Mapelli, M. Tags: LGN spindle orientation fusion proteins research communications Source Type: research

The role of mass transport in protein crystallization
Mass transport takes place within the mesoscopic to macroscopic scale range and plays a key role in crystal growth that may affect the result of the crystallization experiment. The influence of mass transport is different depending on the crystallization technique employed, essentially because each technique reaches supersaturation in its own unique way. In the case of batch experiments, there are some complex phenomena that take place at the interface between solutions upon mixing. These transport instabilities may drastically affect the reproducibility of crystallization experiments, and different outcomes may be obtaine...
Source: Acta Crystallographica Section F - January 26, 2016 Category: Biochemistry Authors: García-Ruiz, J.M.Otálora, F.García-Caballero, A. Tags: mass transport crystal growth protein crystallization research communications Source Type: research

Crystallographic observation of the movement of the membrane-distal domain of the T7SS core component EccB1 from Mycobacterium tuberculosis
The protein EccB1, a core component of the type VII secretion system (T7SS) of Mycobacterium tuberculosis, has been identified as an ATPase and is essential for the secretion of virulence factors by the ESX-1 system. In a previous study, EccB1 structures were determined in two different conformations. Here, two new conformations are identified and described. These four conformations present snapshots of the swinging movement of the membrane-distal domain A2. The movement of this domain involves conformational changes in two flexible loops (loop A, residues 243–264, and loop B, residues 324–341) which are rich i...
Source: Acta Crystallographica Section F - January 22, 2016 Category: Biochemistry Authors: Xie, X.-Q.Zhang, X.-L.Qi, C.Li, D.-F.Fleming, J.Wang, D.-C.Bi, L.-J. Tags: Rv3869 type VII secretion system Mycobacterium tuberculosis ESX-1 research communications Source Type: research

Cloning, expression, purification, crystallization and initial crystallographic analysis of FleN from Pseudomonas aeruginosa
The assembly of bacterial flagella requires the coordinated expression of a large number of genes in a hierarchical manner. These genes code for structural components of flagella, regulatory components and components that are required for chemotaxis. Stringent spatial and numerical control of flagella biosynthesis is essential for promoting motility and pathogenesis in bacteria. These genes are regulated at the level of transcription. FleN, a P-loop-containing ATPase, plays an important role in maintaining flagellar number in Pseudomonas aeruginosa. FleN exhibits anti-activator activity against FleQ, the global transcripti...
Source: Acta Crystallographica Section F - January 22, 2016 Category: Biochemistry Authors: HarshitaChanchalJain, D. Tags: FleN transcription flagella Pseudomonas research communications Source Type: research

Crystal structure of microtubule affinity-regulating kinase 4 catalytic domain in complex with a pyrazolopyrimidine inhibitor
Microtubule-associated protein/microtubule affinity-regulating kinase 4 (MARK4) is a serine/threonine kinase involved in the phosphorylation of MAP proteins that regulate microtubule dynamics. Abnormal activity of MARK4 has been proposed to contribute to neurofibrillary tangle formation in Alzheimer's disease. The crystal structure of the catalytic and ubiquitin-associated domains of MARK4 with a potent pyrazolopyrimidine inhibitor has been determined to 2.8 Å resolution with an Rwork of 22.8%. The overall structure of MARK4 is similar to those of the other known MARK isoforms. The inhibitor is located in the ATP-b...
Source: Acta Crystallographica Section F - January 22, 2016 Category: Biochemistry Authors: Sack, J.S.Gao, M.Kiefer, S.E.Myers, J.E.Newitt, J.A.Wu, S.Yan, C. Tags: MARK4 PAR-1 serine/threonine protein kinase transferase research communications Source Type: research

Structure of the human histone chaperone FACT Spt16 N-terminal domain
The histone chaperone FACT plays an important role in facilitating nucleosome assembly and disassembly during transcription. FACT is a heterodimeric complex consisting of Spt16 and SSRP1. The N-terminal domain of Spt16 resembles an inactive aminopeptidase. How this domain contributes to the histone chaperone activity of FACT remains elusive. Here, the crystal structure of the N-terminal domain (NTD) of human Spt16 is reported at a resolution of 1.84 Å. The structure adopts an aminopeptidase-like fold similar to those of the Saccharomyces cerevisiae and Schizosaccharomyces pombe Spt16 NTDs. Isothermal titration calo...
Source: Acta Crystallographica Section F - January 22, 2016 Category: Biochemistry Authors: Marcianò, G.Huang, D.T. Tags: FACT Spt16 histone chaperone histones pita-bread fold aminopeptidase research communications Source Type: research

Structural and functional analysis of the RNA helicase Prp43 from the thermophilic eukaryote Chaetomium thermophilum
RNA helicases are indispensable for all organisms in each domain of life and have implications in numerous cellular processes. The DEAH-box RNA helicase Prp43 is involved in pre-mRNA splicing as well as rRNA maturation. Here, the crystal structure of Chaetomium thermophilum Prp43 at 2.9 Å resolution is revealed. Furthermore, it is demonstrated that Prp43 from C. thermophilum is capable of functionally replacing its orthologue from Saccharomyces cerevisiae in spliceosomal disassembly assays. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - January 22, 2016 Category: Biochemistry Authors: Tauchert, M.J.Fourmann, J.-B.Christian, H.Lührmann, R.Ficner, R. Tags: spliceosome RNA helicase DEAH-box protein DHX15 research communications Source Type: research

Crystal structure of the flavoenzyme PA4991 from Pseudomonas aeruginosa
The locus PA4991 in Pseudomonas aeruginosa encodes an open reading frame that has been identified as essential for the virulence and/or survival of this pathogenic organism in the infected host. Here, it is shown that this gene encodes a monomeric FAD-binding protein of molecular mass 42.2 kDa. The structure of PA4991 was determined by a combination of molecular replacement using a search model generated with Rosetta and phase improvement by a low-occupancy heavy-metal derivative. PA4991 belongs to the GR2 family of FAD-dependent oxidoreductases, comprising an FAD-binding domain typical of the glutathione reductase famil...
Source: Acta Crystallographica Section F - January 22, 2016 Category: Biochemistry Authors: Jacewicz, A.Schnell, R.Lindqvist, Y.Schneider, G. Tags: molecular replacement Rosetta model phasing oxidoreductase FAD research communications Source Type: research

The power of three
(Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - December 1, 2015 Category: Biochemistry Authors: Weiss, M.S. Hunter, W.N. Tags: editorial Section Editor Source Type: research

Crystallographic analysis of RsmA, a ribosomal RNA small subunit methyltransferase A from Staphylococcus aureus. Corrigendum
The paper by Liu et al. [(2015), Acta Cryst. F71, 1063 – 1066] is corrected. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - November 27, 2015 Category: Biochemistry Authors: Liu, Y. Zhu, Y. Teng, M. Li, X. Tags: RsmA ribosomal RNA methyltransferase A Staphylococcus aureus corrigendum addenda and errata Source Type: research

Structures of plasmepsin II from Plasmodium falciparum in complex with two hydroxyethylamine-based inhibitors
Plasmepsin II (PMII) is one of the ten plasmepsins (PMs) identified in the genome of Plasmodium falciparum, the causative agent of the most severe and deadliest form of malaria. Owing to the emergence of P. falciparum strains that are resistant to current antimalarial agents such as chloroquine and sulfadoxine/pyrimethamine, there is a constant pressure to find new and lasting chemotherapeutic drug therapies. Previously, the crystal structure of PMII in complex with NU655, a potent antimalarial hydroxyethylamine-based inhibitor, and the design of new compounds based on it have been reported. In the current study, two of th...
Source: Acta Crystallographica Section F - November 27, 2015 Category: Biochemistry Authors: Recacha, R. Leitans, J. Akopjana, I. Aprupe, L. Trapencieris, P. Jaudzems, K. Jirgensons, A. Tars, K. Tags: malaria Plasmodium falciparum plasmepsin hydroxyethylamine inhibitor research communications Source Type: research

Crystallographic insights into the structure – activity relationships of diazaborine enoyl-ACP reductase inhibitors
Enoyl-ACP reductase, the last enzyme of the fatty-acid biosynthetic pathway, is the molecular target for several successful antibiotics such as the tuberculosis therapeutic isoniazid. It is currently under investigation as a narrow-spectrum antibiotic target for the treatment of several types of bacterial infections. The diazaborine family is a group of boron heterocycle-based synthetic antibacterial inhibitors known to target enoyl-ACP reductase. Development of this class of molecules has thus far focused solely on the sulfonyl-containing versions. Here, the requirement for the sulfonyl group in the diazaborine scaffold w...
Source: Acta Crystallographica Section F - November 27, 2015 Category: Biochemistry Authors: Jordan, C.A. Sandoval, B.A. Serobyan, M.V. Gilling, D.H. Groziak, M.P. Xu, H.H. Vey, J.L. Tags: antibiotics binding sites boron heterocycle enzyme inhibitors NAD Escherichia coli crystallography enoyl-ACP reductase research communications Source Type: research

Expression, purification, crystallization and X-ray diffraction analysis of ChiL, a chitinase from Chitiniphilus shinanonensis
Chitin, a linear polysaccharide consisting of β -1,4-linked N-acetyl-d-glucosamine (GlcNAc), is widely used because of its biochemical properties. GlcNAc oligomers prepared from chitin have useful biological activities, such as immunostimulation and the induction of plant defence responses. Microbial chitinolytic enzymes have been investigated extensively for their potential use in the eco-friendly enzymatic production of GlcNAc and its oligomers. Chitiniphilus shinanonensis SAY3T is a recently found bacterium with a strong chitinolytic activity. The chitinolytic enzymes from this strain are potentially useful for the...
Source: Acta Crystallographica Section F - November 27, 2015 Category: Biochemistry Authors: Ueda, M. Shimosaka, M. Arai, R. Tags: chitinase chitinolytic enzyme Chitiniphilus shinanonensis N-acetylglucosamine family 18 glycoside hydrolases research communications Source Type: research

Crystallographic analysis of a cupin superfamily enzyme from Microcystis aeruginosa involved in aeruginosin biosynthesis
In this study, recombinant AerE from Microcystis aeruginosa, which was overexpressed in Escherichia coli and purified by Ni2+-chelating affinity and gel-filtration chromatography, was successfully crystallized and X-ray diffraction analysis was performed. The crystal belonged to the orthorhombic space group P212121, with unit-cell parameters a = 34.770, b = 62.133, c = 87.401   Å . The diffraction data from the crystal were scaled to a maximum resolution of 1.60   Å . The calculated Matthews coefficient of the crystal is 1.93   Å 3   Da − 1, suggesting that there is one molecule in the asymm...
Source: Acta Crystallographica Section F - November 27, 2015 Category: Biochemistry Authors: Qiu, X. Xu, C. Chen, X. Tags: cupin domain AerE 2-carboxy-6-hydroxyoctahydroindole moiety aeruginosin research communications Source Type: research

Seeding for sirtuins: microseed matrix seeding to obtain crystals of human Sirt3 and Sirt2 suitable for soaking
Sirtuins constitute a family of NAD+-dependent enzymes that catalyse the cleavage of various acyl groups from the ∊ -amino group of lysines. They regulate a series of cellular processes and their misregulation has been implicated in various diseases, making sirtuins attractive drug targets. To date, only a few sirtuin modulators have been reported that are suitable for cellular research and their development has been hampered by a lack of structural information. In this work, microseed matrix seeding (MMS) was used to obtain crystals of human Sirt3 in its apo form and of human Sirt2 in complex with ADP ribose (ADPR). Cry...
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Rumpf, T. Gerhardt, S. Einsle, O. Jung, M. Tags: sirtuins deacylase deacetylase seeding – inhibitor complex research communications Source Type: research

Structure of Spo0M, a sporulation-control protein from Bacillus subtilis
Spo0M is a sporulation-control protein that is thought to play an essential role in the early stage of endospore formation. While little is known about the functions of Spo0M, a recent phylogenetic study suggests that, based on its amino-acid sequence, Spo0M might belong to the arrestin clan. The crystal structure of the Spo0M protein was determined at a resolution of 2.3   Å . Ten amino acids at the end of the N-terminus were removed to improve the thermal stability of the purified Spo0M protein and the crystal structure of Spo0M was determined by SAD. Spo0M has a well conserved N-terminal domain with an arrestin-...
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Sonoda, Y. Mizutani, K. Mikami, B. Tags: sporulation-control protein Spo0M arrestin FP domain PI31 research communications Source Type: research

Combining dehydration, construct optimization and improved data collection to solve the crystal structure of a CRM1 – RanGTP – SPN1 – Nup214 quaternary nuclear export complex
High conformational flexibility is an intrinsic and indispensable property of nuclear transport receptors, which makes crystallization and structure determination of macromolecular complexes containing exportins or importins particularly challenging. Here, the crystallization and structure determination of a quaternary nuclear export complex consisting of the exportin CRM1, the small GTPase Ran in its GTP-bound form, the export cargo SPN1 and an FG repeat-containing fragment of the nuclear pore complex component nucleoporin Nup214 fused to maltose-binding protein is reported. Optimization of constructs, seeding and the dev...
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Monecke, T. Dickmanns, A. Weiss, M.S. Port, S.A. Kehlenbach, R.H. Ficner, R. Tags: crystal dehydration HC1c crystal humidifier CRM1 nucleoporin maltose-binding protein research communications Source Type: research

Crystallization behaviour of glyceraldehyde dehydrogenase from Thermoplasma acidophilum
The glyceraldehyde dehydrogenase from Thermoplasma acidophilum (TaAlDH) is a microbial enzyme that catalyzes the oxidation of d-glyceraldehyde to d-glycerate in the artificial enzyme cascade designed for the conversion of glucose to the organic solvents isobutanol and ethanol. Various mutants of TaAlDH were constructed by a random approach followed by site-directed and saturation mutagenesis in order to improve the properties of the enzyme that are essential for its functioning within the cascade. Two enzyme variants, wild-type TaAlDH (TaAlDHwt) and an F34M+S405N variant (TaAlDH F34M+S405N), were successfully crystallized....
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Iermak, I. Degtjarik, O. Steffler, F. Sieber, V. Kuta Smatanova, I. Tags: glyceraldehyde dehydrogenase cell-free enzyme cascade bioproduction Thermoplasma acidophilum TaAlDH research communications Source Type: research

Expression, purification and crystallization of a plant polyketide cyclase from Cannabis sativa
Plant polyketides are a structurally diverse family of natural products. In the biosynthesis of plant polyketides, the construction of the carbocyclic scaffold is a key step in diversifying the polyketide structure. Olivetolic acid cyclase (OAC) from Cannabis sativa L. is the only known plant polyketide cyclase that catalyzes the C2 – C7 intramolecular aldol cyclization of linear pentyl tetra- β -ketide-CoA to generate olivetolic acid in the biosynthesis of cannabinoids. The enzyme is also thought to belong to the dimeric α + β barrel (DABB) protein family. However, because of a lack of functional ana...
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Yang, X. Matsui, T. Mori, T. Taura, F. Noguchi, H. Abe, I. Morita, H. Tags: cannabinoids Cannabis sativa cyclase olivetolic acid research communications Source Type: research

Incorporation of copper ions into crystals of T2 copper-depleted laccase from Botrytis aclada
Laccases belong to the class of multicopper oxidases catalyzing the oxidation of phenols accompanied by the reduction of molecular oxygen to water without the formation of hydrogen peroxide. The activity of laccases depends on the number of Cu atoms per enzyme molecule. The structure of type 2 copper-depleted laccase from Botrytis aclada has been solved previously. With the aim of obtaining the structure of the native form of the enzyme, crystals of the depleted laccase were soaked in Cu+- and Cu2+-containing solutions. Copper ions were found to be incorporated into the active site only when Cu+ was used. A comparative ana...
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Osipov, E.M. Polyakov, K.M. Tikhonova, T.V. Kittl, R. Dorovatovskii, P.V. Shleev, S.V. Popov, V.O. Ludwig, R. Tags: laccase copper(I) complex full copper content research communications Source Type: research

Heterogeneous nucleation is required for crystallization of the ZnuA domain of pneumococcal AdcA
This study demonstrates the utility of heterogeneous nucleation. The solution of the crystal structures will lead to further understanding of Zn2+ acquisition by S. pneumoniae. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Luo, Z. Morey, J.R. McDevitt, C.A. Kobe, B. Tags: heterogeneous nucleation seaweed ZnuA AdcA Zn2 Streptococcus pneumoniae solute-binding protein research communications Source Type: research

Production, crystallization and X-ray diffraction analysis of the protease CT441 from Chlamydia trachomatis
The prokaryotic obligate intracellular pathogen Chlamydia trachomatis is the most prevalent cause of preventable blindness, affecting approximately six million people worldwide. In addition, C. trachomatis is the most commonly reported sexually transmitted pathogen in Europe and the US, causing pelvic inflammation, ectopic pregnancy and infertility. As in other intracellular pathogens, proteases play crucial roles during most stages of the complex life cycle of Chlamydia. CT441 is a chlamydial protease that has been reported to interfere with oestrogen signalling of the host cell. Here, the recombinant production, purifica...
Source: Acta Crystallographica Section F - November 18, 2015 Category: Biochemistry Authors: Kohlmann, F. Shima, K. Rupp, J. Solbach, W. Hilgenfeld, R. Hansen, G. Tags: tail-specific protease Ser/Lys protease chlamydial infections research communications Source Type: research

Structure of DnmZ, a nitrososynthase in the Streptomyces peucetius anthracycline biosynthetic pathway
The anthracyclines are a class of highly effective natural product chemotherapeutics and are used to treat a range of cancers, including leukemia. The toxicity of the anthracyclines has stimulated efforts to further diversify the scaffold of the natural product, which has led to renewed interest in the biosynthetic pathway responsible for the formation and modification of this family of molecules. DnmZ is an N-hydroxylating flavin monooxygenase (a nitrososynthase) that catalyzes the oxidation of the exocyclic amine of the sugar nucleotide dTDP-l-epi-vancosamine to its nitroso form. Its specific role in the anthracycline bi...
Source: Acta Crystallographica Section F - October 1, 2015 Category: Biochemistry Authors: Sartor, L.Ibarra, C.Al-Mestarihi, A.Bachmann, B.O.Vey, J.L. Tags: flavin monooxygenase nitrososynthase N-oxidation biosynthetic pathway protein crystal structure research communications Source Type: research

1.25   Å resolution structure of an RNA 20-mer that binds to the TREX2 complex
The 1.25   Å resolution H32:R crystal structure of a 20   nt ribonucleotide that binds to the TREX-2 complex with high affinity shows a double-stranded RNA duplex arranged along a crystallographic 31 axis in which the antiparallel chains overlap by 18 nucleotides and are related by a crystallographic twofold axis. The duplex shows C – A, U – U and C – C noncanonical base pairings together with canonical Watson – Crick A – U and G – C pairs and a G – U wobble. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Valkov, E. Stewart, M. Tags: RNA noncanonical base pair research communications Source Type: research

Structure of a catalytic dimer of the α - and β -subunits of the F-ATPase from Paracoccus denitrificans at 2.3   Å resolution
The structures of F-ATPases have predominantly been determined from mitochondrial enzymes, and those of the enzymes in eubacteria have been less studied. Paracoccus denitrificans is a member of the α -proteobacteria and is related to the extinct protomitochondrion that became engulfed by the ancestor of eukaryotic cells. The P. denitrificans F-ATPase is an example of a eubacterial F-ATPase that can carry out ATP synthesis only, whereas many others can catalyse both the synthesis and the hydrolysis of ATP. Inhibition of the ATP hydrolytic activity of the P. denitrificans F-ATPase involves the ζ inhibitor protein,...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Morales-R í os, E. Montgomery, M.G. Leslie, A.G.W. Garc í a-Trejo, J.J. Walker, J.E. Tags: α -proteobacteria Paracoccus denitrificans F-ATPase structure catalytic β dimer research communications Source Type: research

An active site – tail interaction in the structure of hexahistidine-tagged Thermoplasma acidophilum citrate synthase
Citrate synthase (CS) plays a central metabolic role in aerobes and many other organisms. The CS reaction comprises two half-reactions: a Claisen aldol condensation of acetyl-CoA (AcCoA) and oxaloacetate (OAA) that forms citryl-CoA (CitCoA), and CitCoA hydrolysis. Protein conformational changes that `close' the active site play an important role in the assembly of a catalytically competent condensation active site. CS from the thermoacidophile Thermoplasma acidophilum (TpCS) possesses an endogenous Trp fluorophore that can be used to monitor the condensation reaction. The 2.2   Å resolution crystal structure of TpC...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Murphy, J.R. Donini, S. Kappock, T.J. Tags: thermophile euryarchaeon conformation change carbon – carbon bond formation research communications Source Type: research

Crystallization and preliminary X-ray diffraction studies of frutalin, an α -d-galactose-specific lectin from Artocarpus incisa seeds
Frutalin is an α -d-galactose-specific carbohydrate-binding glycoprotein with antitumour properties and is a powerful tool for tumour biomarker discovery. The crystallization and preliminary X-ray diffraction analysis of this lectin, which was isolated from Artocarpus incisa seeds, are reported here. Frutalin was purified and submitted to mass-spectrometric analysis. Diverse masses at approximately 16   kDa were observed in the deconvoluted spectra, which support the presence of isoforms. The best frutalin crystals were grown within a week in 0.1   M citric acid pH 3.5 which contained 25% PEG 3350 as a precipitan...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Monteiro-Moreira, A.C.O. D-Muniz Pereira, H. Vieira Neto, A.E. Mendes Batista Moreno, F.B. Duarte Pinto Lobo, M. Sousa, F.D. Moreira, R.A. Tags: frutalin jacalin lectin-related family Artocarpus incisa research communications Source Type: research

The structure of the death receptor 4 – TNF-related apoptosis-inducing ligand (DR4 – TRAIL) complex
The structure of death receptor 4 (DR4) in complex with TNF-related apoptosis-inducing ligand (TRAIL) has been determined at 3   Å resolution and compared with those of previously determined DR5 – TRAIL complexes. Consistent with the high sequence similarity between DR4 and DR5, the overall arrangement of the DR4 – TRAIL complex does not differ substantially from that of the DR5 – TRAIL complex. However, subtle differences are apparent. In addition, solution interaction studies were carried out that show differences in the thermodynamics of binding DR4 or DR5 with TRAIL. (Source: Acta Crystallographica Section F)
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Ramamurthy, V. Yamniuk, A.P. Lawrence, E.J. Yong, W. Schneeweis, L.A. Cheng, L. Murdock, M. Corbett, M.J. Doyle, M.L. Sheriff, S. Tags: death receptor 4 death receptor 5 TNF-related apoptosis-inducing ligand research communications Source Type: research

Extension of resolution and oligomerization-state studies of 2,4 ′ -dihydroxyacetophenone dioxygenase from Alcaligenes sp. 4HAP
The enzyme 2,4 ′ -dihydroxyacetophenone dioxygenase (DAD) catalyses the conversion of 2,4 ′ -dihydroxyacetophenone to 4-hydroxybenzoic acid and formic acid. This enzyme is a very unusual dioxygenase in that it cleaves a C — C bond in a substituent of the aromatic ring rather than within the ring itself. Whilst it has been shown that DAD is a tetramer in solution, the recently solved crystal structure of the Alcaligenes sp. 4HAP enzyme was in fact dimeric rather than tetrameric. Since the use of limited chymotrypsinolysis, which apparently results in removal of the first 20 or so N-terminal residues of DAD...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Guo, J. Erskine, P. Coker, A.R. Gor, J. Perkins, S.J. Wood, S.P. Cooper, J.B. Tags: 2,4 ′ -dihydroxyacetophenone dioxygenase protein X-ray structure oligomerization analytical ultracentrifugation gel filtration docking research communications Source Type: research

Formulation screening by differential scanning fluorimetry: how often does it work?
There is strong evidence to suggest that a protein sample needs to be well folded and uniform in order to form protein crystals, and it is accepted knowledge that the formulation can have profound effects on the behaviour of the protein sample. The technique of differential scanning fluorimetry (DSF) is a very accessible method to determine protein stability as a function of the formulation chemistry and the temperature. A diverse set of 252 soluble protein samples was subjected to a standard formulation-screening protocol using DSF. Automated analysis of the DSF results suggest that in over 35% of cases buffer screening s...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Ristic, M.Rosa, N.Seabrook, S.A.Newman, J. Tags: differential scanning fluorimetry protein stability buffer formulation Thermofluor research communications Source Type: research

A sucrose-binding site provides a lead towards an isoform-specific inhibitor of the cancer-associated enzyme carbonic anhydrase IX
Human carbonic anhydrase (CA; EC 4.2.1.1) isoform IX (CA IX) is an extracellular zinc metalloenzyme that catalyzes the reversible hydration of CO2 to HCO3−, thereby playing a role in pH regulation. The majority of normal functioning cells exhibit low-level expression of CA IX. However, in cancer cells CA IX is upregulated as a consequence of a metabolic transition known as the Warburg effect. The upregulation of CA IX for cancer progression has drawn interest in it being a potential therapeutic target. CA IX is a transmembrane protein, and its purification, yield and crystallization have proven challenging to structu...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Pinard, M.A.Aggarwal, M.Mahon, B.P.Tu, C.McKenna, R. Tags: α -carbonic anhydrase CA IX mimic isoform-specific drug design sugar approach carbonic anhydrase IX sucrose research communications Source Type: research

Stabilization of porcine pancreatic elastase crystals by glutaraldehyde cross-linking
Elastase is a serine protease from the chymotrypsin family of enzymes with the ability to degrade elastin, an important component of connective tissues. Excessive elastin proteolysis leads to a number of pathological diseases. Porcine pancreatic elastase (PPE) is often used for drug development as a model for human leukocyte elastase (HLE), with which it shares high sequence identity. Crystals of PPE were grown overnight using sodium sulfate and sodium acetate at acidic pH. Cross-linking the crystals with glutaraldehyde was needed to resist the soaking procedure with a diethyl N-(methyl)pyridinyl-substituted oxo-β-lac...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Hofbauer, S.Brito, J.A.Mulchande, J.Nogly, P.Pessanha, M.Moreira, R.Archer, M. Tags: porcine pancreatic elastase serine protease streak-seeding sensitive protein crystals glutaraldehyde cross-linking soaking of hydrophobic ligand research communications Source Type: research

Structure of the dual-function fructose-1,6/sedoheptulose-1,7-bisphosphatase from Thermosynechococcus elongatus bound with sedoheptulose-7-phosphate
The dual-function fructose-1,6/sedoheptulose-1,7-bisphosphatase (FBP/SBPase) in cyanobacteria carries out two activities in the Calvin cycle. Structures of this enzyme from the cyanobacterium Synechocystis sp. PCC 6803 exist, but only with adenosine monophosphate (AMP) or fructose-1,6-bisphosphate and AMP bound. The mechanisms which control both selectivity between the two sugars and the structural mechanisms for redox control are still unresolved. Here, the structure of the dual-function FBP/SBPase from the thermophilic cyanobacterium Thermosynechococcus elongatus is presented with sedoheptulose-7-phosphate bound and in t...
Source: Acta Crystallographica Section F - September 23, 2015 Category: Biochemistry Authors: Cotton, C.A.R.Kabasakal, B.V.Miah, N.A.Murray, J.W. Tags: fructose-1,6/sedoheptulose-1,7-bisphosphatase Calvin cycle cyanobacteria photosynthesis research communications Source Type: research