PMCC PMCC

Search tips
Search criteria

Advanced
Results 1-25 (1064627)

Clipboard (0)
None

Related Articles

1.  Effect of Algae and Plant Lectins on Planktonic Growth and Biofilm Formation in Clinically Relevant Bacteria and Yeasts 
BioMed Research International  2014;2014:365272.
This study aimed to evaluate the abilities of plant and algae lectins to inhibit planktonic growth and biofilm formation in bacteria and yeasts. Initially, ten lectins were tested on Staphylococcus epidermidis, Staphylococcus aureus, Klebsiella oxytoca, Pseudomonas aeruginosa, Candida albicans, and C. tropicalis at concentrations of 31.25 to 250 μg/mL. The lectins from Cratylia floribunda (CFL), Vatairea macrocarpa (VML), Bauhinia bauhinioides (BBL), Bryothamnion seaforthii (BSL), and Hypnea musciformis (HML) showed activities against at least one microorganism. Biofilm formation in the presence of the lectins was also evaluated; after 24 h of incubation with the lectins, the biofilms were analyzed by quantifying the biomass (by crystal violet staining) and by enumerating the viable cells (colony-forming units). The lectins reduced the biofilm biomass and/or the number of viable cells to differing degrees depending on the microorganism tested, demonstrating the different characteristics of the lectins. These findings indicate that the lectins tested in this study may be natural alternative antimicrobial agents; however, further studies are required to better elucidate the functional use of these proteins.
doi:10.1155/2014/365272
PMCID: PMC4058506  PMID: 24982871
2.  Purification, crystallization and preliminary X-ray crystallographic analysis of rice lectin from Oryza sativa  
Rice lectin was crystallized and analyzed by X-ray crystallography.
Lectins with sugar-binding specificity are widely distributed in higher plants and various other species. The expression of rice lectin from Oryza sativa is up-regulated in the growing coleoptile when anaerobic stress persists. A rice lectin of molecular weight 15.2 kDa has been crystallized using the hanging-drop vapour-diffusion method. From the diffraction of the lectin crystals at 1.93 Å resolution, the unit cell belongs to space group P31, with unit-cell parameters a = 98.58, b = 98.58, c = 44.72 Å. Preliminary analysis indicates that there are two lectin molecules in an asymmetric unit with a large solvent content, 70.1%.
doi:10.1107/S1744309105040698
PMCID: PMC2150942  PMID: 16511272
lectins; rice
3.  Expression, purification, crystallization and preliminary X-ray analysis of the polysaccharide lyase RB5312 from the marine planctomycete Rhodopirellula baltica  
This study describes the crystallization and preliminary X-ray analysis of the family PL1 polysaccharide lyase RB5312 from the marine bacterium R. baltica. Purified recombinant protein was crystallized; the crystals belonged to space group P212121 and diffracted X-rays to a resolution of 1.8 Å.
Polysaccharide lyases belonging to family PL1 act on pectins. These anionic polymers are usually produced by terrestrial plants and therefore pectinolytic enzymes are not frequently observed in marine microorganisms. The protein RB5312 from the marine bacterium Rhodopirellula baltica is distantly related to family PL1 pectate lyases, but its exact function is unclear. In this study, the expression and purification of a recombinant form of RB5312 are described. This protein was crystallized using the hanging-drop vapour-diffusion method. The crystals belongs to space group P212121, with unit-cell parameters a = 39.05, b = 144.05, c = 153.97 Å, α = β = γ = 90°. A complete data set was collected to 1.8 Å resolution from a native crystal.
doi:10.1107/S1744309108004387
PMCID: PMC2374145  PMID: 18323615
marine bacteria; pectins; family PL1
4.  Expression, purification and preliminary X-ray diffraction analysis of the catalytic module of a β-­agarase from the flavobacterium Zobellia galactanivorans  
A novel family GH16 β-agarase from the marine bacterium Zobellia galactanivorans was expressed, purified and crystallized. Hexagonal crystals belonging to space group P3121 diffracted to 2.2 Å resolution, whereas orthorhombic crystals belonging to space group P212121 diffracted to 1.5–Å resolution.
Marine bacteria secrete specific glycoside hydrolases such as agarases to access polysaccharides from algal cell walls as a carbon and energy source. In an attempt to identify agarases with variable degradation patterns, a novel family GH16 β-agarase from the marine bacterium Zobellia galactanivorans was expressed, purified and crystallized. The purified enzyme crystallized in two distinct forms that were grown by the hanging-drop vapour-diffusion method using polyethylene glycol as a precipitant. Hexagonal crystals belonging to space group P3121 diffracted to 2.2 Å resolution, whereas orthorhombic crystals belonging to space group P212121 diffracted to 1.5 Å resolution.
doi:10.1107/S174430911000429X
PMCID: PMC2852333  PMID: 20383011
β-agarases; glycoside hydrolases; heterologous expression; Zobellia galactanivorans
5.  Crystallization and preliminary X-ray diffraction analysis of the lectin from Dioclea rostrata Benth seeds 
D. rostrata lectin was crystallized by hanging-drop vapor diffusion. The crystal belongs to the orthorhombic space group I222 and diffracted to 1.87 Å resolution.
Lectins from the Diocleinae subtribe (Leguminosae) are highly similar proteins that promote various biological activities with distinctly differing potencies. The structural basis for this experimental data is not yet fully understood. Dioclea rostrata lectin was purified and crystallized by hanging-drop vapour diffusion at 293 K. The crystal belongs to the orthorhombic space group I222, with unit-cell parameters a = 61.51, b = 88.22, c = 87.76 Å.  Assuming the presence of one monomer per asymmetric unit, the solvent content was estimated to be about 47.9%. A complete data set was collected at 1.87 Å resolution.
doi:10.1107/S1744309106001801
PMCID: PMC2150952  PMID: 16511292
lectins; Dioclea rostrata
6.  Crystallization and preliminary X-ray diffraction analysis of the Fab fragment of WO2, an antibody specific for the Aβ peptides associated with Alzheimer’s disease 
Crystallization and X-ray diffraction data collection of the Fab fragment of the monoclonal antibody WO2 in the absence or presence of amyloid β peptides associated with Alzheimer’s disease are reported.
The murine monoclonal antibody WO2 specifically binds the N-terminal region of the amyloid β peptide (Aβ) associated with Alzheimer’s disease. This region of Aβ has been shown to be the immunodominant B-cell epitope of the peptide and hence is considered to be a basis for the development of immunotherapeutic strategies against this prevalent cause of dementia. Structural studies have been undertaken in order to characterize the molecular basis for antibody recognition of this important epitope. Here, details of the crystallization and X-ray analysis of the Fab fragment of the unliganded WO2 antibody in two crystal forms and of the complexes that it forms with the truncated Aβ peptides Aβ1–16 and Aβ1–28 are presented. These crystals were all obtained using the hanging-drop vapour-diffusion method at 295 K. Crystals of WO2 Fab were grown in polyethylene glycol solutions containing ZnSO4; they belonged to the orthorhombic space group P212121 and diffracted to 1.6 Å resolution. The complexes of WO2 Fab with either Aβ1–­16 or Aβ1–28 were cocrystallized from polyethylene glycol solutions. These two complex crystals grew in the same space group, P212121, and diffracted to 1.6 Å resolution. A second crystal form of WO2 Fab was grown in the presence of the sparingly soluble Aβ1–42 in PEG 550 MME. This second form belonged to space group P21 and diffracted to 1.9 Å resolution.
doi:10.1107/S1744309108011718
PMCID: PMC2376392  PMID: 18453721
Aβ peptides; Alzheimer’s disease; antibody Fab fragments; immunotherapy
7.  Crystallization and preliminary X-ray crystallographic analysis of a helicase-like domain from a tomato mosaic virus replication protein 
Crystals of the helicase domain from a tomato mosaic virus replication protein obtained using the hanging-drop vapour-diffusion method at 285 K diffracted X-rays to 2.05 Å resolution. They belonged to the orthorhombic space group P212121, with unit-cell parameters a = 85.8, b = 128.3, c = 40.7 Å.
Tomato mosaic virus belongs to the genus Tobamovirus in the alphavirus-like superfamily of positive-strand RNA viruses. The alphavirus-like superfamily includes many plant and animal viruses of agronomical and clinical importance. These viruses encode replication-associated proteins that contain a putative superfamily 1 helicase domain. No three-dimensional structures for this domain have been determined to date. Here, the crystallization and preliminary X-ray diffraction analysis of the 130K helicase domain are reported. Diffraction data were collected and processed to 2.05 and 1.75 Å resolution from native and selenomethionine-labelled crystals, respectively. The crystals belonged to the orthorhombic space group P212121, with unit-cell parameters a = 85.8, b = 128.3, c = 40.7 Å.
doi:10.1107/S174430911104231X
PMCID: PMC3232162  PMID: 22139189
tomato mosaic virus; replicase protein; helicase domain
8.  Crystallization and preliminary X-ray crystallographic studies of an exo-β-d-glucosaminidase from Trichoderma reesei  
An exo-β-d-glucosaminidase from T. reesei (Gls93) has been crystallized by the hanging-drop vapour-diffusion method. Diffraction data have been collected using synchrotron radiation.
Chitosan is degraded to glucosamine (GlcN) by chitosanase and exo-β-d-glucosaminidase (GlcNase). GlcNase from Trichoderma reesei (Gls93) is a 93 kDa extracellular protein composed of 892 amino acids. The enzyme liberates GlcN from the nonreducing end of the chitosan chain in an exo-type manner and belongs to glycoside hydrolase family 2. For crystallographic investigations, Gls93 was overexpressed in Pichia pastoris cells. The recombinant Gls93 had two molecular forms of ∼105 kDa (Gls93-F1) and ∼100 kDa (Gls93-F2), with the difference between them being caused by N-glycosylation. Both forms were crystallized by the hanging-drop vapour-diffusion method. Crystals of Gls93-F1 belonged to the orthorhombic space group P212121, with unit-cell parameters a = 98.27, b = 98.42, c = 108.28 Å, and diffracted to 1.8 Å resolution. Crystals of Gls93-F2 belonged to the ortho­rhombic space group P212121, with unit-cell parameters a = 67.84, b = 81.62, c = 183.14 Å, and diffracted to 2.4 Å resolution. Both crystal forms were suitable for X-ray structure analysis at high resolution.
doi:10.1107/S1744309110000606
PMCID: PMC2833044  PMID: 20208168
exo-β-d-glucosaminidases; exochitosanases; Gls93; Trichoderma reesei
9.  Larvicidal Activity against Aedes aegypti and Molluscicidal Activity against Biomphalaria glabrata of Brazilian Marine Algae 
This study investigated the biological activities of five benthic marine algae collected from Northeastern Region of Brazil. The tested activities included larvicidal activity against Aedes aegypti, molluscicidal activity against Biomphalaria glabrata, and toxicity against Artemia salina. Extracts of Ulva lactuca (Chlorophyta), Padina gymnospora, Sargassum vulgare (Phaeophyta), Hypnea musciformis, and Digenea simplex (Rhodophyta) were prepared using different solvents of increasing polarity, including dichloromethane, methanol, ethanol, and water. Of the extracts screened, the dichloromethane extracts of H. musciformis and P. gymnospora exhibited the highest activities and were subjected to bioassay-guided fractionation in hexane and chloroform. The chloroform fractions of the P. gymnospora and H. musciformis extracts showed molluscicidal activity at values below 40 μg·mL−1 (11.1460 μg·mL−1 and 25.8689 μg·mL−1, resp.), and the chloroform and hexane fractions of P. gymnospora showed larvicidal activity at values below 40 μg·mL−1 (29.018 μg·mL−1 and 17.230 μg·mL−1, resp.). The crude extracts were not toxic to A. salina, whereas the chloroform and hexane fractions of P. gymnospora (788.277 μg·mL−1 and 706.990 μg·mL−1) showed moderate toxicity, indicating that the toxic compounds present in these algae are nonpolar.
doi:10.1155/2014/501328
PMCID: PMC3943412  PMID: 24688787
10.  Crystallization and preliminary X-ray analysis of ZHE1, a hatching enzyme from the zebrafish Danio rerio  
The hatching enzyme of zebrafish, ZHE1, was expressed, purified and crystallized using the hanging-drop vapour-diffusion method. The crystal belonged to space group P212121 and diffracted X-rays to a resolution of 1.14 Å.
The hatching enzyme of the zebrafish, ZHE1 (29.3 kDa), is a zinc metallo­protease that catalyzes digestion of the egg envelope (chorion). ZHE1 was heterologously expressed in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion method using PEG 3350 as the precipitant. Two diffraction data sets with resolution ranges 50.0–1.80 and 50.0–1.14 Å were independently collected from two crystals and were merged to give a highly complete data set over the full resolution range 50.0–1.14 Å. The space group was assigned as primitive orthorhombic P212121, with unit-cell parameters a = 32.9, b = 62.5, c = 87.4 Å. The crystal contained one ZHE1 molecule in the asymmetric unit.
doi:10.1107/S1744309109033016
PMCID: PMC2765890  PMID: 19851011
ZHE1; hatching enzymes; zebrafish
11.  Crystallization and preliminary X-ray crystallographic analysis of a galactose-specific lectin from Dolichos lablab  
The galactose-specific lectin from the seeds of a leguminous plant, D. lablab, has been crystallized. Molecular-replacement solution using 3.0 Å X-ray diffraction data showed the lectin to be a tetramer.
The galactose-specific lectin from the seeds of Dolichos lablab has been crystallized using the hanging-drop vapour-diffusion technique. The crystals belong to space group P1, with unit-cell parameters a = 73.99, b = 84.13, c = 93.15 Å, α = 89.92, β = 76.01, γ = 76.99°. X-ray diffraction data to a resolution of 3.0 Å have been collected under cryoconditions (100 K) using a MAR imaging-plate detector system mounted on a rotating-anode X-ray generator. Molecular-replacement calculations carried out using the available structures of legume lectins as search models revealed that the galactose-specific lectin from D. lablab forms a tetramer similar to soybean agglutinin; two such tetramers are present in the asymmetric unit.
doi:10.1107/S1744309106001448
PMCID: PMC2150945  PMID: 16511291
Dolichos lablab; galactose-specific lectins; legume lectins
12.  Purification, crystallization and preliminary X-ray characterization of a haemagglutinin from the seeds of Jatropha curcas  
A novel haemagglutinin from Jatropha curcas seeds is purified and crystallized. X-ray diffraction data collected from the rod-shaped crystals were processed in the orthorhombic space group P212121 and the crystals diffracted to 2.8 Å resolution at 103 K.
The plant Jatropha curcas (Euphorbiaceae) is an important source of biofuel from the inedible oil present in its toxic seeds. The toxicity arises from the presence of curcin, a ribosome-inactivating protein showing haemagglutination activity. In this communication, the purification, crystallization and preliminary X-ray characterization are reported of a small protein isolated from J. curcas seeds with a molecular mass of ∼10 kDa that agglutinates rabbit erythrocytes. The protein was crystallized using the hanging-drop vapour-diffusion method and also by the microbatch method in 72-well HLA plates, using PEG 8000 as the precipitant in both conditions. X-ray diffraction data collected from the rod-shaped crystals were processed in the orthorhombic space group P212121. The crystals diffracted to 2.8 Å resolution at 103 K.
doi:10.1107/S1744309111038218
PMCID: PMC3232132  PMID: 22139159
Jatropha curcas; haemagglutinins
13.  Crystallization and preliminary X-ray diffraction studies of trypsin-like proteases from the gastric fluid of the marine crab Cancer pagurus  
Two trypsins from the gastric fluid of the marine crab C. pagurus were purified and crystallized and X-ray data were collected to 0.97 and 3.2 Å resolution.
The digestive fluid of the marine crab Cancer pagurus (Decapoda, Brachyura) contains highly stable proteases which display enhanced activity in aqueous mixtures of organic solvents. Three trypsins were isolated from the gastric fluid and two of them, C.p.TryII and C.p.TryIII, were purified to homogeneity by anion-exchange chromatography and crystallized by hanging-drop vapour diffusion. Diffraction data were collected at a synchrotron to 0.97 and 3.2 Å resolution, respectively. The crystal of C.p.TryII belongs to the orthorhombic space group P212121, with unit-cell parameters a = 52.06, b = 62.00, c = 71.66 Å. Based on the Matthews coefficient, one protein molecule per asymmetric unit is suggested. In contrast, crystals of C.p.TryIII, which belong to the cubic space group P213 with unit-cell parameters a = b = c = 215.4 Å, are assumed to contain 12 molecules per asymmetric unit.
doi:10.1107/S1744309107008524
PMCID: PMC2330177  PMID: 17329824
Crustacea; crabs; Cancer pagurus; gastric fluid; digestive enzymes; trypsin
14.  Crystallization and preliminary X-ray diffraction studies of vitamin D3 hydroxylase, a novel cytochrome P450 isolated from Pseudonocardia autotrophica  
The purification, crystallization and preliminary X-ray diffraction studies of vitamin D3 hydroxylase isolated from P. autotrophica are reported.
Vitamin D3 hydroxylase (Vdh) is a novel cytochrome P450 monooxygenase isolated from the actinomycete Pseudonocardia autotrophica and consisting of 403 amino-acid residues. Vdh catalyzes the activation of vitamin D3 via sequential hydroxylation reactions: these reactions involve the conversion of vitamin D3 (cholecalciferol or VD3) to 25-hydroxyvitamin D3 [25(OH)VD3] and the subsequent conversion of 25(OH)VD3 to 1α,25-dihydroxyvitamin D3 [calciferol or 1α,25(OH)2VD3]. Overexpression of recombinant Vdh was carried out using a Rhodococcus erythropolis expression system and the protein was subsequently purified and crystallized. Two different crystal forms were obtained by the hanging-drop vapour-diffusion method at 293 K using polyethylene glycol as a precipitant. The form I crystal belonged to the trigonal space group P31, with unit-cell parameters a = b = 61.7, c = 98.8 Å. There is one Vdh molecule in the asymmetric unit, with a solvent content of 47.6%. The form II crystal was grown in the presence of 25(OH)VD3 and belonged to the orthorhombic system P212121, with unit-cell parameters a = 63.4, b = 65.6 c = 102.2 Å. There is one Vdh molecule in the asymmetric unit, with a solvent content of 46.7%. Native data sets were collected to resolutions of 1.75 and 3.05 Å for form I and form II crystals, respectively, using synchrotron radiation. The structure solution was obtained by the molecular-replacement method and model refinement is in progress for the form I crystal.
doi:10.1107/S1744309109007829
PMCID: PMC2664763  PMID: 19342783
cholecalciferol; CYP107; cytochrome P450 monooxygenase; hydroxylation; Pseudonocardia autotrophica; vitamin D3
15.  Crystallization and preliminary X-ray diffraction analysis of a new chitin-binding protein from Parkia platycephala seeds 
Crystals of P. platycephala chintinase/lectin (PPL-2) belong to the orthorhombic space group P212121, with unit-cell parameters a = 55.19, b = 59.95, c = 76.60 Å. The preliminary cystal structure of PPL-2 was solved at a resolution of 1.73 Å by molecular replacement, presenting a correlation coefficient of 0.558 and an R factor of 0.439.
A chitin-binding protein named PPL-2 was purified from Parkia platycephala seeds and crystallized. Crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 55.19, b = 59.95, c = 76.60 Å, and grew over several days at 293 K using the hanging-drop method. Using synchrotron radiation, a complete structural data set was collected to 1.73 Å resolution. The preliminary crystal structure of PPL-2, determined by molecular replacement, presents a correlation coefficient of 0.558 and an R factor of 0.439. Crystallographic refinement is in progress.
doi:10.1107/S1744309105024462
PMCID: PMC1978108  PMID: 16511174
chitin-binding proteins; chitinases; Parkia platycephala; lectins
16.  Purification, crystallization and preliminary X-ray analysis of haemoglobin from ostrich (Struthio camelus) 
Purification, crystallization and preliminary X-ray analysis of haemoglobin from ostrich (Struthio camelus) has been carried out under 293 K temperature conditions. The ostrich is a large flightless bird which contains inositol tetrakisphosphate in erythrocytes and its whole blood oxygen affinity is higher. Efforts have been made to explore the structure–function relationship of ostrich heamoglobin.
Haemoglobin is a tetrameric protein that carries oxygen from the lungs to tissues and carbon dioxide from tissues back to the lungs. The oxygen-binding properties of haemoglobin are regulated through the binding of allosteric effectors. The respiratory system of avian species is unique and complex in nature when compared with that of mammals. In avian species, inositol pentaphosphate (inositol-P5) is present in the erythrocytes of the adult and is thought to be the major factor responsible for the relatively high oxygen affinity of the whole blood. The ostrich (Struthio camelus) is a large flightless bird which contains inositol tetrakisphosphate (inositol-P4) in its erythrocytes and its whole blood oxygen affinity is higher. Efforts have been made to explore the structure–function relationship of ostrich haemoglobin. Ostrich haemoglobin was purified using ion-exchange chromatography. Haemoglobin crystals were grown by the hanging-drop vapour-diffusion method using PEG 3350 as the precipitant in 50 mM phosphate buffer pH 7.2. Data were collected using a MAR345 image-plate detector system. The crystals of ostrich haemo­globin diffracted to 2.2 Å resolution. They belonged to the ortho­rhombic space group P212121 with one whole biological molecule in the asymmetric unit; the unit-cell parameters were a = 80.93, b = 81.68, c = 102.05 Å.
doi:10.1107/S1744309109019009
PMCID: PMC2705633  PMID: 19574638
haemoglobin; Struthio camelus
17.  Cloning, overexpression, purification, crystallization, and preliminary X-ray studies of SP_0149, the substrate binding protein of an ABC transporter from Streptococcus pneumoniae  
The substrate binding protein (SP_0149) of an ABC transporter from Streptococcus pneumoniae was molecularly cloned, overexpressed and purified. Diffraction quality crystals were grown using the hanging-drop vapour-diffusion technique.
A truncated (29 residues from the N-terminus) and N-terminal His-tagged form of SP_0149 from pneumococcal strain ATCC BAA-334 was overexpressed and purified to homogeneity using affinity and gel-filtration chromatography. Diffraction quality crystals were grown at 293 K using the hanging-drop vapour-diffusion technique. X-ray diffraction data were collected to 2.3 Å resolution from a single-crystal that belonged to the orthorhombic space group P212121 with the unit-cell parameters a = 54.56, b = 75.61, c = 75.52 Å. The calculated values of the Matthews coefficient assuming one molecule (with calculated molecular weight of 30 400 Da) in the crystal asymmetric unit and the corresponding solvent content were 2.56 Å3 Da−1 and 52.0%, respectively.
doi:10.1107/S1744309111018422
PMCID: PMC3144799  PMID: 21795797
ABC transporter; Streptococcus pneumoniae; SP_0149; ATCC BAA-334
18.  Crystallization and X-ray diffraction analysis of human CLEC-2 
Recombinant human CLEC-2 was crystallized in the orthorhombic space group P212121 and X-ray diffraction data were collected to 2.0 Å.
The human C-type lectin-like protein CLEC-2 has recently been shown to be expressed on the surface of platelets and to function as a receptor for the snake-venom protein rhodocytin. The C-type lectin-like domain (CTLD) of CLEC-2 was expressed in Escherichia coli, refolded and purified. Crystals of this recombinant CLEC-2 were grown by sitting-drop vapour diffusion using polyethylene glycol (PEG) 6000 as a precipitant. After optimization, crystals were grown which diffracted to 2.0 Å using in-house radiation (λ = 1.5418 Å). These crystals belonged to the orthorhombic space group P212121, with unit-cell parameters a = 35.407, b = 55.143, c = 56.078 Å. The presence of one molecule per asymmetric unit is consistent with a crystal volume per unit weight (V M) of 1.82 Å3 Da−1 and a solvent content of 32.6%. These results suggest that crystals producing diffraction of this quality will be suitable for the structural determination of human CLEC-2.
doi:10.1107/S1744309105037991
PMCID: PMC1978148  PMID: 16511244
CLEC-2; CLEC1B; rhodocytin; aggretin; C-type lectins; platelets; thrombosis
19.  Crystallization and preliminary X-ray diffraction analysis of a galactose-specific lectin from the seeds of Butea monosperma  
A galactose specific lectin was purified from the seeds of a tropical tree, Butea monosperma. Its X-ray structure was solved by molecular replacement.
The galactose-specific lectin from the seeds of Butea monosperma has been crystallized by the hanging-drop vapour-diffusion technique. The crystals belonged to space group P1, with unit-cell parameters a = 78.45, b = 78.91, c = 101.85 Å, α = 74.30, β = 76.65, γ = 86.88°. X-ray diffraction data were collected to a resolution of 2.44 Å under cryoconditions (100 K) using a MAR image-plate detector system mounted on a rotating-anode X-ray generator. Molecular-replacement calculations carried out using the coordinates of several structures of legume lectins as search models indicate that the galactose-specific lectin from B. monosperma forms an octamer.
doi:10.1107/S1744309111006853
PMCID: PMC3080167  PMID: 21505258
galactose-specific lectin; Butea monosperma
20.  Crystallization and preliminary X-ray diffraction analysis of the lectin from Canavalia boliviana Piper seeds 
Canavalia boliviana lectin (Cbol) was purified using a Sephadex G-50 column and crystallized in the presence of X-Man by hanging-drop vapour diffusion at 293 K. After optimization, crystals suitable for diffraction were obtained using 0.1 M HEPES pH 7.5 and 3.0 M sodium formate.
Plant lectins are the most studied group of carbohydrate-binding proteins. Despite the high similarity between the members of the Diocleinae subtribe (Leguminosae) group, they present differing biological activities. Canavalia boliviana lectin (Cbol) was purified using a Sephadex G-50 column and crystallized in the presence of X-Man by hanging-drop vapour diffusion at 293 K. After optimization, crystals suitable for diffraction were obtained under the condition 0.1 M HEPES pH 7.5 and 3.0 M sodium formate. The crystal belonged to the monoclinic space group C2, with unit-cell parameters a = 126.70, b = 66.64, c = 64.99 Å, α = 90.0, β = 120.8, γ = 90.0°. Assuming the presence of a dimer in the asymmetric unit, the solvent content was estimated to be about 46%. A complete data set was collected at 1.5 Å resolution.
doi:10.1107/S1744309109000797
PMCID: PMC2650465  PMID: 19255467
lectins; Canavalia boliviana Piper
21.  Crystallization and preliminary crystallographic analysis of cyanide-insensitive alternative oxidase from Trypanosoma brucei brucei  
Cyanide-insensitive alternative oxidase from T. brucei brucei has been purified and crystallized for X-ray structure analysis.
Cyanide-insensitive alternative oxidase (AOX) is a mitochondrial membrane protein and a non-proton-pumping ubiquinol oxidase that catalyzes the four-electron reduction of dioxygen to water. In the African trypanosomes, tryp­anosome alternative oxidase (TAO) functions as a cytochrome-independent terminal oxidase that is essential for survival in the mammalian host; hence, the enzyme is considered to be a promising drug target for the treatment of trypanosomiasis. In the present study, recombinant TAO (rTAO) overexpressed in haem-deficient Escherichia coli was purified and crystallized at 293 K by the hanging-drop vapour-diffusion method using polyethylene glycol 400 as a precipitant. X-ray diffraction data were collected at 100 K and were processed to 2.9 Å resolution with 93.1% completeness and an overall R merge of 9.5%. The TAO crystals belonged to the orthorhombic space group I222 or I212121, with unit-cell parameters a = 63.11, b = 136.44, c = 223.06 Å. Assuming the presence of two rTAO molecules in the asymmetric unit (2 × 38 kDa), the calculated Matthews coefficient (V M) was 3.2 Å3 Da−1, which corresponds to a solvent content of 61.0%. This is the first report of a crystal of the membrane-bound diiron proteins, which include AOXs.
doi:10.1107/S1744309109054062
PMCID: PMC2833035  PMID: 20208159
alternative oxidases; membrane proteins; Trypanosoma brucei brucei; ascofuranone
22.  Crystallization and preliminary X-ray analysis of human S100A13 
Human S100A13 protein was cloned, expressed, purified and crystallized by the hanging-drop vapour-diffusion method. The crystals obtained belonged to space group P212121 and diffracted to a resolution of 1.8 Å.
S100A13 is a member of the S100 family of EF-hand-containing calcium-binding proteins and plays an important role in the secretion of fibroblast growth factor-1 and interleukin 1α, two pro-angiogenic factors released by the endoplasmic reticulum/Golgi-independent non-classical secretory pathway. Human S100A13 was heterologously expressed in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion method using PEG 3350 as the precipitant. The crystals diffracted X-rays from a synchrotron-radiation source to 1.8 Å resolution and the space group was assigned as primitive orthorhombic P212121.
doi:10.1107/S1744309106042473
PMCID: PMC2225202  PMID: 17077500
S100A13; EF-hand calcium-binding proteins
23.  Expression, crystallization and preliminary X-ray analysis of a ferric binding protein from Thermus thermophilus HB8 
The cloning, expression, purification, crystallization and preliminary X-ray analysis of a ferric binding protein encoded by T. thermophilus HB8 in apo and iron-bound holo states are presented. Four different crystal forms were obtained.
A ferric binding protein from Thermus thermophilus HB8 (TtFbpA) was expressed, purified and crystallized using the hanging-drop vapour-diffusion method. Four different crystal forms were obtained and characterized by X-ray diffraction. Two crystal forms with TtFbpA in the apo state belonged to the orthorhombic space group P212121 (unit-cell parameters a = 42.1, b = 139.3, c = 326.5 Å and a = 42.1, b = 139.3, c = 218.9 Å). The third form with TtFbpA also in the apo state belonged to the monoclinic space group P21 (unit-cell parameters a = 66.5, b = 61.7, c = 73.9 Å, β = 111.7°). The fourth form, with TtFbpA in the iron-bound holo state as confirmed by an atomic absorption spectrophotometry assay, belonged to the trigonal space group P3121 or P3221 (unit-cell parameters a = 63.6, b = 63.6, c = 266.7 Å, α = β = 90.0, γ = 120.0°).
doi:10.1107/S1744309111012929
PMCID: PMC3107153  PMID: 21636922
ferric binding protein; Thermus thermophilus HB8; apo state; iron-bound holo state
24.  Cloning, expression, purification, crystallization and preliminary X-ray diffraction studies of a major group 7 allergen, Der f 7, from the dust mite Dermatophagoides farinae  
The major group 7 allergen, Der f 7, from the dust mite Dermatophagoides farinae has been crystallized and diffracted X-rays to a resolution of 2.24 Å.
Der f 7 is a major group 7 allergen from the dust mite Dermatophagoides farinae that shows 86% sequence identity to the homologous allergen Der p 7 from D. pteronyssinus. Der f 7 was successfully overexpressed in an Escherichia coli expression system and purified to homogeneity using Ni–NTA affinity and size-exclusion column chromatography. SeMet-labelled Der f 7 was crystallized by the hanging-drop vapour-diffusion method using a reservoir solution consisting of 0.1 M bis-tris pH 7.4 and 28% polyethylene glycol monomethyl ether 2000 at 293 K. X-ray diffraction data were collected to 2.24 Å resolution using synchrotron radiation. The crystals belonged to the orthorhombic system, space group P212121, with unit-cell parameters a = 50.19, b = 58.67, c = 123.81 Å. Based on the estimated Matthews coefficient (2.16 Å3 Da−1), two molecules of Der f 7 could be present in the asymmetric unit of the crystal lattice.
doi:10.1107/S174430911103836X
PMCID: PMC3232152  PMID: 22139179
group 7 allergens; Der f 7; Dermatophagoides farinae
25.  Crystallization and preliminary X-ray diffraction studies of a ferredoxin reductase component of carbazole 1,9a-dioxygenase from Novosphingobium sp. KA1 
The ferredoxin reductase component of carbazole 1,9a-dioxygenase (Red) is involved in electron transfer from NAD(P)H to ferredoxin. The class IIA Red from Novosphingobium sp. KA1 was crystallized and the crystal diffracted to a resolution of 1.58 Å.
Carbazole 1,9a-dioxygenase (CARDO) is the initial enzyme of the carbazole-degradation pathway. The CARDO of Novosphingobium sp. KA1 consists of a terminal oxygenase, a putidaredoxin-type ferredoxin and a ferredoxin-NADH oxidoreductase (Red) and is classified as a class IIA Rieske oxygenase. Red from KA1 was crystallized at 278 K by the hanging-drop vapour-diffusion method using PEG 4000. The crystal diffracted to 1.58 Å resolution and belonged to space group P32, with unit-cell parameters a = b = 92.2, c = 78.6 Å, α = γ = 90, β = 120°. Preliminary analysis of the X-ray diffraction data revealed that the asymmetric unit contained two Red monomers. The crystal appeared to be a merohedral twin, with a twin fraction of 0.32 and twin law (−h, −k, l).
doi:10.1107/S1744309110014491
PMCID: PMC2882777  PMID: 20516607
carbazole; Rieske nonhaem iron oxygenases; ferredoxin reductases

Results 1-25 (1064627)