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Preliminary X-ray Crystallographic Analysis (preliminary + x-ray_crystallographic_analysis)
Selected AbstractsPreliminary X-ray crystallographic analysis of a soluble form of MntC, a periplasmic manganese-binding component of an ABC-type Mn transporter from Synechocystis sp.ACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2002PCC 680 Manganese is recruited in microorganisms by way of ABC-type transporter systems. Here, the expression, purification and preliminary crystallographic analysis of a soluble form of the MntC solute-binding protein component of the MntABC manganese-import system from the cyanobacterium Synechococystis sp. PCC 6803 is reported. The protein (321 amino-acid residues) was expressed exclusively in inclusion bodies, which required unfolding and refolding in the presence of manganese prior to purification. The purified protein was crystallized in the presence of PEG and zinc. The crystals belong to space group P6222, with unit-cell parameters a = b = 128.1, c = 90.0,Å and a single molecule in the asymmetric unit. The crystals diffract to 2.6,Å under cryoconditions using synchrotron radiation. [source] Preliminary X-ray crystallographic analysis of the d -xylulose 5-phosphate phosphoketolase from Lactococcus lactisACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2010Georgiana Petrareanu Phosphoketolases are thiamine diphosphate-dependent enzymes which play a central role in the pentose-phosphate pathway of heterofermentative lactic acid bacteria. They belong to the family of aldehyde-lyases and in the presence of phosphate ion cleave the carbon,carbon bond of the specific substrate d -xylulose 5-phosphate (or d -fructose 6-phosphate) to give acetyl phosphate and d -glyceraldehyde 3-phosphate (or d -erythrose 4-phosphate). Structural information about phosphoketolases is particularly important in order to fully understand their mechanism as well as the steric course of phosphoketolase-catalyzed reactions. Here, the purification, preliminary crystallization and crystallographic characterization of d -xylulose 5-phosphate phosphoketolase from Lactococcus lactis are reported. The presence of thiamine diphosphate during purification was essential for the enzymatic activity of the purified protein. The crystals belonged to the monoclinic space group P21. Diffraction data were obtained to a resolution of 2.2,Å. [source] Preliminary X-ray crystallographic analysis of SMU.2055 protein from the caries pathogen Streptococcus mutansACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 5 2010Wang-Hong Zhao The SMU.2055 gene from the major caries pathogen Streptococcus mutans is annotated as a putative acetyltransferase with 163 amino-acid residues. In order to identify its function via structural studies, the SMU.2055 gene was cloned into the expression vector pET28a. Native and SeMet-labelled SMU.2055 proteins with a His6 tag at the N-terminus were expressed at a high level in Escherichia coli strain BL21 (DE3) and purified to homogeneity by Ni2+ -chelating affinity chromatography. Diffraction-quality crystals of SeMet-labelled SMU.2055 were obtained using the sitting-drop vapour-diffusion method and diffracted to a resolution of 2.5,Å on beamline BL17A at the Photon Factory, Tsukuba, Japan. The crystals belong to the orthorhombic space group C2221, with unit-cell parameters a = 92.0, b = 95.0, c = 192.2,Å. The asymmetric unit contained four molecules, with a solvent content of 57.1%. [source] Preliminary X-ray crystallographic analysis of the breakage,reunion domain of the GyrA subunit of DNA gyrase from Colwellia psychrerythraea strain 34HACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2010Ha Yun Jung DNA gyrase is a type II topoisomerase that is essential for chromosome segregation and cell division owing to its ability to modify the topological forms of bacterial DNA. In this study, the N-terminal breakage,reunion domain of the GyrA subunit of DNA gyrase from Colwellia psychrerythraea 34H was overexpressed in Escherichia coli, purified and crystallized. Diffraction data were collected to 2.60,Å resolution using a synchrotron-radiation source. The crystal belonged to space group P212121, with unit-cell parameters a = 98.98, b = 101.56, c = 141.83,Å. The asymmetric unit contained two molecules, with a corresponding VM of 3.18,Å3,Da,1 and a solvent content of 59.9%. [source] Preliminary X-ray crystallographic analysis of 2-methylcitrate synthase from Salmonella typhimuriumACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 4 2010Sagar Chittori Analysis of the genomic sequences of Escherichia coli and Salmonella typhimurium has revealed the presence of several homologues of the well studied citrate synthase (CS). One of these homologues has been shown to code for 2-methylcitrate synthase (2-MCS) activity. 2-MCS catalyzes one of the steps in the 2-methylcitric acid cycle found in these organisms for the degradation of propionate to pyruvate and succinate. In the present work, the gene coding for 2-MCS from S. typhimurium (StPrpC) was cloned in pRSET-C vector and overexpressed in E. coli. The protein was purified to homogeneity using Ni,NTA affinity chromatography. The purified protein was crystallized using the microbatch-under-oil method. The StPrpC crystals diffracted X-rays to 2.4,Å resolution and belonged to the triclinic space group P1, with unit-cell parameters a = 92.068, b = 118.159, c = 120.659,Å, , = 60.84, , = 67.77, , = 81.92°. Computation of rotation functions using the X-ray diffraction data shows that the protein is likely to be a decamer of identical subunits, unlike CSs, which are dimers or hexamers. [source] Crystallization and preliminary crystallographic analysis of eukaryotic transcription and mRNA export factor Iws1 from Encephalitozoon cuniculiACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2010Michael Koch Transcription elongation by eukaryotic RNA polymerase II requires the coupling of mRNA synthesis and mRNA processing and export. The essential protein Iws1 is at the interface of these processes through its interaction with histone chaperone and elongation factor Spt6 as well as with complexes involved in mRNA processing and export. Upon crystallization of the evolutionarily conserved domain of Iws1 from Encephalitozoon cuniculi, four different crystal forms were obtained. Three of the crystal forms belonged to space group P21 and one belonged to space group P2221. Preliminary X-ray crystallographic analysis of one of the crystal forms allowed the collection of data to 2.5,Å resolution. [source] Preliminary X-ray crystallographic analysis of a nitric oxide-inducible lactate dehydrogenase from Staphylococcus aureusACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 10 2009Jia-Bin Dong Recent studies have indicated that Staphylococcus aureus can survive the nitrosative stress (caused by the radical nitric oxide; NO·) mounted by the immune system of the infected host. It does this by expressing a nitric oxide-inducible l -lactate dehydrogenase (Sa-LDH-1). Therefore, if efficient inhibitors of Sa-LDH-1 can be designed then Sa-LDH-1 could be a potential drug target against the pathogen S. aureus. For this purpose, the nitric acid-inducible LDH-1 from S. aureus COL strain has been cloned into the expression vector pET-28a(+) and the protein has been expressed, purified and crystallized. The Sa-LDH-1 crystal diffracted to 2.4,Å resolution at a home X-ray source and belonged to space group C2, with unit-cell parameters a = 131.4, b = 74.4, c = 103.2,Å, , = 133.4°. [source] Preliminary X-ray crystallographic analysis of sulfide:quinone oxidoreductase from Acidithiobacillus ferrooxidansACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2009Yanfei Zhang The gene product of open reading frame AFE_1293 from Acidithiobacillus ferrooxidans ATCC 23270 is annotated as encoding a sulfide:quinone oxidoreductase, an enzyme that catalyses electron transfer from sulfide to quinone. Following overexpression in Escherichia coli, the enzyme was purified and crystallized using the hanging-drop vapour-diffusion method. The native crystals belonged to the tetragonal space group P42212, with unit-cell parameters a = b = 131.7, c = 208.8,Å, and diffracted to 2.3,Å resolution. Preliminary crystallographic analysis indicated the presence of a dimer in the asymmetric unit, with an extreme value of the Matthews coefficient (VM) of 4.53,Å3,Da,1 and a solvent content of 72.9%. [source] Preliminary X-ray crystallographic analysis of ornithine acetyltransferase (Rv1653) from Mycobacterium tuberculosisACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2009R. Sankaranarayanan The gene product of open reading frame Rv1653 from Mycobacterium tuberculosis is annotated as encoding a probable ornithine acetyltransferase (OATase; EC 2.3.1.35), an enzyme that catalyzes two steps in the arginine-biosynthesis pathway. It transfers an acetyl group from N -acetylornithine to l -glutamate to produce N -acetylglutamate and l -ornithine. Rv1653 was crystallized using the sitting-drop vapour-diffusion method. The native crystals diffracted to a resolution of 1.7,Å and belonged to space group P212121, with unit-cell parameters a = 60.1, b = 99.7, c = 155.3,Å. The preliminary X-ray study showed the presence of a dimer in the asymmetric unit of the crystals, which had a Matthews coefficient VM of 2.8,Å3,Da,1. [source] Preliminary X-ray crystallographic analysis of SMU.573, a putative sugar kinase from Streptococcus mutansACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2008Yan-Feng Zhou SMU.573 from Streptococcus mutans is a structurally and functionally uncharacterized protein that was selected for structural biology studies. Native and SeMet-labelled proteins were expressed with an N-His tag in Escherichia coli BL21 (DE3) and purified by Ni2+ -chelating and size-exclusion chromatography. Crystals of the SeMet-labelled protein were obtained by the hanging-drop vapour-diffusion method and a 2.5,Å resolution diffraction data set was collected using an in-house chromium radiation source. The crystals belong to space group I4, with unit-cell parameters a = b = 96.53, c = 56.26,Å, , = , = , = 90°. [source] Crystallization and preliminary X-ray crystallographic analyses of CMY-1 and CMY-10, plasmidic class C ,-lactamases with extended substrate spectrumACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2004Sun-Joo Lee Plasmid-encoded class C ,-lactamases, including CMY-1 and CMY-10, hydrolyze the lactam bonds of ,-lactam antibiotics, inducing therapeutic failure and a lack of eradication of clinical isolates by third-generation cephalosporins or cephamycins. Therefore, the enzymes are potential targets for developing agents against pathogens isolated from patients suffering from wound infection, urinary tract infection or pneumonia. CMY-1 and CMY-10 were purified and crystallized at 298,K. X-ray diffraction data from CMY-1 and CMY-10 crystals have been collected to 2.5 and 1.5,Å resolution, respectively, using synchrotron radiation. The crystals of the two proteins are isomorphous and belong to the primitive monoclinic space group P21. [source] Production, purification and preliminary X-ray crystallographic studies of adeno-associated virus serotype 9ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2009Matthew Mitchell Adeno-associated virus (AAV) serotype 9, which is under development for gene-delivery applications, shows significantly enhanced capsid-associated transduction efficiency in muscle compared with other AAV serotypes. With the aim of characterizing the structural determinants of this property, the purification, crystallization and preliminary X-ray crystallographic analyses of the AAV9 viral capsid are reported. The crystals diffracted X-rays to 2.8,Å resolution using synchrotron radiation and belonged to the trigonal space group P32, with unit-cell parameters a = b = 251.0, c = 640.0,Å. There are three complete viral capsids in the crystal unit cell. The orientation and position of the asymmetric unit capsid have been determined by molecular-replacement methods and structure determination is in progress. [source] Crystallization and preliminary X-ray crystallographic analysis of a non-specific lipid-transfer protein with antipathogenic activity from Phaseolus mungoACTA CRYSTALLOGRAPHICA SECTION D, Issue 12-2 2004Shao-Yun Wang A 9,kDa non-specific lipid-transfer protein (nsLTP) from mung bean (Phaseolus mungo) seeds, displaying antifungal activity, antibacterial activity and lipid-transfer activity, was crystallized at 297,K using ammonium sulfate as a precipitant by means of the hanging-drop vapour-diffusion method. Native X-ray diffraction data were collected to a resolution of 2.4,Å. The crystals are rhombohedral, belonging to space group P212121, with unit-cell parameters a = 38.671, b = 51.785, c = 55.925,Å. Assuming the presence of one molecule in the crystallographic asymmetric unit results in a Matthews coefficient (VM) of approximately 3.0,Å3,Da,1, corresponding to a solvent content of about 58%. [source] Crystallization and preliminary X-ray crystallographic analysis of the laminarinase endo-,-1,3-glucanase from Pyrococcus furiosusACTA CRYSTALLOGRAPHICA SECTION D, Issue 12-2 2004Andrea Ilari Laminarinase endo-,-1,3 glucanase (LamA) from Pyrococcus furiosus is an enzyme which displays its main hydrolytic activity on the ,-1,3-glucose polymer laminarin. This laminarinase is remarkably resistant to denaturation: its secondary structure is unchanged in 8,M guanidinium chloride. This protein belongs to the family 16 glycosyl hydrolases, which are enzymes that are widely distributed among bacteria, fungi and higher plants. Single crystals of P. furiosus LamA have been obtained by the hanging-drop vapour-diffusion method using 2-methyl-2,4-pentanediol as a precipitant agent. A complete data set has been collected under cryocooling at a synchrotron source. The crystals belong to the monoclinic space group P21, with unit-cell parameters a = 44.36, b = 84.76, c = 69.23,Å, , = 90, , = 104.97, , = 90°, and diffract to 2.15,Å resolution. [source] Crystallization and preliminary X-ray crystallographic analysis of chorismate synthase from Mycobacterium tuberculosisACTA CRYSTALLOGRAPHICA SECTION D, Issue 11 2004Marcio Vinicius Bertacine Dias The enzymes of the shikimate pathway are potential targets for the development of new therapies because they are essential for bacteria but absent from mammals. The last step in this pathway is performed by chorismate synthase (CS), which catalyzes the conversion of 5-enolpyruvylshikimate-3-phosphate to chorismate. Optimization of crystallization trials allowed the crystallization of homogeneous recombinant CS from Mycobacterium tuberculosis (MtCS). The crystals of MtCS belong to space group P6422 (or P6222) and diffract to 2.8,Å resolution, with unit-cell parameters a = b = 129.7, c = 156.8,Å. There are two molecules in the asymmetric unit. Molecular-replacement trials were not sucessful. Heavy-atom derivative screening is in progress. [source] Purification, crystallization and preliminary X-ray crystallographic analysis of a cysteine-rich secretory protein (CRISP) from Naja atra venomACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2004Yu-Ling Wang Cysteine-rich secretory proteins (CRISPs) play an important role in the innate immune system and are transcriptionally regulated by androgens in several tissues. The proteins are mostly found in the epididymis and granules of mammals, whilst a number of snake venoms also contain CRISP-family proteins. The natrin protein from the venom of Naja atra (Taiwan cobra), which belongs to a family of CRISPs and has a cysteine-rich C-terminal amino-acid sequence, has been purified using a three-stage chromatography procedure and crystals suitable for X-ray analysis have been obtained using the hanging-drop vapour-diffusion method. X-ray diffraction data were collected to 1.58,Å resolution using synchrotron radiation; the crystals belong to space group C2221, with unit-cell parameters a = 59.172, b = 65.038, c = 243.156,Å. There are two protein molecules in the asymmetric unit and the Matthews coefficient is estimated to be 2.35,Å3,Da,1, corresponding to a solvent content of 47.60%. [source] Crystallization and preliminary X-ray crystallographic analysis of nicotinic acid mononucleotide adenylyltransferase from Pseudomonas aeruginosaACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2004Hye-Lee Kim The enzyme nicotinic acid mononucleotide adenylyltransferase (NaMN AT; EC 2.7.7.18) is essential for the synthesis of nicotinamide adenine dinucleotide and is a potential target for antibiotics. It catalyzes the transfer of an adenyl group from ATP to nicotinic acid mononucleotide to form nicotinic acid adenine dinucleotide. NaMN AT from Pseudomonas aeruginosa was overexpressed in Escherichia coli and crystallized at 291,K using 100,mM bis,Tris propane pH 7.0, 700,mM trisodium citrate and 15%(v/v) glycerol. X-ray diffraction data have been collected to 1.70,Å. The crystals are tetragonal, belonging to space group P4122 (or P4322), with unit-cell parameters a = b = 65.02, c = 109.80,Å. The presence of one monomer in the asymmetric unit gives a reasonable VM of 2.15,Å3,Da,1, with a solvent content of 42.7%. [source] Crystallization and preliminary X-ray crystallographic analysis of the RecR protein from Deinococcus radiodurans, a member of the RecFOR DNA-repair pathwayACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2004Byung Il Lee The RecR protein plays a key role in the RecFOR pathway of recombination, which is necessary for the repair of ssDNA gaps. RecR from Deinococcus radiodurans has been overexpressed in Escherichia coli and crystallized at 297,K using polyethylene glycol 1000 as a precipitant. X-ray diffraction data to 2.90,Å resolution have been collected at 100,K using Cu,K, X-rays from a mercury-soaked crystal. The crystal belongs to space group C2221, with unit-cell parameters a = 106.96, b = 122.25, c = 156.01,Å. The asymmetric unit contains four monomers of RecR, with a crystal volume per protein weight (VM) of 2.57,Å3,Da,1 and a solvent content of 51.0%. [source] Crystallization and preliminary X-ray crystallographic analysis of the trimer core from measles virus fusion proteinACTA CRYSTALLOGRAPHICA SECTION D, Issue 3 2003Jieqing Zhu Two heptad-repeat regions (HR1 and HR2) are highly conserved in paramyxovirus fusion proteins and form a stable helical trimer of heterodimers [(HR1,HR2)3] after the fusion between viral and cellular membranes. In this study, two HR regions of the fusion protein of measles virus, a member of the paramyxoviruses, were selected and overexpressed as a single chain (named 2-Helix) connected by an amino-acid linker using a GST-fusion expression system in Escherichia coli. Crystals of 2-Helix protein (GST removed) could be obtained from many conditions using the sitting- or hanging-drop vapour-diffusion method. A complete data set was collected in-house to 1.9,Å resolution from a single crystal. The crystal belongs to space group P6, with unit-cell parameters a = b = 51.637, c = 67.058,Å. To facilitate the crystal structure solution, SeMet-substituted 2-Helix crystals, grown under similar conditions to the native, were also obtained and diffracted X-rays to 1.8,Å using synchrotron radiation. [source] Crystallization and preliminary X-ray crystallographic analysis of peptide deformylase from Pseudomonas aeruginosaACTA CRYSTALLOGRAPHICA SECTION D, Issue 10-2 2002Hyung-Wook Kim Peptide deformylase (PDF) from the pathogenic bacterium Pseudomonas aeruginosa has been overexpressed in Escherichia coli and crystallized in the presence of its inhibitor actinonin at 297,K using polyethylene glycol (PEG) 4000 as a precipitant. The diffraction limit and the spot shape of the crystals could be slightly improved by the crystal annealing/dehydration procedure. X-ray diffraction data to 1.85,Å have been collected using synchrotron radiation. The crystal belongs to the orthorhombic space group P212121, with unit-cell parameters a = 68.75, b = 74.46, c = 77.18,Å. The asymmetric unit contains two subunits of peptide deformylase, with a corresponding crystal volume per protein mass (VM) of 2.45,Å3,Da,1 and a solvent content of 49.8%. [source] Crystallization and preliminary X-ray crystallographic analysis of p24, a component of the potato nuclear factor PBF-2ACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2002Darrell Desveaux The Solanum tuberosum (potato) nuclear factor PBF-2 is implicated in pathogen-induced expression of the pathogenesis-related gene PR-10a. Crystals of the DNA-binding component of PBF-2, p24, have been obtained at 277,K in 20,mM Tris,HCl pH 8.0. Recombinant protein with a His tag at its C-terminus was overexpressed in Escherichia coli in the presence and absence of selenomethionine and was purified using a combination of HiTrap affinity columns and gel-filtration chromatography. Crystals suitable for structural analysis were obtained for both native and selenomethionine-labelled proteins and yielded diffraction data at 100,K that were processed to 2.3 and 2.8,Å resolution, respectively. The p24 protein crystals belong to space group P212121, with unit-cell parameters a = 69.4,(69.1), b = 89.4,(90.5), c = 144.1,(144.3),Å. The asymmetric unit contains four protomers, giving a crystal volume per protein mass (VM) of 2.23,Å3,Da,1 and a solvent content of 45% by volume. [source] Crystallization and preliminary X-ray crystallographic analysis of the Rv2002 gene product from Mycobacterium tuberculosis, a ,-ketoacyl carrier protein reductase homologueACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2002Jin Kuk Yang A 260-residue protein (FabG3) encoded by the Rv2002 gene of Mycobacterium tuberculosis shows amino-acid sequence similarity to ,-ketoacyl carrier protein (ACP) reductase, FabG. A soluble mutant (I6T/V47M/T69M) was produced by the green fluorescent protein-based directed-evolution method. It was crystallized at 296,K using the hanging-drop vapour-diffusion method. The diffraction quality of the crystal improved significantly after annealing/dehydration. X-ray diffraction data were collected to 1.8,Å resolution using synchrotron radiation. The crystal belongs to the space group P3121 (or P3221), with unit-cell parameters a = b = 70.38, c = 148.93,Å. The asymmetric unit contains two subunits, with a corresponding VM of 1.90,Å3,Da,1 and a solvent content of 35.3%. [source] Crystallization and preliminary X-ray crystallographic analysis of deoxyuridine triphosphate nucleotidohydrolase from Saccharomyces cerevisiaeACTA CRYSTALLOGRAPHICA SECTION D, Issue 8 2001Byung Woo Han Deoxyuridine triphosphate nucleotidohydrolase (dUTPase) from Saccharomyces cerevisiae is essential for cell viability. It has been overexpressed in Escherichia coli and has been crystallized at 296,K using polyethylene glycol (PEG) 1500 as a precipitant. The crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 59.48, b = 138.54, c = 157.91,Å, , = , = , = 90°. Two molecules of trimeric dUTPase from S. cerevisiae are present in the asymmetric unit, giving a crystal volume per protein mass (VM) of 3.36,Å3,Da,1 and a solvent content of 63%. The diffraction limit of the crystals could be significantly extended by the crystal-annealing procedure. A set of native data extending to 2.7,Å resolution has been collected at 100,K using synchrotron X-rays. [source] Crystallization and preliminary X-ray crystallographic analysis of thioesterase I from Escherichia coliACTA CRYSTALLOGRAPHICA SECTION D, Issue 6 2000Yu-Chih Lo The Escherichia coli thioesterase I specifically catalyzes the deacylation of fatty acyl,CoA thioesters, especially those with long acyl groups (C12,C18). Single crystals of thioesterase I (E.C. 3.1.2.2) from E. coli have been obtained using methoxypolyethylene glycol 5000 (PEG,MME 5K) as a precipitant at room temperature in 21,d. The crystals belong to the tetragonal space group P41212 or its enantiomorph P43212, with unit-cell parameters a = b = 50.85,(7), c = 171.5,(1),Å. The crystals diffract to beyond 2.4,Å resolution. There is one molecule of molecular weight 20.5,kDa in the asymmetric unit, with a solvent content of 55%. [source] Crystallization and preliminary X-ray crystallographic analysis of the ArsM arsenic(III) S -adenosylmethionine methyltransferaseACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010Kavitha Marapakala Arsenic is the most ubiquitous environmental toxin and carcinogen and consequently ranks first on the Environmental Protection Agency's Superfund Priority List of Hazardous Substances. It is introduced primarily from geochemical sources and is acted on biologically, creating an arsenic biogeocycle. A common biotransformation is methylation to monomethylated, dimethylated and trimethylated species. Methylation is catalyzed by the ArsM (or AS3MT) arsenic(III) S -adenosylmethionine methyltransferase, an enzyme (EC 2.1.1.137) that is found in members of every kingdom from bacteria to humans. ArsM from the thermophilic alga Cyanidioschyzon sp. 5508 was expressed, purified and crystallized. Crystals were obtained by the hanging-drop vapor-diffusion method. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 84.85, b = 46.89, c = 100.35,Å, , = 114.25° and one molecule in the asymmetric unit. Diffraction data were collected at the Advanced Light Source and were processed to a resolution of 1.76,Å. [source] Cloning, expression, crystallization and preliminary X-ray crystallographic analysis of a human condensin SMC2 hinge domain with short coiled coilsACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010Kazuki Kawahara In higher eukaryotes, the condensin complex, which mainly consists of two structural maintenance of chromosomes (SMC) subunits, SMC2 (CAP-E) and SMC4 (CAP-C), plays a critical role in the formation of higher order chromosome structures during mitosis. Biochemical and electron-microscopic studies have revealed that the SMC2 and SMC4 subunits dimerize through the interaction of their hinge domains, forming a characteristic V-shaped heterodimer. However, the details of their function are still not fully understood owing to a lack of structural information at the atomic level. In this study, the human SMC2 hinge domain with short coiled coils was cloned, expressed, purified and crystallized in the orthorhombic space group C222 in native and SeMet-derivatized forms. Because of the poor diffraction properties of these crystals, the mutant Leu68,SeMet was designed and crystallized in order to obtain the experimental phases. The SeMet-derivatized crystals of the mutant belonged to space group P3212, with unit-cell parameters a = b = 128.8, c = 91.4,Å. The diffraction data obtained from a crystal that diffracted to 2.4,Å resolution were suitable for SAD phasing. [source] Crystallization and preliminary X-ray crystallographic analysis of a bacterial tyrosinase from Bacillus megateriumACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 9 2010Mor Sendovski Tyrosinases are type 3 copper enzymes that are involved in the production of melanin and have two copper ions in the active site. Here, the crystallization and primary analysis of a tyrosinase from Bacillus megaterium is reported. The purified protein was crystallized in the absence or presence of zinc ions and the crystals diffracted to a resolution of 2.0,Å. Crystals obtained in the presence of zinc belonged to space group P212121, while crystals grown in the absence of zinc belonged to space group P21. In both space groups the asymmetric unit contained a dimer, with minor differences in the crystal density and in packing interactions. [source] Overexpression, crystallization and preliminary X-ray crystallographic analysis of Pseudomonas aeruginosa MnmE, a GTPase involved in tRNA modificationACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2010Hyung Ho Lee MnmE, an evolutionarily conserved GTPase, is involved in modification of the uridine base (U34) at the wobble position of certain tRNAs. Previous crystal structures of MnmE suggest that it is a dimer with considerable conformational flexibility. To facilitate structural comparisons among MnmE proteins, structural analysis of MnmE from Pseudomonas aeruginosa encoded by the PA5567 gene was initiated. It was overexpressed in Escherichia coli and crystallized at 297,K using a reservoir solution consisting of 100,mM sodium ADA pH 6.5, 12%(w/v) polyethylene glycol 4000, 100,mM lithium sulfate, 2%(v/v) 2-propanol and 2.5,mM dithiothreitol. X-ray diffraction data were collected to 2.69,Å resolution. The crystals belonged to the orthorhombic space group C2221, with unit-cell parameters a = 96.74, b = 204.66, c = 120.90,Å. Two monomers were present in the asymmetric unit, resulting in a crystal volume per protein mass (VM) of 2.99,Å3,Da,1 and a solvent content of 58.8%. [source] Cloning, purification, crystallization and preliminary X-ray crystallographic analysis of SET/TAF-I,,N from Homo sapiensACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2010Zhen Xu The histone chaperone SET encoded by the SET gene, which is also known as template-activating factor I, (TAF-I,), is a multifunctional molecule that is involved in many biological phenomena such as histone binding, nucleosome assembly, chromatin remodelling, replication, transcription and apoptosis. A truncated SET/TAF-I,,N protein that lacked the first 22 residues of the N-terminus but contained the C-terminal acidic domain and an additional His6 tag at the C-terminus was overexpressed in Escherichia coli and crystallized by the hanging-drop vapour-diffusion method using sodium acetate as precipitant at 283,K. The crystals diffracted to 2.7,Å resolution and belonged to space group P43212. [source] Crystallization and preliminary X-ray crystallographic analysis of BxlA, an intracellular ,- d -xylosidase from Streptomyces thermoviolaceus OPC-520ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 7 2010Hideaki Morioka BxlA from Streptomyces thermoviolaceus OPC-520, together with the extracellular BxlE and the integral membrane proteins BxlF and BxlG, constitutes a xylanolytic system that participates in the intracellular transport of xylan-degradation products and the production of xylose. To elucidate the mechanism of the hydrolytic degradation of xylooligosaccharides to xylose at the atomic level, X-ray structural analysis of BxlA was attempted. The recombinant BxlA protein (molecular weight 82,kDa) was crystallized by the hanging-drop vapour-diffusion method at 289,K. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 142.2, b = 129.5, c = 101.4,Å, , = 119.8°, and contained two molecules per asymmetric unit (VM = 2.47,Å3,Da,1). Diffraction data were collected to a resolution to 2.50,Å and provided a data set with an overall Rmerge of 8.3%. [source] | ||||||||||