Crystallization and preliminary crystallographic analysis of the snake muscle fructose 1,6-bisphosphatase

1999 ◽  
Vol 55 (7) ◽  
pp. 1342-1344 ◽  
Author(s):  
D.-W. Zhu ◽  
G.-J. Xu ◽  
P. H. Rehse ◽  
A. Azzi ◽  
F.-K. Zhao ◽  
...  
Keyword(s):  
Polyethylene Glycol ◽  
Magnesium Chloride ◽  
Crystallographic Analysis ◽  
Molecular Replacement ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
Allosteric Enzyme ◽  
X Ray ◽  
Cell Parameters ◽  
Fructose 1,6 Bisphosphatase

The snake muscle fructose 1,6-bisphosphatase, a typical allosteric enzyme which plays important roles in gluconeogenesis, was crystallized in the presence of polyethylene glycol 3350 and magnesium chloride at pH 8.5. The crystals diffract to 2.3 Å on a rotating-anode X-ray source. The space group was determined to be either P3121 or its enantiomorph P3221, with unit-cell parameters a = b = 83.7, c = 202.41 Å, α = β = 90 and γ = 120°. There are two subunits in the asymmetric unit. Preliminary molecular-replacement studies indicate that the first enantiomorph is the correct one.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of human myotubularin-related protein 1

2015 ◽  
Vol 71 (3) ◽  
pp. 261-265 ◽  
Author(s):  
Seoung Min Bong ◽  
Seung Won Yang ◽  
Ji-Woong Choi ◽  
Seung Jun Kim ◽  
Byung Il Lee
Keyword(s):  
Escherichia Coli ◽  
Polyethylene Glycol ◽  
Synchrotron Radiation ◽  
Crystallographic Analysis ◽  
Asymmetric Unit ◽  
Related Protein ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
X Ray ◽  
Cell Parameters

Myotubularin-related protein 1 is a phosphatase that dephosphorylates phospholipids such as phosphatidylinositol 3-phosphate or phosphatidylinositol 3,5-bisphosphate. In this study, human MTMR1 was overexpressed inEscherichia coli, purified and crystallized at 277 K using polyethylene glycol 20 000 as a precipitant. Diffraction data were collected to 2.0 Å resolution using synchrotron radiation. The crystals belonged to space groupP1, with unit-cell parametersa= 67.219,b= 96.587,c= 97.581 Å, α = 87.597, β = 86.072, γ = 77.327°. Assuming the presence of four molecules in the asymmetric unit, the calculated Matthews coefficient value was 2.61 Å3 Da−1and the corresponding solvent content was 52.9%.

scholarly journals Characterization, crystallization and preliminary X-ray crystallographic analysis of the Uba5 fragment necessary for high-efficiency activation of Ufm1

2014 ◽  
Vol 70 (6) ◽  
pp. 765-768 ◽  
Author(s):  
Shutao Xie
Keyword(s):  
High Efficiency ◽  
Crystallographic Analysis ◽  
Terminal Region ◽  
Adenylation Domain ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Multicellular Organisms

Uba5 is the smallest ubiquitin-like molecule-activating enzyme and contains an adenylation domain and a C-terminal region. This enzyme only exists in multicellular organisms. The mechanism through which the enzyme recognizes and activates ubiquitin-fold modifier 1 (Ufm1) remains unknown. In this study, Uba5 adenylation domains with different C-terminal region lengths were cloned, expressed and purified. The results of anin vitrotruncation assay suggest that Uba5 residues 57–363 comprise the minimal fragment required for the high-efficiency activation of Ufm1. Crystallization of Uba5 residues 57–363 was performed at 277 K using PEG 3350 as the precipitant, and crystals optimized by microseeding diffracted to 2.95 Å resolution, with unit-cell parametersa=b= 97.66,c= 144.83 Å, α = β = 90, γ = 120°. There is one molecule in the asymmetric unit; the Matthews coefficient and the solvent content were calculated to be 2.93 Å3 Da−1and 58.1%, respectively.

scholarly journals Purification, crystallization and preliminary X-ray crystallographic analysis of the UDP-N-acetylmuramoyl-tripeptide-D-alanyl-D-alanine ligase (MurF) fromAcinetobacter baumannii

2014 ◽  
Vol 70 (7) ◽  
pp. 976-978 ◽  
Author(s):  
Young Jun An ◽  
Chang-Sook Jeong ◽  
Jeong Hee Yu ◽  
Kyung Min Chung ◽  
Sun-Shin Cha
Keyword(s):  
Multidrug Resistant ◽  
Crystallographic Analysis ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
X Ray ◽  
Cell Parameters ◽  
Peptidoglycan Biosynthesis ◽  
Crystallization Method ◽  
Global Spread

The emergence and global spread of multidrug-resistantAcinetobacter baumanniistrains are major threats to public health. Inhibition of peptidoglycan biosynthesis is an effective strategy for the development of antibiotics. The ATP-dependent UDP-N-acetylmuramoyl-tripeptide-D-alanyl-D-alanine ligase (MurF) that is responsible for the last step of peptidoglycan biosynthesis is a validated target for the development of antibiotics. Crystals ofA. baumanniiMurF in complex with ATP were grown by the microbatch crystallization method at 295 K. The crystals belonged to space groupP3221, with unit-cell parametersa=b= 85.42,c= 129.86 Å. Assuming the presence of one molecule in the asymmetric unit, the solvent content was estimated to be about 54.32%.

scholarly journals Cloning, purification and preliminary X-ray crystallographic analysis of the OmpA-like domain of peptidoglycan-associated lipoprotein fromAcinetobacter baumannii

2012 ◽  
Vol 68 (11) ◽  
pp. 1351-1353 ◽  
Author(s):  
Jung Hyun Song ◽  
Woo Cheol Lee ◽  
Jeong Soon Park ◽  
Seung Il Kim ◽  
Je Chul Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acinetobacter Baumannii ◽  
Crystallographic Analysis ◽  
Diffusion Method ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Vapour Diffusion ◽  
Strain Bl21

Peptidoglycan-associated lipoprotein (Pal) is one component of the Tol–Pal system that is involved in maintaining the integrity and stability of the outer membrane. The C-terminal OmpA-like domain of Pal interacts noncovalently with peptidoglycan. In this study, the OmpA-like domain of Pal fromAcinetobacter baumanniiwas overexpressed inEscherichia colistrain BL21 (DE3), purified and crystallized using the vapour-diffusion method. A native crystal diffracted to 1.4 Å resolution and belonged to space groupP61orP65, with unit-cell parametersa=b= 72.58,c= 44.65 Å, a calculated Matthews coefficient of 2.64 Å3 Da−1and one molecule per asymmetric unit.

scholarly journals Overproduction, crystallization and preliminary X-ray crystallographic analysis ofEscherichia colitRNAN6-threonylcarbamoyladenosine dehydratase

2014 ◽  
Vol 70 (11) ◽  
pp. 1517-1520 ◽  
Author(s):  
Sunmin Kim ◽  
Keon Young Kim ◽  
Jeong Kuk Park ◽  
Byung Il Lee ◽  
Yun-Gon Kim ◽  
...  
Keyword(s):  
Crystallographic Analysis ◽  
X Rays ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
E Coli ◽  
Protein Mass ◽  
Crystal Volume

Escherichia colitRNAN6-threonylcarbamoyladenosine dehydratase (TcdA), previously called CsdL or YgdL, was overproduced and purified fromE. coliand crystallized using polyethylene glycol 3350 as a crystallizing agent. X-ray diffraction data were collected to 2.70 Å resolution under cryoconditions using synchrotron X-rays. The crystals belonged to space groupP21, with unit-cell parametersa= 65.4,b= 96.8,c= 83.3 Å, β = 111.7°. According to the Matthews coefficient, the asymmetric unit may contain up to four subunits of the monomeric protein, with a crystal volume per protein mass (VM) of 2.12 Å3 Da−1and 42.1% solvent content.

scholarly journals Crystallization and preliminary X-ray crystallographic analysis of a bacterial Asn-transamidosome

2014 ◽  
Vol 70 (6) ◽  
pp. 790-793 ◽  
Author(s):  
Tateki Suzuki ◽  
Keitaro Yamashita ◽  
Yoshikazu Tanaka ◽  
Isao Tanaka ◽  
Min Yao
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Thermus Thermophilus ◽  
Crystallographic Analysis ◽  
Asymmetric Unit ◽  
Ribonucleoprotein Particle ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Trna Synthetases ◽  
Aminoacyl Trna Synthetases

Most canonical aminoacyl-tRNAs are synthesized directly by their cognate aminoacyl-tRNA synthetases (aaRSs), but glutaminyl-tRNAGlnand asparaginyl-tRNAAsnare synthesized indirectly by two-step processes. These processes are catalyzed by the transamidosome, a large ribonucleoprotein particle composed of GatA, GatB, GatC, aaRS and tRNA. In this study, the Asn-transamidosome fromPseudomonas aeruginosawas reconstructed and crystallized by mixing purified GatCAB complex, AspRS and tRNAAsn. The crystal of the Asn-transamidosome belonged to space groupP21, with unit-cell parametersa= 93.3,b= 186.0,c= 287.8 Å, β = 93.3°, and diffracted to 3.73 Å resolution. Preliminary X-ray crystallographic analysis showed that the asymmetric unit contained two Asn-transamidosomes, each composed of two GatCABs, one AspRS dimer and two tRNAAsns, indicating that the construction of the current Asn-transamidosome differs from that ofThermus thermophilus.

scholarly journals Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of a ferredoxin/flavodoxin-NADP(H) oxidoreductase (Bc0385) fromBacillus cereus

2014 ◽  
Vol 70 (6) ◽  
pp. 777-780 ◽  
Author(s):  
Silje Skråmo ◽  
Hans-Petter Hersleth ◽  
Marta Hammerstad ◽  
K. Kristoffer Andersson ◽  
Åsmund K. Røhr
Keyword(s):  
Electron Transfer ◽  
Thioredoxin Reductase ◽  
Free Form ◽  
Molecular Replacement ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Orthorhombic Space Group ◽  
X Ray ◽  
Cell Parameters

Ferredoxin/flavodoxin-NADP(H) oxidoreductases (FNRs) are key enzymes involved in catalysing electron transfer between ferredoxins/flavodoxins and NAD(P)H/NAD(P)+. InBacillus cereusthere are three genes that may encode FNRs, and the Bc0385 FNR has been cloned, overexpressed, purified and successfully crystallized in its NADPH/NADP+-free form. Diffraction data have been collected to 2.5 Å resolution from crystals belonging to the orthorhombic space groupP21212, with unit-cell parametersa= 57.2,b= 164.3,c= 95.0 Å, containing two FNR molecules in the asymmetric unit. The structure of the Bc0385 FNR has been solved by molecular replacement, and is a member of the homodimeric thioredoxin reductase-like class of FNRs.

scholarly journals Expression, crystallization and preliminary X-ray crystallographic analysis ofD-alanine-D-alanine ligase from OXA-23-producingAcinetobacter baumanniiK0420859

2014 ◽  
Vol 70 (4) ◽  
pp. 505-508 ◽  
Author(s):  
Kim-Hung Huynh ◽  
Huyen-Thi Tran ◽  
Tan-Viet Pham ◽  
Ho-Phuong-Thuy Ngo ◽  
Sun-Shin Cha ◽  
...  
Keyword(s):  
Crystallographic Analysis ◽  
Antibacterial Drug ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
Orthorhombic Space Group ◽  
Solvent Content ◽  
X Ray ◽  
Cell Parameters ◽  
Novel Target ◽  
Tract Infections

Acinetobacter baumanniicauses bacteraemia, pneumonia, other respiratory-tract and urinary-tract infections in humans. OXA-23 carbapenemase-producingA. baumanniiK0420859 (A. baumanniiOXA-23) is resistant to carbapenem, a common antibacterial drug. To develop an efficient and novel antibacterial drug againstA. baumanniiOXA-23, D-alanine-D-alanine ligase, which is essential in bacterial cell-wall synthesis, is of interest. Here, the D-alanine-D-alanine ligase (AbDdl) gene fromA. baumanniiOXA-23 was cloned and expressed, and theAbDdl protein was purified and crystallized; this enzyme can be used as a novel target for an antibacterial drug againstA. baumanniiOXA-23. TheAbDdl crystal diffracted to a resolution of 2.8 Å and belonged to the orthorhombic space groupP212121, with unit-cell parametersa= 113.4,b= 116.7,c= 176.5 Å, a correspondingVMof 2.8 Å3 Da−1and a solvent content of 56.3%, and six protomers in the asymmetric unit.

scholarly journals The LRR-Roc-COR module of theChlorobium tepidumRoco protein: crystallization and X-ray crystallographic analysis

2017 ◽  
Vol 73 (9) ◽  
pp. 520-524 ◽  
Author(s):  
Egon Deyaert ◽  
Arjan Kortholt ◽  
Wim Versées
Keyword(s):  
Protein Crystallization ◽  
Crystallographic Analysis ◽  
Chlorobium Tepidum ◽  
Molecular Replacement ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Protein Dimer ◽  
Map Analysis ◽  
Lrr Domain

Roco proteins are characterized by the presence of a Roc-COR supradomain harbouring GTPase activity, which is often preceded by an LRR domain. The most notorious member of the Roco protein family is the Parkinson's disease-associated LRRK2. The Roco protein from the bacteriumChlorobium tepidumhas been used as a model system to investigate the structure and mechanism of this class of enzymes. Here, the crystallization and crystallographic analysis of the LRR-Roc-COR construct of theC. tepidumRoco protein is reported. The LRR-Roc-COR crystals belonged to space groupP212121, with unit-cell parametersa= 95.6,b= 129.8,c= 179.5 Å, α = β = γ = 90°, and diffracted to a resolution of 3.3 Å. Based on the calculated Matthews coefficient, Patterson map analysis and an initial molecular-replacement analysis, one protein dimer is present in the asymmetric unit. The crystal structure of this protein will provide valuable insights into the interaction between the Roc-COR and LRR domains within Roco proteins.

Crystallization and preliminary X-ray analysis of a complex between the Bowman–Birk trypsin inhibitor from barley and porcine pancreatic trypsin

1999 ◽  
Vol 55 (6) ◽  
pp. 1244-1246 ◽  
Author(s):  
Young Sil Kim ◽  
Hyun Kyu Song ◽  
Se Won Suh
Keyword(s):  
Polyethylene Glycol ◽  
Trypsin Inhibitor ◽  
Unit Cell ◽  
X Rays ◽  
Asymmetric Unit ◽  
Unit Cell Parameters ◽  
Orthorhombic Space Group ◽  
Solvent Content ◽  
X Ray ◽  
Cell Parameters

A 1:2 complex between the Bowman–Birk trypsin inhibitor from barley seeds and porcine pancreatic trypsin has been crystallized at 291 K using polyethylene glycol as precipitant. The crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 67.10, b = 88.38 and c = 203.65 Å. The asymmetric unit contains two monomers of the complex, with a corresponding Vm of 2.41 Å3 Da−1 and a solvent content of 49%. Native data to 2.2 Å resolution have been collected at 100 K using synchrotron X-rays.

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