scholarly journals Crystallization and preliminary X-ray diffraction analysis of a novel β-L-arabinofuranosidase (HypBA1) fromBifidobacterium longum

2014 ◽  
Vol 70 (5) ◽  
pp. 636-638 ◽  
Author(s):  
Zhen Zhu ◽  
Miao He ◽  
Chun-Hsiang Huang ◽  
Tzu-Ping Ko ◽  
Yi-Fang Zeng ◽  
...  
Keyword(s):  
Glycoside Hydrolase ◽  
Bifidobacterium Longum ◽  
Data Bank ◽  
Diffusion Method ◽  
Glycoside Hydrolase Family ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Hydrolase Family

The β-L-arabinofuranosidase (HypBA1) fromBifidobacterium longumJCM 1217 hydrolyzes the β-1,2-linked arabinofuranose disaccharide to release L-arabinoses. HypBA1 was classified into glycoside hydrolase family 127 (GH127) by the CAZy website (http://www.cazy.org/). The enzyme was expressed inEscherichia coliand the purified recombinant protein was crystallized. Crystals belonging to the primitive hexagonal space groupP3x21, with unit-cell parametersa=b= 75.9,c= 254.0 Å, were obtained by the sitting-drop vapour-diffusion method and diffracted to 2.78 Å resolution. ABLASTPsearch (http://blast.ncbi.nlm.nih.gov/) of the Protein Data Bank did not reveal any similar crystal structures. Structural determination by using SeMet MAD and MIR methods is in progress.

scholarly journals Structure of recombinant prolidase fromThermococcus sibiricusin space groupP21221

2015 ◽  
Vol 71 (8) ◽  
pp. 951-957 ◽  
Author(s):  
Vladimir Timofeev ◽  
Elvira Slutskaya ◽  
Marina Gorbacheva ◽  
Konstantin Boyko ◽  
Tatiana Rakitina ◽  
...  
Keyword(s):  
Data Bank ◽  
High Variability ◽  
Protein Crystal ◽  
Diffusion Method ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Counter Diffusion

The crystal structure of recombinant prolidase fromThermococcus sibiricuswas determined by X-ray diffraction at a resolution of 2.6 Å and was found to contain a tetramer in the asymmetric unit. A protein crystal grown in microgravity using the counter-diffusion method was used for X-ray studies. The crystal belonged to space groupP21221, with unit-cell parametersa= 97.60,b= 123.72,c= 136.52 Å, α = β = γ = 90°. The structure was refined to anRcrystof 22.1% and anRfreeof 29.6%. The structure revealed flexible folding of the N-terminal domain of the protein as well as high variability in the positions of the bound metal ions. The coordinates of the resulting model were deposited in the Protein Data Bank as entry 4rgz.

scholarly journals Expression, purification, crystallization and preliminary X-ray diffraction analysis ofAspergillus terreusendo-β-1,4-glucanase from glycoside hydrolase family 12

2014 ◽  
Vol 70 (2) ◽  
pp. 267-270 ◽  
Author(s):  
Fernando Segato ◽  
Gabriela L. Berto ◽  
Evandro Ares de Araújo ◽  
João Renato Muniz ◽  
Igor Polikarpov
Keyword(s):  
Glycoside Hydrolase ◽  
Aspergillus Terreus ◽  
Glycoside Hydrolases ◽  
Glycoside Hydrolase Family ◽  
Molecular Replacement ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
Replacement Solution ◽  
Hydrolase Family

Endoglucanases are important enzymes that are involved in the modification and degradation of cellulose. Filamentous fungi such asAspergillus terreusare effective biomass degraders in nature owing to their capacity to produce an enzymatic arsenal of glycoside hydrolases, including endoglucanase from glycoside hydrolase family 12 (GH12). TheA. terreusGH12 endoglucanase was cloned and overexpressed inA. nidulans, purified and crystallized. A single crystal was obtained from a solution consisting of 2 Mammonium sulfate, 5%(v/v) 2-propanol. X-ray diffraction data were collected to a resolution of 1.85 Å using synchrotron radiation and a preliminary molecular-replacement solution was obtained in the trigonal space groupP3221. The unit-cell parameters werea=b= 103.24,c= 48.96 Å.

scholarly journals Crystallization and preliminary X-ray diffraction analysis of the Csu pili CsuC–CsuA/B chaperone–major subunit pre-assembly complex fromAcinetobacter baumannii

2015 ◽  
Vol 71 (6) ◽  
pp. 770-774 ◽  
Author(s):  
Natalia Pakharukova ◽  
Minna Tuittila ◽  
Sari Paavilainen ◽  
Anton Zavialov
Keyword(s):  
Diffusion Method ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Unit Cell Parameters ◽  
Gram Negative ◽  
X Ray ◽  
Cell Parameters ◽  
Abiotic Surfaces ◽  
Vapour Diffusion ◽  
Fimbrial Adhesins

The attachment of many Gram-negative pathogens to biotic and abiotic surfaces is mediated by fimbrial adhesins, which are assembledviathe classical, alternative and archaic chaperone–usher (CU) pathways. The archaic CU fimbrial adhesins have the widest phylogenetic distribution, yet very little is known about their structure and mechanism of assembly. To elucidate the biogenesis of archaic CU systems, structural analysis of the Csu fimbriae, which are used byAcinetobacter baumanniito form stable biofilms and cause nosocomial infection, was focused on. The major fimbriae subunit CsuA/B complexed with the CsuC chaperone was purified from the periplasm ofEscherichia colicells co-expressing CsuA/B and CsuC, and the complex was crystallized in PEG 3350 solution using the hanging-drop vapour-diffusion method. Selenomethionine-labelled CsuC–CsuA/B complex was purified and crystallized under the same conditions. The crystals diffracted to 2.40 Å resolution and belonged to the hexagonal space groupP6422, with unit-cell parametersa=b= 94.71,c = 187.05 Å, α = β = 90, γ = 120°. Initial phases were derived from a single anomalous diffraction (SAD) experiment using the selenomethionine derivative.

scholarly journals Crystallization and X-ray diffraction analysis of native and selenomethionine-substituted PhyH-DI fromBacillussp. HJB17

2017 ◽  
Vol 73 (11) ◽  
pp. 607-611
Author(s):  
Fang Lu ◽  
Bei Zhang ◽  
Yong Liu ◽  
Ying Song ◽  
Gangxing Guo ◽  
...  
Keyword(s):  
Unit Cell ◽  
Diffusion Method ◽  
Data Sets ◽  
Asymmetric Unit ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Solvent Content ◽  
Space Group ◽  
X Ray ◽  
Cell Parameters

Phytases are phosphatases that hydrolyze phytates to less phosphorylatedmyo-inositol derivatives and inorganic phosphate. β-Propeller phytases, which are very diverse phytases with improved thermostability that are active at neutral and alkaline pH and have absolute substrate specificity, are ideal substitutes for other commercial phytases. PhyH-DI, a β-propeller phytase fromBacillussp. HJB17, was found to act synergistically with other single-domain phytases and can increase their efficiency in the hydrolysis of phytate. Crystals of native and selenomethionine-substituted PhyH-DI were obtained using the vapour-diffusion method in a condition consisting of 0.2 Msodium chloride, 0.1 MTris pH 8.5, 25%(w/v) PEG 3350 at 289 K. X-ray diffraction data were collected to 3.00 and 2.70 Å resolution, respectively, at 100 K. Native PhyH-DI crystals belonged to space groupC121, with unit-cell parametersa = 156.84,b = 45.54,c = 97.64 Å, α = 90.00, β = 125.86, γ = 90.00°. The asymmetric unit contained two molecules of PhyH-DI, with a corresponding Matthews coefficient of 2.17 Å3 Da−1and a solvent content of 43.26%. Crystals of selenomethionine-substituted PhyH-DI belonged to space groupC2221, with unit-cell parametersa = 94.71,b= 97.03,c= 69.16 Å, α = β = γ = 90.00°. The asymmetric unit contained one molecule of the protein, with a corresponding Matthews coefficient of 2.44 Å3 Da−1and a solvent content of 49.64%. Initial phases for PhyH-DI were obtained from SeMet SAD data sets. These data will be useful for further studies of the structure–function relationship of PhyH-DI.

scholarly journals Preliminary X-ray diffraction analysis of a thermophilic β-1,3–1,4-glucanase fromClostridium thermocellum

2014 ◽  
Vol 70 (7) ◽  
pp. 946-948 ◽  
Author(s):  
Lilan Zhang ◽  
Puya Zhao ◽  
Chun-Chi Chen ◽  
Chun-Hsiang Huang ◽  
Tzu-Ping Ko ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Specific Activity ◽  
High Specific Activity ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Glycosidic Bonds ◽  
Hydrolysis Of

β-1,3–1,4-Glucanases catalyze the specific hydrolysis of internal β-1,4-glycosidic bonds adjacent to the 3-O-substituted glucose residues in mixed-linked β-glucans. The thermophilic glycoside hydrolase CtGlu16A fromClostridium thermocellumexhibits superior thermal profiles, high specific activity and broad pH adaptability. Here, the catalytic domain of CtGlu16A was expressed inEscherichia coli, purified and crystallized in the trigonal space groupP3121, with unit-cell parametersa=b= 74.5,c= 182.9 Å, by the sitting-drop vapour-diffusion method and diffracted to 1.95 Å resolution. The crystal contains two protein molecules in an asymmetric unit. Further structural determination and refinement are in progress.

scholarly journals Crystallization and preliminary X-ray diffraction analysis of AntE, a crotonyl-CoA carboxylase/reductase fromStreptomycessp. NRRL 2288

2014 ◽  
Vol 70 (6) ◽  
pp. 734-737 ◽  
Author(s):  
Lihan Zhang ◽  
Jing Chen ◽  
Takahiro Mori ◽  
Yan Yan ◽  
Wen Liu ◽  
...  
Keyword(s):  
Diffusion Method ◽  
Antimycin A ◽  
X Ray Diffraction ◽  
Unit Cell Parameters ◽  
Data Set ◽  
X Ray ◽  
Cell Parameters ◽  
Vapour Diffusion ◽  
Unsaturated Acyl ◽  
Reductive Carboxylation

AntE fromStreptomycessp. NRRL 2288 is a crotonyl-CoA carboxylase/reductase that catalyzes the reductive carboxylation of various α,β-unsaturated acyl-CoAs to provide the building block at the C7 position for antimycin A biosynthesis. Recombinant AntE expressed inEscherichia coliwas crystallized by the sitting-drop vapour-diffusion method. The crystals belonged to space groupI222 orI212121, with unit-cell parametersa= 76.4,b= 96.7,c= 129.6 Å, α = β = γ = 90.0°. A diffraction data set was collected at the KEK Photon Factory to 2.29 Å resolution.

Crystallization and preliminary X-ray diffraction studies on the N-utilizing substance-B (NusB) from Mycobacterium tuberculosis

2000 ◽  
Vol 56 (1) ◽  
pp. 64-66 ◽  
Author(s):  
B. Gopal ◽  
R. A. Cox ◽  
M. J. Colston ◽  
G. G. Dodson ◽  
S. J. Smerdon ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Ribosomal Rna ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Core Complex ◽  
Unit Cell Parameters ◽  
Data Set ◽  
X Ray ◽  
Cell Parameters

N-utilizing substance B (NusB) is a protein which forms part of a complex assembly in transcriptional antitermination in Mycobacterium tuberculosis. It forms a heterodimer with the product of the NusE gene (identical to the ribosomal protein S10) and mediates the process of transcriptional antitermination by forming the core complex with the nut site of the ribosomal RNA along with other protein factors. NusB has been cloned and overexpressed in Escherichia coli and crystallized using the hanging-drop vapour-diffusion method. The space group is P212121, with unit-cell parameters a = 46.6, b = 64.2, c = 90.1 Å. A native data set complete to 1.6 Å resolution has been collected from a single crystal.

scholarly journals Crystallization and preliminary X-ray diffraction analysis of Trap1 complexed with Hsp90 inhibitors

2014 ◽  
Vol 70 (12) ◽  
pp. 1683-1687 ◽  
Author(s):  
Hanbin Jeong ◽  
Byoung Heon Kang ◽  
Changwook Lee
Keyword(s):  
Diffraction Analysis ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Hanging Drop ◽  
Hsp90 Inhibitors ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Middle Domain ◽  
Client Proteins

Hsp90 is a molecular chaperone responsible for the assembly and regulation of many cellular client proteins. In particular, Trap1, a mitochondrial Hsp90 homologue, plays a pivotal role in maintaining mitochondrial integrity, protecting against apoptosis in cancer cells. The N (N-terminal)-M (middle) domain of human Trap1 was crystallized in complex with Hsp90 inhibitors (PU-H71 and BIIB-021) by the hanging-drop vapour-diffusion method at pH 6.5 and 293 K using 15% PEG 8K as a precipitant. Diffraction data were collected from crystals of the Trap1–PU-H71 (2.7 Å) and Trap1–BIIB-021 (3.1 Å) complexes to high resolution at a synchrotron-radiation source. Preliminary X-ray diffraction analysis revealed that both crystals belonged to space groupP41212 orP43212, with unit-cell parametersa=b= 69.2,c= 252.5 Å, and contained one molecule per asymmetric unit according to Matthews coefficient calculations.

scholarly journals Crystallization of the C-terminal redox domain of the sulfur-assimilatory enzyme APR1 fromArabidopsis thaliana

2014 ◽  
Vol 70 (9) ◽  
pp. 1211-1214 ◽  
Author(s):  
Fang-Fang Chen ◽  
Yu-Yung Chang ◽  
Chao-Cheng Cho ◽  
Chun-Hua Hsu
Keyword(s):  
Biochemical Properties ◽  
Diffusion Method ◽  
X Ray Diffraction ◽  
Plant Type ◽  
Unit Cell Parameters ◽  
X Ray ◽  
Cell Parameters ◽  
Functional Studies ◽  
Aps Reductase ◽  
Crystal Volume

Plant-type APS reductase (APR), which catalyzes the reduction of activated sulfate to sulfite in plants, consists of a reductase domain and a C-terminal redox domain showing sequence homology to thioredoxin but possessing the activity of glutaredoxin. In order to understand the structural and biochemical properties of the redox domain of plant-type APS reductase, the C-terminal domain of APR1 (APR1C) fromArabidopsis thalianawas crystallized using the sitting-drop vapour-diffusion method. X-ray diffraction data were collected to a resolution of 2.70 Å on the SPXF beamline BL13B1 at the NSRRC, Taiwan. The crystals belonged to space groupP43212 orP41212, with unit-cell parametersa=b= 58.2,c= 86.7 Å. With one molecule per asymmetric unit, the crystal volume per unit protein weight (VM) is 2.64 Å3 Da−1, which corresponds to a solvent content of approximately 53.49%. Further structure-based functional studies of APR1C would extend knowledge of the molecular mechanism and regulation of APR.

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