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B C Wang

Publications and source records attributed to B C Wang.

At least 37 records · Page 2Linked to original sources
PubMed2000

Crystallization and preliminary X-ray diffraction analysis of the mitochondrial transcription factor sc-mtTFB from Saccharomyces cerevisiae.

Eukaryotic mitochondria contain a distinct mini-chromosome. In yeast, transcription of the mitochondrial genome is mediated by a nuclear-encoded RNA polymerase consisting of a single polypeptide core enzyme and a specificity factor termed sc-mtTFB which bears some similarity to bacterial sigma-factors. sc-mtTFB from Saccharomyces cerevisiae has been cloned, expressed, purified and crystallized. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 89.7, b = 44.6, c = 98.9 A, beta = 110 degrees. Based on one molecule per asymmetric unit, the solvent content is estimated to be 48%. Small crystals of dimensions 0.01 x 0.05 x 0.13 mm diffract to at least 2.7 A resolution on a rotating-anode X-ray source.

Crystallization↗
PubMed2000

Crystallization and preliminary X-ray analysis of the Clostridium thermocellum cellulosome xylanase Z feruloyl esterase domain.

Feruloyl esterases cleave ferulic acid from arabinoxylan and pectin. Feruloyl groups are believed to crosslink the polysaccharide chain within the polymer and to link hemicellulose to lignin, which may play a role in controlling the growth of plants. The Clostridium thermocellum cellulosome xylanase Z feruloyl esterase was expressed in Escherichia coli, purified and crystallized. The crystals diffract to 2.4 A resolution and belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 43.14, b = 63.77, c = 79.57 A. Assuming one molecule per asymmetric unit, the Matthews coefficient is calculated to be 1.87 A(3) Da(-1), which corresponds to a solvent content of 34%.

Carbohydrate Sequence↗
PubMed2000

Structure of the Ca2+-regulated photoprotein obelin at 1.7 A resolution determined directly from its sulfur substructure.

The crystal structure of the photoprotein obelin (22.2 kDa) from Obelia longissima has been determined and refined to 1.7 A resolution. Contrary to the prediction of a peroxide, the noncovalently bound substrate, coelenterazine, has only a single oxygen atom bound at the C2-position. The protein-coelenterazine 2-oxy complex observed in the crystals is photo-active because, in the presence of calcium ion, bioluminescence emission within the crystal is observed. This structure represents only the second de novo protein structure determined using the anomalous scattering signal of the sulfur substructure in the crystal. The method used here is theoretically different from that used for crambin in 1981 (4.72 kDa) and represents a significant advancement in protein crystal structure determination.

Aequorin↗
PubMed2000

[Determination of ibandronate by high performance ion exchange chromatography].

A method for the determination of Ibandronate by high performance ion exchange chromatography is reported. Ibandronate can form a complex compound with the copper ion added in the mobile phase. The complex can be detected at UV 240 nm. The effects of pH of mobile phase and concentration of methanol to the retention time were studied. Concentration of 10% methanol and pH 2.5 were suitable for the detection in less than 5 min. The flow rate was 1.0 mL/min. The relative standard deviation of peak area was 0.8%. The linear correlation was observed from 300 mg/L to 1,500 mg/L of Ibandronate (r = 0.9989). The lowest detectable concentration of Ibandronate was 6 mg/L. The method is simple, rapid and reproducible.

Bone Resorption↗
PubMed1999

Purification, crystallization and preliminary X-ray analysis of Drosophila melanogaster ferrochelatase.

Ferrochelatase (protoheme ferrolyase, E.C. 4.99.1.1), the terminal enzyme in the heme biosynthetic pathway, catalyzes the insertion of ferrous iron into protoporphyrin IX to form protoheme. In eukaryotes, the protein is associated with the inner surface of the inner mitochondrial membrane, and in higher animals the enzyme contains a [2Fe-2S] cluster. This cluster is highly sensitive to NO and is coordinated by four Cys residues whose spacing in the primary sequence is unique. Ferrochelatase from Drosophila melanogaster has been expressed in Escherichia coli with an amino-terminal six-histidine tag and purified to homogeneity. The protein has been crystallized with the [2Fe-2S] cluster intact. The crystals belong to space group I422, with unit-cell dimensions a = b = 158.1, c = 171.2 A and two molecules in the asymmetric unit, and diffract to 3. 0 A resolution.

Animals↗
PubMed1999

Low-salt crystallization of T7 RNA polymerase: a first step towards the transcription bubble complex.

DNA-dependent RNA polymerase is the key enzyme responsible for the biosynthesis of RNA, a process known as transcription. This process, which decodes the genetic information from DNA, is one of the most significant events in a biological system. The crystallization of both native and a chimeric T7/T3 RNAP using high salt conditions has been reported previously but these conditions proved unsuitable for DNA-RNAP complex formation since at high salt concentrations the DNA binding affinity to RNAP is reduced. A search for low-salt crystallization conditions has yielded new low-salt crystals of native T7-RNAP, a chimeric T7-RNAP (T7/T3 RNAP) which contains the T3 promoter recognition sequence, and a T7-RNAP containing an N-terminal histidine tag. The crystals, which are better suited for DNA-RNAP complex formation, belong to space group P3121 with a = 136, c = 156 A, contain a single molecule per asymmetric unit and diffract to 2.7 A resolution. Packing analysis shows that the new low-salt crystals have packing contacts similar to those observed in the high-salt T7-RNAP crystals reported previously. The diffraction anisotropicity observed in crystals of T7 RNAP is explained in term of crystal packing.

Base Sequence↗
PubMed1999

Preparation and preliminary study of crystals of the recombinant calcium-regulated photoprotein obelin from the bioluminescent hydroid Obelia longissima.

Crystals of recombinant obelin, the Ca(2+)-regulated photoprotein from the marine hydroid Obelia longissima, have been grown from sodium citrate solutions. Crystals grow as hexagonal light-yellow rods (0.1 x 0.1 x 1.0 mm) which diffract to beyond 1.8 A with synchrotron radiation of 1.0 A wavelength. The crystals have a primitive hexagonal lattice with unit-cell parameters a = 81.55, c = 86.95 A. The asymmetric unit contains two molecules. This represents the successful preparation of single crystals of a photoprotein obelin which have promising diffraction properties.

Animals↗
PubMed1999

Relationships within the aldehyde dehydrogenase extended family.

One hundred-forty-five full-length aldehyde dehydrogenase-related sequences were aligned to determine relationships within the aldehyde dehydrogenase (ALDH) extended family. The alignment reveals only four invariant residues: two glycines, a phenylalanine involved in NAD binding, and a glutamic acid that coordinates the nicotinamide ribose in certain E-NAD binary complex crystal structures, but which may also serve as a general base for the catalytic reaction. The cysteine that provides the catalytic thiol and its closest neighbor in space, an asparagine residue, are conserved in all ALDHs with demonstrated dehydrogenase activity. Sixteen residues are conserved in at least 95% of the sequences; 12 of these cluster into seven sequence motifs conserved in almost all ALDHs. These motifs cluster around the active site of the enzyme. Phylogenetic analysis of these ALDHs indicates at least 13 ALDH families, most of which have previously been identified but not grouped separately by alignment. ALDHs cluster into two main trunks of the phylogenetic tree. The largest, the "Class 3" trunk, contains mostly substrate-specific ALDH families, as well as the class 3 ALDH family itself. The other trunk, the "Class 1/2" trunk, contains mostly variable substrate ALDH families, including the class 1 and 2 ALDH families. Divergence of the substrate-specific ALDHs occurred earlier than the division between ALDHs with broad substrate specificities. A site on the World Wide Web has also been devoted to this alignment project.

Aldehyde Dehydrogenase↗
PubMed1999

Structural basis of neurophysin hormone specificity: Geometry, polarity, and polarizability in aromatic ring interactions.

The structural origins of the specificity of the neurophysin hormone-binding site for an aromatic residue in peptide position 2 were explored by analyzing the binding of a series of peptides in the context of the crystal structure of liganded neurophysin. A new modeling method for describing the van der Waals surface of binding sites assisted in the analysis. Particular attention was paid to the unusually large (5 kcal/mol) difference in binding free energy between Phe and Leu in position 2, a value representing more than three times the maximum expected based on hydrophobicity alone, and additionally remarkable since modeling indicated that the Leu side chain was readily accommodated by the binding pocket. Although evidence was obtained of a weak thermodynamic linkage between the binding interactions of the residue 2 side chain and of the peptide alpha-amino group, two factors are considered central. (1) The bound Leu side chain can establish only one-third of the van der Waals contacts available to a Phe side chain. (2) The bound Phe side chain appears to be additionally stabilized relative to Leu by more favorable dipole and induced dipole interactions with nonaromatic polar and sulfur ligands in the binding pocket, as evidenced by examination of its interactions in the pocket, analysis of the detailed energetics of transfer of Phe and Leu side chains from water to other phases, and comparison with thermodynamic and structural data for the binding of residue 1 side chains in this system. While such polar interactions of aromatic rings have been previously observed, the present results suggest their potential for significant thermodynamic contributions to protein structure and ligand recognition.

Amino Acids↗
PubMed1999

Pharmacology study of the neutral myocardial imaging agent technetium-99m-N(NOEt)2.

OBJECTIVE: The biological properties of a new neutral myocardial imaging agent 99mTcN(NOEt)2 were evaluated. METHODS: Blood clearance in rabbits, biodistribution in rats, and initial myocardial imaging in dogs were performed. RESULTS: Radiochemical purity of 99mTcN(NOEt)2 was more than 90% and stable for 6 h at room temperature. Blood disappearance was analyzed with a biexponential model and T1/2(alpha) = 2.53 min, T1/2(beta) = 330 min and CI = 378 ml/h were obtained. Biodistribution studies demonstrated that 99mTcN(NOEt)2 localized selectively in the rat myocardium. Cardiac uptakes were 4.69, 4.20, 3.95 and 3.43% ID/g at 5, 30, 60 and 90 min postinjection, respectively. The mean heart-to-lung activity ratios were 1.69, 2.40 and 2.55 at 10 min, 30 min and 60 min postinjection, respectively. CONCLUSION: Technetium-99m-N(NOEt)2 exhibited favorable stability and biological properties. Further study in humans is required.

Animals↗
PubMed1999

Increased phosphorylation of DARPP-32 by D1 agonistic action of l-stepholidine in the 6-OHDA-lesioned rat striatum.

In order to explore the characteristics of l-stepholidine (SPD) activating the postsynaptic D(1) receptors, the effects of SPD on DARPP-32 phosphorylation in vivo with back-phosphorylation assay and on the postsynaptic D(1) receptor densities with radioligand assay were observed in the striatum of 6-OHDA-lesioned rat. The results showed that following subcutaneously administration of 20 or 40 mg/kg SPD for 21 d, (32)P phosphate incorporation into the DARPP-32 protein in the denervated striatum showed a 50% reduction (P<0.01) vs the intact striatum, indicating an increase of DARPP-32 phosphorylation in vivo in the denervated striatum. However,the D(1) receptor B(max) was decreased from 385.0+/-26.1 to 319.7+/-20.1 fmol/mg protein. It is suggested that D(1) agonist action of SPD decreases the D(1) receptor density but increases the phosphorylation of DARPP-32 in the striatum of 6-OHDA-lesioned rat, which may be responsible for the regulation of D(1) receptor signal transduction in brain neurons.

Animals↗
PubMed1998

The structure of glutamine-binding protein complexed with glutamine at 1.94 A resolution: comparisons with other amino acid binding proteins.

The crystal structure of the glutamine-binding protein (GlnBP) complexed with its ligand (Gln) was determined and refined to 1.94 A resolution. This ellipsoidal protein has two globular domains and is approximately 52 Ax40 Ax35 A in size. The glutamine ligand is located in the cleft between the two domains and stablized by hydrogen bondings and ionic interactions with Asp10, Gly68, Thr70, Ala67, Asp157, Arg75, Lys115, Gly119 and His156. The aliphatic portion of the glutamine ligand is sandwiched in a hydrophobic pocket formed between Phe13 and Phe50 and has 21 van der Waals contacts with GlnBP. Lys115 and His156, that are unique to GlnBP among amino acid binding proteins, apparently contribute to the ligand binding specificity of GlnBP. Asp10 is within 3 A of Lys115. These two residues are over 10 A apart in the ligand-free form of the GlnBP. In addition, GlnBP-Gln exhibits a large-scale movement of the two hinges connecting the two globular domains upon ligand binding. The most significant changes are 41.1 degrees in the phi angle of Gly89 and 34.3 degrees in the psi angle of Glu181 from the first and the second hinge of the protein, respectively. Besides the original six hydrogen bonds, three extra hydrogen bonds can be observed between the two hinge strands upon ligand binding. A hydrogen bond network connects the large domain to the second hinge and a second hydrogen bond network coalesces the small domain to the same strand, both via interaction with the glutamine ligand. Although the two strands of the hinge connecting the domains do not directly participate in the ligand binding, Gln183 and Tyr185 from the second hinge may be involved in the cascade of the conformational change that is induced by ligand binding.

Amino Acid Sequence↗
PubMed1998

Structural modeling of the pro-ocytocin-neurophysin precursor.

The hormonal precursor pro-ocytocin-neurophysin is activated by selective cleavage at Arg2-Ala13, producing mature ocytocin and neurophysin. To understand the cleavage mechanism better, and in particular the recognition of the cleavage site, it is necessary to characterize the three-dimensional structure of the precursor molecule. Here we combine a variety of experimental data with molecular modeling and dynamics calculations to derive possible precursor conformations. In the models obtained, the N-terminus of the precursor, corresponding to the ocytocin segment, is hydrogen bonded in a pocket of the neurophysin moiety in a similar manner to a crystallographically obtained non-covalent complex between the two molecules. The calculations suggest that although the ocytocin segment is relatively flexible, it adopts a stable, broad loop structure in the vicinity of the cleavage region, which may constitute the structural element recognized by the cleaving enzyme. The calculations also suggest a possible widening of the distance between the two neurophysin domains in the precursor relative to that in the non-covalent neurophysin-ocytocin complex.

Amino Acid Sequence↗
PubMed1998

Enhancement of (-)-stepholidine on protein phosphorylation of a dopamine- and cAMP-regulated phosphoprotein in denervated striatum of oxidopamine-lesioned rats.

AIM: To study effects of (-)-stepholidine (SPD) on the phosphorylation of a dopamine- and cAMP-regulated phosphoprotein (DARPP-32) in the striatum of oxidopamine-lesioned rats. METHODS: The amount of dephospho-DARPP-32 was measured by a back-phosphorylation assay. RESULTS: In the striatum of control rats, SPD per se had no effect on the phosphorylation of DARPP-32, but it antagonized the decrease by 28% of dephospho-DARPP-32 induced by the D1 agonist SK&F-38393. In the denervated striatum of oxidopamine-lesioned rats, SPD decreased the amount of dephospho-DARPP-32 by 44%. The effect of SPD was completely counteracted by the concomitant administration of the D1 antagonist Sch-23390. CONCLUSION: SPD exhibits D1 agonistic action on DARPP-32 phosphorylation in the denervated striatum of oxidopamine-lesioned rats, but it acts as a D1 antagonist in normal striatum.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMed1997

In vitro and in vivo mechanical evaluations of plasma-sprayed hydroxyapatite coatings on titanium implants: the effect of coating characteristics.

This study was undertaken to evaluate the effect of coating characteristics on the mechanical strengths of the plasma-sprayed HA-coated Ti-6Al-4V implant system both in vitro and in vivo. Two types of HA coatings (HACs) with quite different microstructures, concentrations of impurity-phases, and indices-of-crystallinity were used. In vitro testings were done by measuring the bonding-strength at the Ti-6Al-4V-HAC interface, with HACs that had and had not been immersed in a pH-buffered, serum-added simulated body fluid (SBF). The shear-strength at the HAC-bone interface was investigated in a canine transcortical femoral model after 12 and 24 weeks of implantation. The results showed a bonding degradation of approximately 32% or higher of the original strength after 4 weeks of immersion in SBF, and this predominantly depended on the constructed microstructure of the HACs. After the push-out measurements, it was demonstrated that the HACs with higher bonding-strength in vitro would correspondingly result in significantly higher shear-strength at each implant period in vivo. Nevertheless, there were no substantial histological variations between the two types of HACs evaluated. The most important point elucidated in this study was that, among coating characteristics, the microstructure was the key factor in influencing the mechanical stability of the HACs both in vitro and in vivo. As a consequence, a denser HAC was needed to ensure mechanical stability at both interfaces.

Adhesiveness↗
PubMed1997

Crystallization and preliminary crystallographic data for the augmenter of liver regeneration.

A new cellular growth factor termed augmenter of liver regeneration (ALR) has been crystallized. ALR has been shown to have a proliferative effect on liver cells while at the same time producing an immunosuppressive effect on liver-resident natural killer cells and liver-resident mononuclear leukocytes. In addition, ALR appears to play an important role in the synthesis and stabilization of mitochondrial gene transcripts in actively regenerating cells. ALR crystals diffract to beyond 2 A resolution and belong to space group P2(1)2(1)2, with a = 125.1, b = 108.1 and c = 38.5 A. Based on four molecules per asymmetric unit, the Matthews coefficient is calculated to be 2.16 A(3) Da(-1) which corresponds to a solvent content of 43%.

Journal Article↗
PubMed1997

Intramedullary implant of plasma-sprayed hydroxyapatite coating: an interface study.

An intramedullary implant model in the canine femora was developed to evaluate the mechanical and histological responses between cancellous bone and plasma-sprayed hydroxyapatite coatings (HACs) on ti-6A1-4V implants, with 12- and 24-week follow-ups. HACs of different thicknesses were investigated. Results of the mechanical testings revealed that after 24 weeks of implantation, the mean shear strength (2.49 +/- 0.12 MPa) of the 50 microns HACs was significantly higher (p < 0.05) than that of the 200 microns HACs (1.44 +/- 0.19 MPa). However, using backscattered electron images (BEIs) throughout all the implant periods, no substantial histological variations in the extent of new bone apposition between the two HACs were observed. Occasionally, solution-mediated disintegration of the 50 microns HAC was found 24 weeks postimplantation. Histomorphometric studies from the BEIs demonstrated that for both HACs the percentage of the direct HAC-cancellous bone contact was approximately 50% at 12 weeks and 75% at 24 weeks. After the mechanical tests, the 200 microns HACs had fracture sites either inside the coating layers or at the HAC-titanium interfaces, which might explain why the mechanical performance of the 200 microns HACs was inferior to that of the 50 microns HACs even though both HACs had the same histological behaviors.

Alloys↗