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REFERENCE LINKING PLATFORM OF KOREA S&T JOURNALS
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Korean Society for Biochemistry and Molecular Biology
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Volume & Issues
Volume 39, Issue 6 - Nov 2006
Volume 39, Issue 5 - Sep 2006
Volume 39, Issue 4 - Jul 2006
Volume 39, Issue 3 - May 2006
Volume 39, Issue 2 - Mar 2006
Volume 39, Issue 1 - Jan 2006
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Sphingosine Kinase: Biochemical and Cellular Regulation and Role in Disease
Taha, Tarek Assad ; Hannun, Yusuf Awni ; Obeid, Lina Marie ;
BMB Reports , volume 39, issue 2, 2006, Pages 113~131
DOI : 10.5483/BMBRep.2006.39.2.113
Sphingolipids have emerged as molecules whose metabolism is regulated leading to generation of bioactive products including ceramide, sphingosine, and sphingosine-1-phosphate. The balance between cellular levels of these bioactive products is increasingly recognized to be critical to cell regulation; whereby, ceramide and sphingosine cause apoptosis and growth arrest phenotypes, and sphingosine-1-phosphate mediates proliferative and angiogenic responses. Sphingosine kinase is a key enzyme in modulating the levels of these lipids and is emerging as an important and regulated enzyme. This review is geared at mechanisms of regulation of sphingosine kinase and the coming to light of its role in disease.
Vascular Endothelial Cadherin-mediated Cell-cell Adhesion Regulated by a Small GTPase, Rap1
Fukuhra, Shigetomo ; Sakurai, Atsuko ; Yamagishi, Akiko ; Sako, Keisuke ; Mochizuki, Naoki ;
BMB Reports , volume 39, issue 2, 2006, Pages 132~139
DOI : 10.5483/BMBRep.2006.39.2.132
Vascular endothelial cadherin (VE-cadherin), which belongs to the classical cadherin family, is localized at adherens junctions exclusively in vascular endothelial cells. Biochemical and biomechanical cues regulate the VE-cadherin adhesive potential by triggering the intracellular signals. VE-cadherin-mediated cell adhesion is required for cell survival and endothelial cell deadhesion is required for vascular development. It is therefore crucial to understand how VE-cadherin-based cell adhesion is controlled. This review summarizes the inter-endothelial cell adhesions and introduces our recent advance in Rap1-regulated VE-cadherin adhesion. A further analysis of the VE-cadherin recycling system will aid the understanding of cell adhesion/deadhesion mechanisms mediated by VE-cadherin in response to extracellular stimuli during development and angiogenesis.
Purification and Characterization of a Deoxyriboendonuclease from Mycobacterium smegmatis
Mandal, Prajna ; Chakraborty, Phulghuri ; Sau, Subrata ; Mandal, Nitai Chandra ;
BMB Reports , volume 39, issue 2, 2006, Pages 140~144
DOI : 10.5483/BMBRep.2006.39.2.140
A deoxyriboendonuclease has been purified to near homogeneity from a fast growing mycobacterium species, M. smegmatis and characterized to some extent. The size of enzyme is about 43 kDa as determined by a denaturing gel analysis. It shows optimum activity at
in Tris-HCl buffer (pH 7.2) containing 2.5 mM of
. Both EDTA and
inhibit its activity. Evidences show that the enzyme is not a restriction endonuclease but catalyzes the endonucleolytic cleavage of both the double- as well as the single-strand DNA non-specifically. It has been shown that the cleavage by this enzyme generates DNA fragments carrying phosphate groups at 5' ends and hydroxyl group at the 3' ends, respectively. Analysis reveals that no endonuclease having size and property identical to our deoxyriboendonuclease had been purified from M. smegmatis before. The property of our enzymes closely matches with the deoxyriboendonucleases purified from diverse sources including bacteria.
Maltol Inhibits Apoptosis of Human Neuroblastoma Cells Induced by Hydrogen Peroxide
Yang, Yang ; Wang, Jian ; Xu, Caimin ; Pan, Huazhen ; Zhang, Zinan ;
BMB Reports , volume 39, issue 2, 2006, Pages 145~149
DOI : 10.5483/BMBRep.2006.39.2.145
To analyze the effect of Maltol on the apoptosis of Human Neuroblastoma Cells (SH-SY5Y) treated by free radical which was generated from Hydrogen Peroxide (
), flow cytometry analysis on Phosphatidylserine (PS) inverting percentage was applied to determine the apoptosis. MTT (3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl-tetrazolium bromide) assay was employed to analyze the cell viability. DNA electrophoresis was used to detect DNA fragmentation. Moreover intracellular calcium of concentration (
) was measured by fluorescence emission. Flow cytometry analysis on the function of mitochondria and Western blto analysis of NF-
. The results showed that the pretreatment with maltol for 2 hours could prevent the
-induced apoptosis. Maltol could reduce the inverting percentage of PS, DNA fragmentation and
, and enhance the cellular function of mitochondria. NF-
is reduced. The experiments suggest that maltol could effectively inhibit the apoptosis induced by
. As a novel anti-oxidant, maltol is a new promising drug in protecting the neurological cells from the damage by free radical.
Substitution of Serine for Non-disulphide-bond-forming Cysteine in Grass Carp (Ctenopharygodon Idellus) Growth Hormone Improves In Vitro Oxidative Renaturation
Leung, Michael Yiu-Kwong ; Ho, Walter Kwok-Keung ;
BMB Reports , volume 39, issue 2, 2006, Pages 150~157
DOI : 10.5483/BMBRep.2006.39.2.150
Native grass carp (Ctenopharygodon idellus) growth hormone, has 5 cysteine amino acid residues, forms two disulphide bridges in its mature form. Recombinant grass carp growth hormone, when over-expressed in E. coli, forms inclusion bodies. In vitro oxidative renaturation of guanidine-hydrochloride dissolved recombinant grass carp growth hormone was achieved by sequential dilution and stepwise dialysis at pH 8.5. The redox potential of the refolding cocktail was maintained by glutathione disulphide/glutathione couple. The oxidative refolded protein is heterogeneous, and contains multimers, oligomers and monomers. The presence of non-disulphide-bond-forming cysteine in recombinant grass carp growth hormone enhances intermolecular disulphide bond formation and also non-native intramolecular disulphide bond formation during protein folding. The non-disulphide-bond-forming cysteine was converted to serine by PCR-mediated site-directed mutagenesis. The resulting 4-cysteine grass carp growth hormone has improved in vitro oxidative refolding properties when studied by gel filtration and reverse phase chromatography. The refolded 4-cysteine form has less hydrophobic aggregate and has only one monomeric isoform. Both refolded 4-cysteine and 5-cystiene forms are active in radioreceptor binding assay.
Molecular Cloning, Characterization and Expression of a Novel Trehalose-6-phosphate Synthase Homologue from Ginkgo biloba
Wu, Weisheng ; Pang, Yongzhen ; Shen, Guo-An ; Lu, Jie ; Lin, Juan ; Wang, Jin ; Sun, Xiaofen ; Tang, Kexuan ;
BMB Reports , volume 39, issue 2, 2006, Pages 158~166
DOI : 10.5483/BMBRep.2006.39.2.158
In many organisms, trehalose acts as protective metabolite against harsh environmental stresses, such as freezing, drought, nutrient starvation, heat and salt. Herein a cDNA (designated as GbTPS, GenBank Accession Number AY884150) encoding a trehalose-6-phosphate synthase homologue was isolated and characterized from the living fossil plant, Ginkgo biloba, which is highly tolerant to drought and cold. GbTPS encoded an 868-amino-acid polypeptide with a predicted isoelectric point of 5.83 and molecular mass of 97.9 kD. Amino acid sequence alignment revealed that GbTPS shared high identity with class II trehalose-6-phosphate synthase homologues (67% identical to AtTPS7), but had only 17% and 23% of identity with OstA from Escherichia coli and ScTPS1 from S. cerevisiae, respectively. DNA gel blot analysis indicated that GbTPS belonged to a small multi-gene family. The expression analysis by RT-PCR showed that GbTPS expressed in a tissue-specific manner in G biloba and might involve in leaf development. GbTPS was also found to be induced by a variety of stresses including cold, salt, drought and mannitol.
Transition State Characterization of the Low- to Physiological-Temperature Nondenaturational Conformational Change in Bovine Adenosine Deaminase by Slow Scan Rate Differential Scanning Calorimetry
Bodnar, Melissa A. ; Britt, B. Mark ;
BMB Reports , volume 39, issue 2, 2006, Pages 167~170
DOI : 10.5483/BMBRep.2006.39.2.167
Bovine adenosine deaminase undergoes a nondenaturational conformational change at
upon heating which is characterized by a large increase in heat capacity. We have determined the transition state thermodynamics of the conformational change using a novel application of differential scanning calorimetry (DSC) which employs very slow scan rates. DSC scans at the conventional, and arbitrary, scan rate of
show no evidence of the transition. Scan rates from 0.030 to
reveal the transition indicating it is under kinetic control. The transition temperature
and the transition temperature interval
increase with scan rate. A first order rate constant
is calculated at each
is the scan rate, and an Arrhenius plot is constructed. Standard transition state analysis reveals an activation free energy
of 88.1 kJ/mole and suggests that the conformational change has an unfolding quality that appears to be on the direct path to the physiological-temperature conformer.
Preliminary Proteomic Analysis of Indomethacin's Effect on Tumor Transplanted with Colorectal Cancer Cell in Nude Mice
Wang, Yu-Jie ; Zhang, Gui-Ying ; Xiao, Zhi-Qiang ; Wang, Hong-Mei ; Chen, Zhu-Chu ;
BMB Reports , volume 39, issue 2, 2006, Pages 171~177
DOI : 10.5483/BMBRep.2006.39.2.171
Nonsteroidal anti-inflammatory drugs such as indomethacin (IN) can exert anti-colorectal cancer (CRC) activity through cyclooxygenase independent mechanism, but the exactly biological mechanism is not completely known. Here we use proteomic tools to investigate the molecular mechanism of this action. First, nude mice bearing tumors derived from subcutaneous injection with human CRC cell line HCT116 were randomly allocated to groups treated with or without indomethacin. Later, tumor lumps were incised and then total proteins extracted. After separated with two-dimensional electrophoresis, thirty-one differently expressed spots were found between IN-treated and non-IN-treated groups, of which 25 spots decreased and 6 spots increased in abundance in IN-treated group. Through matrix-assisted laser desorption ionization time of flight mass spectrometry and then NCBInr and SWISS-PROT databases searching, 12 protein spots were finally identified including galectin-1, annexin A1, annexin IV, trancription factor BTF3A, calreticulin. Most of the identified proteins are correlated with tumor's biological prosperities of proliferation, invasion, apoptosis and immunity, or take part in cell's signal transduction. From above we thought that indomethacin can exert its effect on colorectal cancer through regulating several proteins' expression directly or indirectly. Further study of these proteins may be helpful in founding new targets of drugs for cancer chemotherapy.
Structure Analysis of 16S rDNA Sequences from Strains of Acidithiobacillus ferrooxidans
Peng, Hong ; Yang, Yu ; Li, Xuan ; Qiu, Guanzhou ; Liu, Xueduan ; Huang, Jufang ; Hu, Yuehua ;
BMB Reports , volume 39, issue 2, 2006, Pages 178~182
DOI : 10.5483/BMBRep.2006.39.2.178
Four strains of Acidithiobacillus ferrooxidans with different iron oxidation capacity were isolated from different mine drainage stations. The 16S rRNA gene of these strains were cloned and sequenced. Based on our sequences analysis on the four strain and the data on the other strains deposited in Genbank, all A. ferrooxidans may be classified into three phylogenetic groups. The analysis data showed that nucleotide variables (signature sites) were detected in 21 positions, and most of them were found in the first 800bp from 5' terminal except position 970 and 1375. Interestingly, the first 13 signature sites were located in two main regions:the first region (position 175-234) located in V2 while the second region (position 390-439) were detected in constant region between V2 and V3. Furthermore, the secondary structure and minimal free energy were determined in two regions among strains of three groups. These results may be useful in characterizing the microevolutionary mechanisms of species formation and monitoring in biohydrometallurgical application.
Confirming Single Nucleotide Polymorphisms from Expressed Sequence Tag Datasets Derived from Three Cattle cDNA Libraries
Lee, Seung-Hwan ; Park, Eung-Woo ; Cho, Yong-Min ; Lee, Ji-Woong ; Kim, Hyoung-Yong ; Lee, Jun-Heon ; Oh, Sung-Jong ; Cheong, Il-Cheong ; Yoon, Du-Hak ;
BMB Reports , volume 39, issue 2, 2006, Pages 183~188
DOI : 10.5483/BMBRep.2006.39.2.183
Using the Phred/Phrap/Polyphred/Consed pipeline established in the National Livestock Research Institute of Korea, we predicted candidate coding single nucleotide polymorphisms (cSNPs) from 7,600 expressed sequence tags (ESTs) derived from three cDNA libraries (liver, M. longissimus dorsi, and intermuscular fat) of Hanwoo (Korean native cattle) steers. From the 7,600 ESTs, 829 contigs comprising more than two EST reads were assembled using the Phrap assembler. Based on the contig analysis, 201 candidate cSNPs were identified in 129 contigs, in which transitions (69%) outnumbered transversions (31%). To verify whether the predicted cSNPs are real, 17 SNPs involved in lipid and energy metabolism were selected from the ESTs. Twelve of these were confirmed to be real while five were identified as artifacts, possibly due to expressed sequence tag sequence error. Further analysis of the 12 verified cSNPs was performed using the program BLASTX. Five were identified as nonsynonymous cSNPs, five were synonymous cSNPs, and two SNPs were located in 3'-UTRs. Our data indicated that a relatively high SNP prediction rate (71%) from a large EST database could produce abundant cSNPs rapidly, which can be used as valuable genetic markers in cattle.
Antibodies against Nitric Oxide Damaged Poly L-Tyrosine and 3-Nitrotyrosine Levels in Systemic Lupus Erythematosus
Khan, Fozia ; Ali, Rashid ;
BMB Reports , volume 39, issue 2, 2006, Pages 189~196
DOI : 10.5483/BMBRep.2006.39.2.189
Alterations in the amino acid structure or sequence can generate neo-epitopes from self-proteins causing autoaggressive immune attack. Reactive nitrogen species are an important factor that induces post-translational modification of proteins by cellular reduction and oxidation mechanism; cysteinyl-nitrosylation or tyrosine nitration leading to potentially pathogenic pathways. It was thought of interest to investigate the immunogenicity of nitrated poly L-tyrosine vis-
-vis its possible role in the induction of antibodies in systemic lupus erythematosus (SLE). Commercially available poly L-tyrosine was exposed to nitrating species and the damage was monitored by UV spectroscopy and alkaline gel electrophoresis. The results indicated the formation of 3-nitrotyrosine. Nitrated poly L-tyrosine induced higher titre antibodies as compared to the native form. Nitrated poly L-tyrosine was recognized by the autoantibodies present in the sera of patients suffering from SLE by enzyme immunoassays and band shift assay. The possible role of nitrated self-proteins has been discussed in the production of circulating anti-DNA antibodies in SLE.
Protective Effect of a 43 kD Protein from the Leaves of the Herb, Cajanus indicus L on Chloroform Induced Hepatic-disorder
Ghosh, Ayantika ; Sarkar, Kasturi ; Sil, Parames C. ;
BMB Reports , volume 39, issue 2, 2006, Pages 197~207
DOI : 10.5483/BMBRep.2006.39.2.197
Cajanus indicus is a herb with medicinal properties and is traditionally used to treat various forms of liver disorders. Present study aimed to evaluate the effect of a 43 kD protein isolated from the leaves of this herb against chloroform induced hepatotoxicity. Male albino mice were intraperitoneally treated with 2mg/kg body weight of the protein for 5 days followed by oral application of chloroform (0.75ml/kg body weight) for 2 days. Different biochemical parameters related to physiology and pathophysiology of liver, such as, serum glutamate pyruvate transaminase and alkaline phosphatase were determined in the murine sera under various experimental conditions. Direct antioxidant role of the protein was also determined from its reaction with Diphenyl picryl hydraxyl radical, superoxide radical and hydrogen peroxide. To find out the mode of action of this protein against chloroform induced liver damage, levels of antioxidant enzymes catalase, superoxide dismutase and glutathione-S-transferase were measured from liver homogenates. Peroxidation of membrane lipids both in vivo and in vitro were also measured as malonaldialdehyde. Finally, histopathological analyses were done from liver sections of control, toxin treated and protein pre- and post-treated (along with the toxin) mice. Levels of serum glutamate pyruvate transaminase and alkaline phosphatase, which showed an elevation in chloroform induced hepatic damage, were brought down near to the normal levels with the protein pretreatment. On the contrary, the levels of anti-oxidant enzymes such as catalase, superoxide dismutase and glutathione-S-transferase that had gone down in mice orally fed with chloroform were significantly elevated in protein pretreated ones. Besides, chloroform induced lipid peroxidation was effectively reduced by protein treatment both in vivo and in vitro. In cell free system the protein effectively quenched diphenyl picryl hydrazyl radical and superoxide radical, though it could not catalyse the breakdown of hydrogen peroxide. Post treatment with the protein for 3 days after 2 days of chloroform administration showed similar results. Histopathological studies indicated that chloroform induced extensive tissue damage was less severe in the mice livers treated with the 43 kD protein prior and post to the toxin administration. Results from all these data suggest that the protein possesses both preventive and curative role against chloroform induced hepatotoxicity and probably acts by an anti-oxidative defense mechanism.
Immunohistochemical Studies of Human Ribosomal Protein S3 (rpS3)
Choi, Soo-Hyun ; Kim, So-Young ; An, Jae-Jin ; Lee, Sun-Hwa ; Kim, Dae-Won ; Won, Moo-Ho ; Kang, Tae-Cheon ; Park, Jin-Seu ; Eum, Won-Sik ; Kim, Joon ; Choi, Soo-Young ;
BMB Reports , volume 39, issue 2, 2006, Pages 208~215
DOI : 10.5483/BMBRep.2006.39.2.208
The human ribosomal protein S3 (rpS3) was expressed in E. coli using the pET-I5b vector and the monoclonal antibodies (mAbs) were produced and characterized. A total of five hybridoma cell lines were established and the antibodies recognized a single band of molecular weight of 33 kDa on immunoblot with purified rpS3. When the purified rpS3 was incubated with the mAbs, the UV endonuclease activity of rpS3 was inhibited up to a maximum of 49%. The binding affinity of mAbs to rpS3 determined by using a biosensor technology showed that they have similar binding affinities. Using the anti-rpS3 antibodies as probes, we investigated the cross-reactivities of various other mammalian brain tissues and cell lines, including human. The immunoreactive bands on Western blots appeared to be the same molecular mass of 33 kDa in all animal species tested. They also appear to be extensively cross-reactive among different organs in rat. These results demonstrated that only one type of immunologically similar rpS3 protein is present in all of the mammalian brain tissues including human. Furthermore, these antibodies were successfully applied in immunohistochemistry in order to detect rpS3 in the gerbil brain tissues. Among the various regions in the brain tissues, the rpS3 positive neurons were predominantly observed in the ependymal cells, hippocampus and substantia nigra pars compacta. The different distributions of rpS3 in brain tissues reply that rpS3 protein may play an important second function in the neuronal cells.
Solubilization of Proteins from Human Lymph Node Tissue and Two-Dimensional Gel Storage
De Marqui, Alessandra Bernadete Trovo ; Vidotto, Alessandra ; Polachini, Giovana Mussi ; De Mattos Bellato, Claudia ; Cabral, Hamilton ; Leopoldino, Andreia Machado ; De Gois Filho, Jose Francisco ; Fukuyama, Erica Erina ; Settanni, Flavio Aurelio Parente ; Cury, Patricia Maluf ; Bonilla-Rodriguez, Gustavo Orlando ; Palma, Mario Sergio ; Tajara, Eloiza Helena ;
BMB Reports , volume 39, issue 2, 2006, Pages 216~222
DOI : 10.5483/BMBRep.2006.39.2.216
In the present study, we compared six different solubilization buffers and optimized two-dimensional electrophoresis (2-DE) conditions for human lymph node proteins. In addition, we developed a simple protocol for 2-D gel storage. Efficient solubilization was obtained with lysis buffers containing (a) 8M urea, 4% CHAPS (3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate), 40 mM Tris base, 65 mM DTT(dithiothreitol) and 0.2% carrier ampholytes; (b) 5M urea, 2M thiourea, 2% CHAPS, 2% SB 3-10 (N-decyl-N, N-dimethyl-3-ammonio-1-propanesulfonate), 40mM Tris base, 65 mM DTT and 0.2% carrier ampholytes or (c) 7M urea, 2M thiourea, 4% CHAPS, 65 mM DTT and 0.2% carrier ampholytes. The optimal protocol for isoelectric focusing (IEF) was accumulated voltage of 16,500 Vh and 0.6% DTT in the rehydration solution. In the experiments conducted for the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), best results were obtained with a doubled concentration (50 mM Tris, 384 mM glycine, 0.2% SDS) of the SDS electrophoresis buffer in the cathodic reservoir as compared to the concentration in the anodic reservoir (25 mM Tris, 192 mM glycine, 0.1% SDS). Among the five protocols tested for gel storing, success was attained when the gels were stored in plastic bags with 50% glycerol. This is the first report describing the successful solubilization and 2D-electrophoresis of proteins from human lymph node tissue and a 2-D gel storage protocol for easy gel handling before mass spectrometry (MS) analysis.
Activity of Human Dihydrolipoamide Dehydrogenase Is Largely Reduced by Mutation at Isoleucine-51 to Alanine
Kim, Hak-Jung ;
BMB Reports , volume 39, issue 2, 2006, Pages 223~227
DOI : 10.5483/BMBRep.2006.39.2.223
Dihydrolipoamide dehydrogenase (E3) belongs to the pyridine nucleotide-disulfide oxidoreductase family including glutathione reductase and thioredoxin reductase. It catalyzes the reoxidation of dihydrolipoyl moiety of the acyltransferase components of three
-keto acid dehydrogenase complexes and of the hydrogen-carrier protein of the glycine cleavage system. Isoleucine-51 of human E3, located near the active disulfide center Cys residues, is highly conserved in most E3s from several sources. To examine the importance of this highly conserved Ile-51 in human E3 function, it was substituted with Ala using site-directed mutagenesis. The mutant was expressed in Escherichia coli and highly purified using an affinity column. Its E3 activity was decreased about 100-fold, indicating that the conservation of the Ile-51 residue in human E3 was very important to the efficient catalytic function of the enzyme. Its altered spectroscopic properties implied that conformational changes could occur in the mutant.