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Biomedical subjects

J Moon

Publications and source records attributed to J Moon.

At least 37 records · Page 2Linked to original sources

Induction of G(1) cell cycle arrest and p27(KIP1) increase by panaxydol isolated from Panax ginseng.

Polyacetylenic compounds of Panax ginseng roots have been shown to inhibit growth of several human malignant tumor cell lines. Panaxydol is known to be one of the cytotoxic polyacetylenic compounds of P. ginseng. In this study, we first showed that panaxydol decreased markedly the proliferation, and to a lesser extent, the number of cells in a human melanoma cell line, SK-MEL-1. Next, the effect of panaxydol on cell cycle progression and its mechanism of action were investigated. Cell cycle analysis revealed that panaxydol inhibited cell cycle progression of a human malignant melanoma cell line, SK-MEL-1, at G(1)-S transition. At the same time, panaxydol increased the protein expression of p27(KIP1) as early as 1 hr after treatment. Cyclin-dependent kinase 2 (Cdk2) activity was decreased in a dose-dependent manner after 24 hr of panaxydol treatment. Protein levels of p21(WAF1), p16(INK4a), p53, pRb (retinoblastoma protein), and E2F-1 were not changed. It was also found that cycloheximide reversed the growth inhibition induced by panaxydol and partially abrogated the increase in p27(KIP1) expression. These results indicate that panaxydol induces G(1) cell cycle arrest by decreasing Cdk2 activity and up-regulating p27(KIP1) protein expression.

Alkynes↗

Crystal structure of NAD(+)-dependent DNA ligase: modular architecture and functional implications.

DNA ligases catalyze the crucial step of joining the breaks in duplex DNA during DNA replication, repair and recombination, utilizing either ATP or NAD(+) as a cofactor. Despite the difference in cofactor specificity and limited overall sequence similarity, the two classes of DNA ligase share basically the same catalytic mechanism. In this study, the crystal structure of an NAD(+)-dependent DNA ligase from Thermus filiformis, a 667 residue multidomain protein, has been determined by the multiwavelength anomalous diffraction (MAD) method. It reveals highly modular architecture and a unique circular arrangement of its four distinct domains. It also provides clues for protein flexibility and DNA-binding sites. A model for the multidomain ligase action involving large conformational changes is proposed.

Amino Acid Sequence↗

Overexpression, crystallization and preliminary X-ray crystallographic analysis of dihydrofolate reductase from bacteriophage T4.

Dihydrofolate reductase (DHFR) from bacteriophage T4 is a homodimer consisting of 193-residue subunits. It has been crystallized in the presence of the cofactor (NADPH) and an inhibitor (aminopterin) at 296 K using sodium chloride as precipitant. The crystals are tetragonal, belonging to the space group P4(1)22 (or P4(3)22), with unit-cell parameters a = b = 61.14, c = 123.23 A under cryogenic conditions. The asymmetric unit contains a single subunit, with a corresponding V(m) of 2.65 A(3) Da(-1) and a solvent content of 53. 6%. Native data have been collected from a crystal to 1.9 A resolution using synchrotron X-rays.

Bacteriophage T4↗

Crystallization and preliminary X-ray diffraction analysis of Saccharomyces cerevisiae Ygr203p, a homologue of Acr2 arsenate reductase.

Ygr203p, a 148-residue protein encoded by the ygr203w gene of Saccharomyces cerevisiae, is a homologue of the yeast Acr2 arsenate reductase encoded by the acr2 (or ypr200c) gene. It also shows significant sequence similarity to the human cell-cycle control Cdc25 phosphatase family. It has been overexpressed in soluble form in Escherichia coli with a His(6) tag at its C-terminus. The recombinant protein has been crystallized at 296 K using sodium chloride as precipitant. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 40.48, b = 50.95, c = 91.95 A. The asymmetric unit contains a monomer, giving a crystal volume per protein mass (V(m)) of 2.61 A(3) Da(-1) and a solvent content of 53.8%. The crystals diffract to better than 1.9 A resolution with Cu Kalpha X-rays. They are therefore suitable for high-resolution structure determination.

Adenosine Triphosphatases↗

Crystallization and preliminary X-ray crystallographic analysis of human nucleoside diphosphate kinase A.

Human nucleoside diphosphate kinase A catalyzes phosphoryl transfer and acts as a suppressor of metastasis. It has been crystallized using 2-methyl-2,4-pentanediol as a precipitant at 288 K. The crystal is monoclinic, belonging to the space group P2(1), with unit-cell parameters a = 74.21, b = 78.11, c = 82.29 A, beta = 101. 33 degrees. The asymmetric unit contains a homohexamer, with a corresponding crystal volume per protein mass (V(m)) of 2.27 A(3) Da(-1) and a solvent content of 46%. Native X-ray data to 2.15 A resolution have been collected using synchrotron X-rays.

Crystallization↗

Crystal structure of a 16 kDa double-headed Bowman-Birk trypsin inhibitor from barley seeds at 1.9 A resolution.

The Bowman-Birk trypsin inhibitor from barley seeds (BBBI) consists of 125 amino acid residues with two inhibitory loops. Its crystal structure in the free state has been determined by the multiwavelength anomalous diffraction (MAD) method and has been refined to a crystallographic R-value of 19.1 % for 8.0-1.9 A data. This is the first report on the structure of a 16 kDa double-headed Bowman-Birk inhibitor (BBI) from monocotyledonous plants and provides the highest resolution picture of a BBI to date. The BBBI structure consists of 11 beta-strands and the loops connecting these beta-strands but it lacks alpha-helices. BBBI folds into two compact domains of similar tertiary structure. Each domain shares the same overall fold with 8 kDa dicotyledonous BBIs. The five disulfide bridges in each domain are a subset of the seven disulfide bridges in 8 kDa dicotyledonous BBIs. Two buried water molecules form hydrogen bonds to backbone atoms in the core of each domain. One interesting feature of this two-domain inhibitor structure is that the two P1 residues (Arg17 and Arg76) are approximately 40 A apart, allowing the two reactive-site loops to bind to and to inhibit two trypsin molecules simultaneously and independently. The conformations of the reactive-site loops of BBBI are highly similar to those of other substrate-like inhibitors. This structure provides the framework for modeling of the 1:2 complex between BBBI and trypsin.

Amino Acid Sequence↗

Insights into eukaryotic multistep phosphorelay signal transduction revealed by the crystal structure of Ypd1p from Saccharomyces cerevisiae.

"Two-component" phosphorelay signal transduction systems constitute a potential target for antibacterial and antifungal agents, since they are found exclusively in prokaryotes and lower eukaryotes (yeast, fungi, slime mold, and plants) but not in mammalian organisms. Saccharomyces cerevisiae Ypd1p, a key intermediate in the osmosensing multistep phosphorelay signal transduction, catalyzes the phosphoryl group transfer between response regulators. Its 1.8 A structure, representing the first example of a eukaryotic phosphorelay protein, contains a four-helix bundle as in the HPt domain of Escherichia coli ArcB sensor kinase. However, Ypd1p has a 44-residue insertion between the last two helices of the helix bundle. The side-chain of His64, the site of phosphorylation, protrudes into the solvent. The structural resemblance between Ypd1p and ArcB HPt domain suggests that both prokaryotes and lower eukaryotes utilize the same basic protein fold for phosphorelay signal transduction. This study sheds light on the best characterized eukaryotic phosphorelay system.

Amino Acid Sequence↗

Heart rate variability of preterm neonates quantified by energy entropy.

Identifying variables predictive of neurobehavioural sequelae is a key objective in the study of high-risk neonates. Examination of heart rate variability (HRV) characteristics may be a finer discriminator of the neonate's response to physiologic stressors than the mean heart rate. The energy entropy of the heart beat tachogram, computed in four different domains, was used to quantify the HRV in 13 preterm neonates. The entropies of energies were computed from 1024 interbeat time intervals obtained once per week from 26 to 35 weeks postconceptional age (PCA). The energy entropy computed in three of the domains, like the standard deviation of intervals, distinguished between the 10 neonates that were measured at 35 weeks PCA with 100% specificity and 67% sensitivity, but did not distinguish between healthy and unhealthy neonates at earlier ages. The findings suggest that energy entropy may be a discerning measure of physiologic stress in the preterm infant, although future research is needed to refine the test and determine statistical significance.

Cardiotocography↗

Crystallization and preliminary X-ray crystallographic analysis of deoxycytidylate hydroxymethylase from bacteriophage T4.

Deoxycytidylate hydroxymethylase from bacteriophage T4 is a homodimeric enzyme in which each polypeptide chain consists of 246 amino-acid residues. It has been crystallized in the presence of its substrate, deoxycytidine monophosphate, at room temperature using sodium citrate as precipitant. The crystals are monoclinic, belonging to space group C2, with unit-cell parameters a = 174.22, b = 53.12, c = 75.17 A, beta = 115.29 degrees. The asymmetric unit contains one homodimer, with a corresponding Vm of 2.65 A3 Da-1 and solvent content of 54%. Native diffraction data to 1.6 A resolution have been collected from two crystals using synchrotron radiation.

Bacteriophage T4↗

Clinical significance of respiratory bronchiolitis on open lung biopsy and its relationship to smoking related interstitial lung disease.

BACKGROUND: Respiratory bronchiolitis-associated interstitial lung disease (RBILD) is a rare form of interstitial lung disease which may present in similar fashion to other types of chronic interstitial pneumonia. The purpose of this study was to undertake a clinicopathological review of 10 patients with RBILD and to examine the clinical and imaging data related to its histopathological pattern, in particular the relationship of RBILD to smoking. METHODS: Thirteen out of 168 retrospectively reviewed patients, from whom biopsy specimens were taken for suspected diffuse lung disease, were identified with a histopathological pattern of RBILD. Three cases were rejected as follow up data were unavailable. The 10 remaining cases constituted the study group and both clinical and imaging data were collected from patients' notes and referring physicians. RESULTS: Histopathologically, four cases of RBILD overlapped with the pattern of desquamative interstitial pneumonitis (DIP) and nine also had microscopic evidence of centrilobular emphysema. Nine patients were smokers, ranging from 3 to 80 pack years. The one non-smoker had an occupational exposure to the fumes of solder flux. The sex distribution was equal with an age range of 32-65 years. Two patients were clubbed. Lung function tests showed both restrictive and obstructive patterns together with severe reductions in carbon monoxide transfer factor in seven patients. Chest radiographs showed reticular or reticulonodular infiltrates in five patients and a ground glass pattern in two. CT scans were consistent with either DIP or RBILD in six of eight patients. Although seven patients remained stable or improved, either with or without treatment, three patients deteriorated. CONCLUSIONS: This study adds weight to the hypothesis that smoking can cause clinically significant interstitial lung disease, with deterioration in pulmonary function despite treatment. Given the overlapping histopathological patterns of RBILD and DIP and their strong association with smoking, the term "smoking related interstitial lung disease" is suggested for those patients who are smokers.

Adult↗

Characterization of the gene encoding catechol 2,3-dioxygenase from Achromobacter xylosoxidans KF701.

Catechol 2,3-dioxygenase (C23O) catalyzes a meta cleavage of the aromatic ring in catechol to form 2-hydroxymuconic semialdehyde. A C23O gene was cloned from chromosomal DNA of A. xylosoxidans KF701, a soil bacterium degrading biphenyl, and expressed in E. coli HB101. In substrate specificity to catechol and its analogs, the C23O exhibited the highest aromatic ring-fission activity to catechol, and its relative activity to other dihydroxylated aromatics was 4-chlorocatechol > 4-methylcatechol > 3-methylcatechol >> 2, 3-dihydroxybiphenyl. Aromatic ring-fission activity of the C23O to catechol was about 40-fold higher than that to 2,3-dihydroxybiphenyl. Nucleotide sequence analysis of the C23O gene from A. xylosoxidans KF701 revealed an open reading frame consisting of 924 base pairs, and identified a putative ribosome-binding sequence (AGGTGA) at about 10 nucleotides upstream from the initiation codon. The open reading frame can encode a polypeptide chain with molecular weight of 34 kDa containing 307 amino acid residues. The deduced amino acid sequence of the C23O exhibited the highest homology with that of C23O from Pseudomonas sp. IC with 96% identity, and the least homology with that of C23O from P. putida F1 with 22% identity among reported C23O sequences. Furthermore, comparison of the C23O sequence with other extradiol dioxygenases has led to identification of evolutionally conserved amino acid residues whose possible catalytic and structural roles are proposed.

Alcaligenes↗

Characterization of the gene encoding catechol 2,3-dioxygenase of Alcaligenes sp. KF711: overexpression, enzyme purification, and nucleotide sequencing.

Catechol 2,3-dioxygenase (C23O) is an extradiol-type dioxygenase that catalyzes the aromatic ring fission of catechol to form 2-hydroxymuconic semialdehyde. The C23O gene of Alcaligenes sp. KF711 was overexpressed in Escherichia coli HB101 by using the lac promoter of pUC18, and its gene product was purified by using immunoaffinity chromatography. The purified C23O exhibited a 35-kDa single band on an SDS-polyacrylamide gel, and its ring-fission activity on dihydroxylated aromatics was 4-methylcatechol > 4-chlorocatechol > catechol > 3-methylcatechol >> 2,3-dihydroxybiphenyl. Nucleotide sequence analysis of the C23O gene revealed an open reading frame of 927 bp, which can encode a polypeptide of 308 amino acid residues. The predicted molecular mass of 35 kDa is in agreement with that of purified C23O on an SDS-polyacrylamide gel. The amino acid sequence of the C23O was compared with those of nine other extradiol-type dioxygenases, including 2,3-dihydroxybiphenyl dioxygenase (2,3-DHBD) and 1,2-dihydroxynaphthalene dioxygenase (1,2-DHND). The C23O of Alcaligenes sp. KF711 exhibited 80 to 94% identity in amino acid sequence with other C23Os, and 20 to 25% identity with 1,2-DHND and 2,3-DHBDs. Furthermore, sequence comparison of 10 extradiol-type dioxygenases has led to identifying 19 evolutionarily conserved amino acid residues whose possible catalytic roles are proposed.

Alcaligenes↗

Hematopoietic potential of cryopreserved and ex vivo manipulated umbilical cord blood progenitor cells evaluated in vitro and in vivo.

The hematopoietic potential of cryopreserved and ex vivo manipulated umbilical cord blood (UCB) samples was evaluated in vitro and in vivo. Phenotypic analysis shows that approximately 1% of cord blood mononuclear cells express high levels of CD34 antigen on their surface (CD34hi), but none of a panel of lineage antigens (Lin-), suggesting that they are hematopoietic progenitor cells that have not yet committed to a specific lineage. Approximately 1% of CD34hi/Lin- cells are primitive hematopoietic progenitors that produce B lymphoid and multiple myeloid progeny for up to 7 weeks in stromal cell cultures. Twenty-one percent (+/- 13%) of CD34hi/Lin- cells also express low levels of the Thy-1 antigen and are threefold to fourfold enriched over CD34hi/Lin- cells in primitive hematopoietic potential as measured by long-term culture and phenotypic analysis. One-week liquid cultures of CD34-enriched UCB progenitor cells in the presence of interleukin (IL)-3, IL-6, and stem cell factor (SCF) results in a two-fold to threefold expansion of progenitors capable of reinitiating long-term stromal cell cultures. Only the CD34hi/Thy-1+/Lin- cell population was capable of maintaining progenitors with secondary transfer potential in long-term stromal cell cultures and is thus postulated to contain all of the primitive hematopoietic stem cells in UCB. The in vivo transplantation potential of UCB was also measured. Ex vivo manipulated UCB progenitor cells were used to engraft irradiated human thymus fragments implanted in severe combined immunodeficiency (SCID) mice. Thymic engraftment with >5% donor-derived cells and a normal CD4/CD8 distribution was observed in 19 of 23 tissues tested. UCB cells from in vitro expansion cultures engrafted with efficiencies comparable to nonexpanded cells. Similar results were obtained for UCB engraftment of human bone fragments implanted in SCID mice. In all cases, engraftment was achieved in competition with endogenous competitor stem cells and across major histocompatibility barriers. Taken together, this data demonstrates that human UCB is a rich source of multipotent hematopoietic progenitors that can be cryopreserved, enriched by physical methods, and expanded in a limited fashion without measurable loss of long-term culture or in vivo engrafting potential as measured in these assays.

Animals↗

Taurine intake of Korean breast-fed infants during lactation.

i. Taurine concentrations of human milk of nonvegetarians and lacto-ovovegetarians decreased significantly during the course of lactation. Taurine concentrations in lacto-ovovegetarians after 90 days postpartum were lower than those of nonvegetarians. Taurine concentrations of human milk by 150 days postpartum were: nonvegetarian 248-434 nmol/ml (31.0-54.4 mg/L); lacto-ovovegetarian 153-418 nmol/ml (19.1-52.3 mg/L). ii. Taurine intakes in infants of lacto-ovovegetarians decreased significantly during lactation, unlike from the pattern in infants of nonvegetarians. Taurine intakes in infants of lacto-ovovegetarians at 90, 120 and 150 days postpartum were lower, compared with those in nonvegetarians. Taurine intakes in infants by 150 days postpartum were: Infants of nonvegetarians 169-229 mumol/day (21.1-28.6 mg/day); Infants of lacto-ovovegetarians 106-210 mumol/day (21.1-28.6 mg/day). iii. This study showed that the taurine concentration in human milk and the taurine intake of infants were different during lactation between nonvegetarians and lacto-ovovegetarians, and that the intake per kg body weight decreased during lactation. Results suggest that the differences observed might be due to the effect of consumption of different types of food.

Breast Feeding↗

Isolation of a root-specific cDNA encoding a ns-LTP-like protein from the roots of bean (Phaseolus vulgaris L.) seedlings.

A root-specific cDNA clone, PVR3, was isolated from a bean (Phaseolus vulgaris L.) root cDNA library by a differential screening procedure. The nucleotide sequence of PVR3 contains an open reading frame coding for an 11.14 kDa polypeptide of 102 amino acid residues; the first 25 amino acids correspond to the sequence characteristic of a signal peptide. Comparison of the deduced PVR3 polypeptide sequence with the polypeptide sequences of previously cloned genes indicates that PVR3 may encode a ns-LTP-like protein. Molecular modelling of the PVR3 protein predicts that it has a three-dimensional structure that is similar to the three-dimensional model determined from the maize ns-LTP. The PVR3 mRNA accumulated mainly in the roots of young seedlings. It can be detected at low levels in flowers, but it is not detected in other organs. Genomic Southern blot analysis indicates that the genomic DNA corresponding to PVR3 cDNA is encoded by a single gene or small gene family in the bean genome.

Amino Acid Sequence↗