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S C Kim

Publications and source records attributed to S C Kim.

At least 145 records · Page 8Linked to original sources
PubMed1998

Frame-shift mutation in hormone binding domain of human androgen receptor gene causes complete androgen insensitivity.

Mutation of the human androgen receptor gene impairs normal sexual differentiation and development in karyotypic males, resulting in a spectrum of external genital phenotypes ranging from complete female to nearly complete male. The androgen insensitivity syndrome (AIS) is an X-linked disorder in which genetic males fail to undergo normal fetal masculinization or pubertal virilization. PCR amplification of AR exons followed by SSCP analysis was performed with the genomic DNA of a patient having complete AIS. Mutation was observed at the exon E, where a single nucleotide deletion of T at the nucleotide 3286 was observed by cyclic sequencing. A single base deletion in Exon E in the hormone binding domain causes a frame-shift mutation, which leads to changes in the open reading frame, and causes the early termination of AR synthesis. The structural change of the hormone binding domain and AR might cause an insensitivity to the androgen thus leading to AIS.

Adult↗
PubMed1998

Cre/loxP-mediated in vivo excision of large segments from yeast genome and their amplification based on the 2microm plasmid-derived system.

In vivo excision and amplification of pre-determined, large genomic segments, directly from the genome of a natural host, provides an alternative to conventional cloning in foreign vectors. Using this approach, we have devised an in vivo procedure for excising large segments of Saccharomyces cerevisiae genome using Cre/loxP system of bacteriophage P1, followed by amplification of excised circles, as based on the yeast 2microm plasmid-derived ori and Flp/FRT machinery. To provide the excision and replication enzymes, trans-acting genes cre and FLP, which were under a very tight control of GAL1 and GAL10 promoters, respectively, were inserted by homologous recombination into the URA3 gene on chromosome V. Two parallel loxP sequences, which serve as the recognition sites for the Cre recombinase, were also integrated into the genome at pre-determined sites that are 50-100kb apart. Moreover, 2microm ori, REP3 and two inverted FRTs, which serve as a conditional replication system, were also integrated between the loxP sites. The strain carrying all these inserted elements was perfectly stable. Only after the induction by galactose of the Cre excision function, the genomic segment flanked by two loxP sites was excised and circularized. Applying this procedure, the 50-kb LEU2-YCR011c and 100-kb LEU2-YCR035c regions of chromosome III were successfully excised from the S. cerevisiae genome, whereas the 2microm ori, as aided by FRT/Flp, provided the amplification function. Such excised and amplified genomic segments can be used for the sequencing and functional analysis of any yeast genes.

Chromosomes↗
PubMed1998

Parasin I, an antimicrobial peptide derived from histone H2A in the catfish, Parasilurus asotus.

In response to epidermal injury, Parasilurus asotus, a catfish, secreted a strong antimicrobial peptide into the epithelial mucosal layer. The molecular mass of the antimicrobial peptide, named parasin I, was 2000.4 Da, as determined by matrix-associated laser desorption ionization mass spectrometry. The complete amino acid sequence of parasin I, which was determined by automated Edman degradation, was Lys-Gly-Arg-Gly-Lys-Gln-Gly-Gly-Lys-Val-Arg-Ala-Lys-Ala-Lys-Thr-Arg-Ser- Ser. Eighteen of the 19 residues in parasin I were identical to the N-terminal of buforin I, a 39-residue antimicrobial peptide derived from the N-terminal of toad histone H2A [Kim et al. (1996) Biochem. Biophys. Res. Commun. 229, 381-387], which implies that parasin I was cleaved off from the N-terminal of catfish histone H2A. Parasin I showed strong antimicrobial activity, about 12-100 times more potent than magainin 2, against a wide spectrum of microorganisms, without any hemolytic activity. Circular dichroism spectra of parasin I indicated a structural content of 11% alpha-helix, 33% beta-sheet, and 56% random coils. The beta-sheet axial projection diagram of parasin I showed an amphipathic structure. Our results indicate that the catfish may produce parasin I from its histone H2A by a specific protease upon injury to protect against invasion by microorganisms.

Amino Acid Sequence↗
PubMed1998

Lumbricin I, a novel proline-rich antimicrobial peptide from the earthworm: purification, cDNA cloning and molecular characterization.

A novel antimicrobial peptide was isolated and characterized from the earthworm, Lumbricus rubellus. The antimicrobial peptide was purified to homogeneity by a heparin-affinity column and C18 reverse-phase HPLC, and named lumbricin I. Lumbricin I was a proline-rich antimicrobial peptide of 62 amino acids (15% proline in molar ratio; molecular mass, 7231 Da), whose complete sequence was determined by a combination of peptide sequence and cDNA analysis. The peptide and cDNA sequence analysis revealed that lumbricin I was produced as a precursor form consisting of 76 amino acids, with 14 residues in a presegment and 62 residues in mature lumbricin I. Lumbricin I showed antimicrobial activity in vitro against a broad spectrum of microorganisms without hemolytic activity. In addition, a 29-amino acid peptide, named lumbricin I(6-34), which was derived from residues 6-34 of lumbricin I, showed marginally stronger antimicrobial activity than lumbricin I. Northern blot analysis on total RNA revealed that expression of lumbricin I gene was not induced by bacterial infection, but was constitutively expressed. Furthermore, the expression of lumbricin I gene was specific in adult L. rubellus: Lumbricin I mRNA was detected only in adult L. rubellus, but not in eggs and young L. rubellus.

Amino Acid Sequence↗
PubMed1998

Reduced microvascular thrombosis and improved outcome in acute murine stroke by inhibiting GP IIb/IIIa receptor-mediated platelet aggregation.

Treatment options in acute stroke are limited by a dearth of safe and effective regimens for recanalization of an occluded cerebrovascular tributary, as well as by the fact that patients present only after the occlusive event is established. We hypothesized that even if the site of major arterial occlusion is recanalized after stroke, microvascular thrombosis continues to occur at distal sites, reducing postischemic flow and contributing to ongoing neuronal death. To test this hypothesis, and to show that microvascular thrombosis occurs as an ongoing, dynamic process after the onset of stroke, we tested the effects of a potent antiplatelet agent given both before and after the onset of middle cerebral arterial (MCA) occlusion in a murine model of stroke. After 45 min of MCA occlusion and 23 h of reperfusion, fibrin accumulates in the ipsilateral cerebral hemisphere, based upon immunoblotting, and localizes to microvascular lumena, based upon immunostaining. In concordance with these data, there is a nearly threefold increase in the ipsilateral accumulation of 111In-labeled platelets in mice subjected to stroke compared with mice not subjected to stroke. When a novel inhibitor of the glycoprotein IIb/IIIa receptor (SDZ GPI 562) was administered immediately before MCA occlusion, platelet accumulation was reduced 48%, and fibrin accumulation was reduced by 47% by immunoblot densitometry. GPI 562 exhibited a dose-dependent reduction of cerebral infarct volumes measured by triphenyltetrazolium chloride staining, as well as improvement in postischemic cerebral blood flow, measured by laser doppler. GPI 562 caused a dose-dependent increase in tail vein bleeding time, but intracerebral hemorrhage (ICH) was not significantly increased at therapeutic doses; however, there was an increase in ICH at the highest doses tested. When given immediately after withdrawal of the MCA occluding suture, GPI 562 was shown to reduce cerebral infarct volumes by 70%. These data support the hypothesis that in ischemic regions of brain, microvascular thrombi continue to accumulate even after recanalization of the MCA, contributing to postischemic hypoperfusion and ongoing neuronal damage.

Animals↗
PubMed1998

Tissue-specific and androgen-repressible regulation of the rat dehydroepiandrosterone sulfotransferase gene promoter.

Dehydroepiandrosterone sulfotransferase (Std) catalyzes sulfonation of androgenic steroids and certain aromatic procarcinogens. In rats, this enzyme is selectively expressed in the liver, and its expression is strongly repressed by androgens. DNase I footprinting and electrophoretic mobility shift analyses revealed two hepatocyte nuclear factor-1 (HNF1), three CCAAT/enhancer-binding protein (C/EBP), and one consensus palindromic thyroid hormone response elements within the first 215 base pairs (bp) of the promoter sequence of rat Std. This promoter is normally inactive in fibroblast-derived NIH 3T3 cells. However, overexpression of HNF1 and C/EBP resulted in synergistic activation of the Std promoter in this cell type, indicating essential roles of these two trans-regulators in liver-selective expression of the rat Std gene. On the other hand, point mutations at any one of five cis elements proximal to the -215 bp region markedly reduced reporter gene expression, suggesting that all of these sites are important for overall promoter function. Androgenic repression of the Std gene in rat liver can be recapitulated in androgen receptor (AR)-negative HepG2 hepatoma cells after cotransfection with an AR expression plasmid. Functional assay of a nested set of 5'-deleted promoters mapped the negative androgen response region between positions -235 and -310. Antibody supershift and oligonucleotide competition identified three OCT-1 and two C/EBP elements between bp -231 and -292. An additional OCT-1 site was found to overlap with a C/EBP element at the -262/-252 position. Mutational inactivation of any one of five cis elements within the -231/-292 region abolished negative androgen response. However, none of these cis elements showed DNase I protection by recombinant AR in footprinting assay, suggesting the absence of a direct AR-DNA interaction. Thus, these studies on rat Std promoter function indicate that (i) HNF1 and C/EBP are responsible for liver specificity of the rat Std gene; (ii) androgenic repression of the gene requires the presence of all of the OCT-1 and C/EBP elements between positions -231 and -292; and (iii) AR may exert its negative regulatory effect indirectly through transcriptional interference of OCT-1 and C/EBP rather than through a direct DNA-AR interaction.

3T3 Cells↗
PubMed1998

Effect of the replication mode of a plasmid on the stability of multimeric endoxylanase genes in Bacillus subtilis.

Effect of the replication mode of a plasmid on the stability of tandemly multimerized endoxylanase genes and a gene dose-dependent expression of the endoxylanase were studied in Bacillus subtilis. The structural genes encoding an endoxylanase, carrying its original promoter and ribosomal binding sequence, were tandemly multimerized and cloned into the Escherichia coli-B. subtilis shuttle plasmid, pJH27 delta 88 or pMTL500e, which has a rolling circle-replicon or a theta (theta)-replicon in B. subtilis, respectively. The cloned dimers in pJH27 delta 88, which has a rolling circle-replicon, spontaneously rearranged to monomers in B. subtilis DB104, whereas those in pMTL500e, having a theta (theta)-replicon, were stably maintained. Expression level of the endoxylanase was proportional to the gene dosage in multimers. The endoxylanase activity in the supernatant increased from 80 U ml-1 with pMTL-1x containing a monomer of the gene to 165 U ml-1 with pMTL-4x containing a tetramer. These results indicate that high level expression of the endoxylanase gene can be obtained by tandemly multimerizing the genes in a plasmid with a theta (theta)-replicon.

Bacillus subtilis↗
PubMed1998

Antimicrobial peptides derived from pepsinogens in the stomach of the bullfrog, Rana catesbeiana.

Three antimicrobial peptides, which had strong antimicrobial activity against a broad spectrum of microorganisms, were isolated from the stomach of the bullfrog, Rana catesbeiana. Two of the antimicrobial peptides were found to be derived from the N-terminal sequences of pepsinogen A and C prosequences. The amino acid sequences of the new antimicrobial peptides, named bullfrog pepsinogen A-derived antimicrobial peptide (bPaAP) and bullfrog pepsinogen C-derived antimicrobial peptide (bPcAP), were Gly-Val-Val-Lys-Val-Ser-Arg-Leu-Lys-Gly-Glu-Ser-Leu-Arg-Ala-Arg-Leu (MW 1865.5) and Ile-Ile-Lys-Val-Pro-Leu-Lys-Lys-Phe-Lys-Ser-Met-Arg-Glu-Val-Met-Arg-A sp-His-Gly-Ile-Lys-Ala-Pro-Val-Val-Asp-Pro-Ala-Thr-Lys-Tyr (MW 3691.6), respectively. The bPaAP and bPcAP adopted 35% and 42% amphipathic alpha-helical structure in 50% trifluoroethanol, respectively, and were non-hemolytic up to a concentration of 200 microg/ml. Synthesized pepsinogen C prosequences of monkey and human, which had similar structural characteristics as bPaAP and bPcAP, also showed antimicrobial activity at concentrations of 10-200 microg/ml. The third peptide was buforin I, previously found in the stomach of the Asian toad, Bufo bufo gargarizans. These findings strongly suggest that peptides derived from the prosequences of pepsinogens, along with buforin I, may contribute to the antimicrobial function of the gastrointestinal mucosa of vertebrates, including human.

Amino Acid Sequence↗
PubMed1998

Molecular cloning and characterization of an endoxylanase gene of Bacillus sp. in Escherichia coli.

A gene encoding an endoxylanase of Bacillus sp. was cloned and expressed in Escherichia coli. The entire nucleotide sequence of a 1,620 bp SmaI fragment containing the endoxylanase gene was determined. The endoxylanase gene was 639 bp long and encoded 213 amino acids which showed up to 96% amino acid homology with other endoxylanases. The endoxylanase produced by E. coli harboring pKJX4 was purified by ion-exchange chromatography (DE-52 and CM-52) and its N-terminal sequence was determined to be Ala-Gly-Thr-Asp-Tyr-Trp-Gln-Asn-Trp-Thr-Asp-Gly-Gly-Gly-Thr. The endoxylanase expressed in E. coli was identical to that of the original Bacillus sp. whose molecular weight was approximately 20,400. Most of the produced endoxylanase was localized in the periplasmic space of E. coli. When the endoxylanase was reacted with 2% oat spelts xylan (w/v) at 40 degrees C for 10 h, the major product was xylobiose which is known to be a selective growth stimulant to one of the healthy intestinal microflora, Bifidobacteria.

Amino Acid Sequence↗
PubMed1998

Ginsenoside Rf, a trace component of ginseng root, produces antinociception in mice.

Ginseng root, a traditional oriental medicine, contains more than a dozen biologically active saponins called ginsenosides, including one present in only trace amounts called ginsenoside-Rf (Rf). Previously, we showed that Rf inhibits Ca2+ channels in mammalian sensory neurons through a mechanism requiring G-proteins, whereas a variety of other ginsenosides were relatively ineffective. Since inhibition of Ca2+ channels in sensory neurons contributes to antinociception by opioids, we tested for analgesic actions of Rf. We find dose-dependent antinociception by systemic administration of Rf in mice using two separate assays of tonic pain: in the acetic acid abdominal constriction test, the ED50 was 56+/-9 mg/kg, a concentration similar to those reported for aspirin and acetaminophen in the same assay; in the tonic phase of the biphasic formalin test, the ED50 was 129+/-32 mg/kg. Rf failed to affect nociception measured in three assays of acute pain: the acute phase of the formalin test, and the thermal (49 degrees C) tail-flick and increasing-temperature (3 degrees C/min) hot-plate tests. The simplest explanation is that Rf inhibits tonic pain without affecting acute pain, but other possibilities exist. Seeking a cellular explanation for the effect, we tested whether Rf suppresses Ca2+ channels on identified nociceptors. Inhibition was seen on large, but not small, nociceptors. This is inconsistent with a selective effect on tonic pain, so it seems unlikely that Ca2+ channel inhibition on primary sensory neurons can fully explain the behavioral antinociception we have demonstrated for Rf.

Abdominal Muscles↗
PubMed1998

Mechanism of action of the antimicrobial peptide buforin II: buforin II kills microorganisms by penetrating the cell membrane and inhibiting cellular functions.

The mechanism of action of buforin II, which is a 21-amino acid peptide with a potent antimicrobial activity against a broad range of microorganisms, was studied using fluorescein isothiocyanate (FITC)-labeled buforin II and a gel-retardation experiment. Its mechanism of action was compared with that of the well-characterized magainin 2, which has a pore-forming activity on the cell membrane. Buforin II killed Esche-richia coli without lysing the cell membrane even at 5 times minimal inhibitory concentration (MIC) at which buforin II reduced the viable cell numbers by 6 orders of magnitude. However, magainin 2 lysed the cell to death under the same condition. FITC-labeled buforin II was found to penetrate the cell membrane and accumulate inside E. coli even below its MIC, whereas FITC-labeled magainin 2 remained outside or on the cell wall even at its MIC. The gel-retardation experiment showed that buforin II bound to DNA and RNA of the cells over 20 times strongly than magainin 2. All these results indicate that buforin II inhibits the cellular functions by binding to DNA and RNA of cells after penetrating the cell membranes, resulting in the rapid cell death, which is quite different from that of magainin 2 even though they are structurally similar: a linear amphipathic alpha-helical peptide.

Anti-Bacterial Agents↗
PubMed1998

Acidic peptide-mediated expression of the antimicrobial peptide buforin II as tandem repeats in Escherichia coli.

Antimicrobial peptides have received increasing attention as a new pharmaceutical substance, because of their broad spectrum of antimicrobial activities and the rapid development of multidrug-resistant pathogenic microorganisms. The main obstacle to the wide application of antimicrobial peptides has been the lack of a cost-effective, mass-production method. A novel mass-production method for an antimicrobial peptide of 21 amino acids, buforin II, which was isolated from the stomach of the amphibian Bufo bufo gargarizans, has been developed. This method is based on the neutralization of the positive charges of buforin II by fusing to an acidic peptide to avoid the lethal effect of the expressed antimicrobial peptide on the host cells. The fusion peptide was expressed in Escherichia coli as tandem repeats to increase the product yield. Multimers of the acidic peptide-buforin II fusion peptide were expressed at high levels without causing damage to the cells. The presence of cysteine residues in the acidic peptide was critical for the high level expression of the fusion peptide multimers. Multimers of this fusion peptide were expressed as inclusion bodies, and about 107 mg of pure buforin II was obtained from 1 L of E. coli culture by cleaving the multimers with CNBr. Recombinant buforin II had an antimicrobial activity identical to that of natural buforin II. These results may lead to a general, cost-effective solution to the mass production of antimicrobial peptides and other basic peptides which are lethal to the host strain.

Amino Acid Sequence↗
PubMed1998

Imaging of the neurological complications of infective endocarditis.

We describe the findings on CT or MRI in five patients with neurological symptoms and underlying infective endocarditis (IE). We noted the size, number, and distribution of lesions, the presence or absence of haemorrhage, and contrast enhancement patterns. The number of lesions ranged from 4 to more than 10 in each patient. Their size varied from punctate to 6 cm; they were distributed throughout the brain. The lesions could be categorized into four patterns based on imaging features. A cortical infarct pattern was seen in all patients. Patchy lesions, which did not enhance, were found in the white matter or basal ganglia in three. Isolated, tiny, nodular or ring-enhancing white matter lesions were seen in three patients, and parenchymal haemorrhages in four. In addition to the occurrence of multiple lesions with various patterns in the same patient, isolated, tiny, enhancing lesions in the white matter seemed to be valuable features which could help to differentiate the neurological complications of IE from other thromboembolic infarcts.

Adult↗
PubMed1998

High-level expression of the angiotensin-converting-enzyme-inhibiting peptide, YG-1, as tandem multimers in Escherichia coli.

To produce a large quantity of the angiotensin-converting- enzyme(ACE)-inhibiting peptide YG-1, which consists of ten amino acids derived from yeast glyceraldehyde-3-phosphate dehydrogenase, a high-level expression was explored with tandem multimers of the YG-1 gene in Escherichia coli. The genes encoding YG-1 were tandemly multimerized to 9-mers, 18-mers and 27-mers, in which each of the repeating units in the tandem multimers was connected to the neighboring genes by a DNA linker encoding Pro-Gly-Arg for the cleavage of multimers by clostripain. The multimers were cloned into the expression vector pET-21b, and expressed in E. coli BL21(DE3) with isopropyl beta-D-thiogalactopyranoside induction. The expressed multimeric peptides encoded by the 9-mer, 18-mer and 27-mer accumulated intracellularly as inclusion bodies and comprised about 67%, 25% and 15% of the total proteins in E. coli respectively. The multimeric peptides expressed as inclusion bodies were cleaved with clostripain, and active monomers were purified to homogeneity by reversed-phase high-performance liquid chromatography. In total, 105 mg pure recombinant YG-1 was obtained from 11 E. coli culture harboring pETYG9 which contained the 9-mer of the YG-1 gene. The recombinant YG-1 was identical to the natural YG-1 in molecular mass, amino acid sequence and ACE-inhibiting activity.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1998

High-level expression of an endoxylanase gene from Bacillus sp. in Bacillus subtilis DB104 for the production of xylobiose from xylan.

To produce xylobiose from xylan, high-level expression of an endoxylanase gene from Bacillus sp. was carried out in Bacillus sabtilis DB104. A 1.62-kb SmaI DNA fragment, coding for an endoxylanase of Bacillus sp., was ligated into the Escherichia coli/B. subtilis shuttle vector pJH27 delta 88, producing pJHKJ4, which was subsequently transformed into B. subtilis DB104. A maximum endoxylanase activity of 105 U/ml was obtained from the supernatant of B. subtilis DB104 harboring pJHKJ4. The endoxylanase was purified to homogeneity by ion-exchange chromatography and the production profile of xylooligosaccharides from xylan by the endoxylanase was examined by HPLC with a carbohydrate analysis column. Xylobiose was the major product from xylan at 40 degrees C and its proportion in the xylan hydrolyzates increased with the reaction time; at 12 h, over 60% of the reaction products was xylobiose. These results suggest that xylobiose, which has a stimulatory effect on the selective growth of the intestinal bacterium Bifidobacterium, can be mass-produced effectively by the endoxylanase of Bacillus sp. cloned in B. subtilis.

Bacillus subtilis↗
PubMed1998

Efficacy and safety of fluoxetine, sertraline and clomipramine in patients with premature ejaculation: a double-blind, placebo controlled study.

PURPOSE: We compared the efficacy and safety of fluoxetine, sertraline, clomipramine and placebo for the oral pharmacotherapy of premature ejaculation. MATERIALS AND METHODS: The study included 36 men (mean age 44 years) who had intravaginal ejaculation latency of less than 2 minutes. Patients took each of 3 drugs and the placebo consecutively during a 4-week period per each agent. Efficacy and side effects data were obtained by a self-reported patient questionnaire that rated intravaginal ejaculation latency, sexual satisfaction of patient and partner, and possible side effects. RESULTS: After 4 weeks of treatment with placebo, fluoxetine, sertraline and clomipramine the mean intravaginal ejaculation latency time was significantly increased from 46 seconds to 2.27 minutes, 2.30 minutes, 4.27 minutes and 5.75 minutes, respectively (all p <0.01). However, treatment with clomipramine or sertraline caused a greater increase in mean intravaginal ejaculation latency time than fluoxetine or placebo (p <0.01). Patient sexual satisfaction rate after treatment with clomipramine was significantly higher (p <0.05) than with sertraline, fluoxetine or placebo. Partner sexual satisfaction rate was also higher with clomipramine than with sertraline or fluoxetine but no statistical difference was found. The incidence of side effects with clomipramine was significantly higher (p <0.05) compared to that of fluoxetine, sertraline and placebo, while no significant difference among sertraline, fluoxetine and placebo was noted. CONCLUSIONS: In men with premature ejaculation clomipramine was the most useful drug in terms of efficacy. Treatment with sertraline was nearly as effective and had a lower incidence of side effects.

1-Naphthylamine↗