• Title/Summary/Keyword: novel enzyme

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Knockdown of cytosolic NADP+-dependent isocitrate dehydrogenase enhances MPP+-induced oxidative injury in PC12 cells

  • Yang, Eun-Sun;Park, Jeen-Woo
    • BMB Reports
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    • v.44 no.5
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    • pp.312-316
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    • 2011
  • 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its toxic metabolite 1-methyl-4-phenylpyridium ion (MPP$^+$) have been shown to induce Parkinson's disease-like symptoms as well as neurotoxicity in humans and animal species. Recently, we reported that maintenance of redox balance and cellular defense against oxidative damage are primary functions of the novel antioxidant enzyme cytosolic NADP$^+$-dependent isocitrate dehydrogenase (IDPc). In this study, we examined the role of IDPc in cellular defense against MPP$^+$-induced oxidative injury using PC12 cells transfected with IDPc small interfering RNA (siRNA). Our results demonstrate that MPP$^+$-mediated disruption of cellular redox status, oxidative damage to cells, and apoptotic cell death were significantly enhanced by knockdown of IDPc.

Therapeutic Effect of Rebamipide on Ammonia-induced Gastric Mucosal Hemorrhagic Lesion in Rats

  • Huh, Keun;Kwon, Tae-Hyub;Kim, Soo-Kyun;Kim, Jin-Sook;Shin, Uk-Seob
    • Archives of Pharmacal Research
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    • v.21 no.1
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    • pp.1-5
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    • 1998
  • Rebamipide, 2-(4-chlorobenzoylamino)-3-[2(1H)-quinolinone-4-yl]-propionic acid, a novel antipeptic ulcer agent, has been reported to prevent various acute experimental gastric mucosal lesions and to accelerate the healing of chronic ulcers. Therapeutic effect of rebamipide was investigated with regard to the inhibitory effect on xanthine oxidase activity and type conversion of the enzyme which play a profound role in oxygen radicals generation system. Intraperitoneal administration of rebamipide at 60 mg/kg body weight reduced the xanthine oxidase activity, lipid peroxide content in ammonia induced hemorrhagic lesion. These results suggest that the therapeutic effect of rebamipide on gastric mucosal lesion may be in part due to the inhibitory activity of xanthine oxidase and type conversion rate of the enzyme.

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Development of E. coli Expression System to Overproduce a Harmful Protein, Carboxypeptidase Taq.

  • Lee, Sang-Hyeon
    • Journal of Life Science
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    • v.11 no.2
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    • pp.108-110
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    • 2001
  • The E. coli expression system to overproduce a harmful protein, carboxypeptidase Taq was developed. Since expression plasmid pCK305N containing the colicin promoter already has the initiation codon on the restriction site, the initiation codon of the CPase Taq gene was removed. Expression plasmid pCP4-col includes the entire CPase Taq gene, which is directed by the colicin promoter. E. coli cells harboring pCP-col produced a high amount of the enzyme when they were cultured in the present of mitomycin C (0.4 ${\mu}g$/ml). An amount of purified enzyme produced by pCP4-col directed by the colicin promoter was 10.5 mg. This result indicated that the novel E. coli expression system controlled by the colicin promoter could produce almost twice amounts of CPase Taq than the conventional system controlled by the tart promoter.

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Isolation of Sulfotransferase Producing Bacteria from Mouse Intestinal Microflora (생쥐의 장내미생물로부터 새로운 슬포트란스훼라제 생산균의 분리)

  • Kim, Byung-Taek;Kim, Eun-Ha;Kim, Dong-Hyun
    • YAKHAK HOEJI
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    • v.36 no.5
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    • pp.455-459
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    • 1992
  • Haemophilus K-12 producing novel sulfotransferase was isolated from mouce intestinal flora. The enzyme catalyzed the transfer of sulfate group from p-nitrophenylsulfate to phenolic compounds. The optimum medium condition for the sulfotransferase production was 0.2% sucrose, 1% yeast extract, $Na_{2}HPO_4$ and 0.5% NaCl. The enzyme production was induced by donor substrate, but was not by accepters. When p-nitrophenylsulfate was used as a donor substrate, 1-naphthol was best substrate, followed by phenol, p-acetaminophenol and tyramine.

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RFLP Analysis of cry1 and cry2 Genes of Bacillus thuringiensis Isolates from India

  • Patel, Ketan D.;Ingle, Sanjay S.
    • Journal of Microbiology and Biotechnology
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    • v.22 no.6
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    • pp.729-735
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    • 2012
  • The PCR-RFLP method has been useful for detection of known genes and identification of novel genes. In the present study, degenerate primers were designed from five groups of cry1 genes for PCR-RFLP analysis. Bacillus thuringiensis (Bt) isolates from different regions were evaluated for PCR amplification of various cry1 genes using newly designed primers and cry2 genes using reported primers. PCR analysis showed an abundance of cry1A genes and especially cry1Ac genes in isolates from all regions. RFLP analysis revealed the presence of multiple cry1A genes in isolates from central and southern regions. Unique digestion patterns of cry1A genes were observed in isolates from each region. Few of the isolates represented a digestion pattern of cry1A genes that did match to any of the known cry1A genes. RFLP analysis suggested an abundance of cry2Ab along with a novel cry2 gene in Bt isolates from different regions of India. Sequence analysis of the novel cry2 gene revealed 95% sequence identity to cry2Ab and cry2Ah genes. Phylogenetic analysis revealed that the novel cry2 gene could have diverged earlier than the other cry2 genes. Our results encourage finding of more diverse cry2 genes in Bt isolates. Rarefaction analysis was used to compare cry1A gene diversity in isolates from different soil types. It showed a higher degree of cry1A gene diversity in isolates from central region. In the present study, we propose the use of novel degenerate primers for cry1 genes and the PCR-RFLP method using a single enzyme to distinguish multiple cry1A and cry2 genes as well as identify novel genes.

Isolation and Characterization of a Novel Bacterium Burkholderia gladioli Bsp-1 Producing Alkaline Lipase

  • Zhu, Jing;Liu, Yanjing;Yanqin, Yanqin;Pan, Lixia;Li, Yi;Liang, Ge;Wang, Qingyan
    • Journal of Microbiology and Biotechnology
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    • v.29 no.7
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    • pp.1043-1052
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    • 2019
  • Active lipase-producing bacterium Burkholderia gladioli Bps-1 was rapidly isolated using a modified trypan blue and tetracycline, ampicillin plate. The electro-phoretically pure enzyme was obtained by purification using ethanol precipitation, ion-exchange chromatography, and gel filtration chromatography. The molecular weight was 34.6 kDa and the specific activity was determined to be 443.9 U/mg. The purified lipase showed the highest activity after hydrolysis with $p-NPC_{16}$ at a pH of 8.5 and $50^{\circ}C$, and the $K_m$, $k_{cat}$, and $k_{cat}/K_m$ values were 1.05 mM, $292.95s^{-1}$ and $279s^{-1}mM^{-1}$, respectively. The lipase was highly stable at $7.5{\leq}pH{\leq}10.0$. $K^+$ and $Na^+$ exerted activation effects on the lipase which had favorable tolerance to short-chain alcohols with its residual enzyme activity being 110% after being maintained in 30% ethanol for 1 h. The results demonstrated that the lipase produced by the strain B. gladioli Bps-1 has high enzyme activity and is an alkaline lipase. The lipase has promising chemical properties for a range of applications in the food-processing and detergent industries, and has particularly high potential for use in the manufacture of biodiesel.

P62 and the Sequestosome, a Novel Mechanism for Protein Metabolism

  • Shin, Jae-Kyoon
    • Archives of Pharmacal Research
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    • v.21 no.6
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    • pp.629-633
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    • 1998
  • In addition to selecting proteins for degradation by the 26S proteasome, ubiqitination appears to serve other regulatory functions, including for endosomal/lysosomal targeting, protein translocation, and enzyme modification. Currently, little is known how multiubiquitin chains are recognized by these cellular mechanisms. Within the 26S proteasome, one subunit (Mcb1/S5a) has been identified that has affinity for multiubiquitin chains and may function as a ubiquitin receptor. We recently found that a non-proteasomal protein p62 also preferentially binds multiubiquitin chains and forms a novel cytoplasmic structure "sequestosome" which serves as a storage place for ubiquitinated proteins. In the present manuscript, the role and regulation of p62 in relation to the sequestosomal function will be reviewed.

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Memory-Enhancing Effects of Silk Fibroin-Derived Peptides in Scopolamine-Treated Mice

  • Kang, Yong Koo;Lee, Woojoo;Kang, Byunghoon;Kang, Hannah
    • Journal of Microbiology and Biotechnology
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    • v.23 no.12
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    • pp.1779-1784
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    • 2013
  • Although enzyme-hydrolyzed silk fibroin has been reported to enhance cognitive function before, it has been still unknown which peptides can improve memory. Here we report that amino acid sequences of three novel peptides were identified from fibroin hydrolysate. Fibroin hydrolysate was obtained by hydrolysis with protease after partial hydrolysis with 5M $CaCl_2$. Synthesized peptides derived from these sequences improved scopolamine-induced memory impairments in mice. We confirmed this hydrolysate had effects that improved learning and memory abilities by performing the Rey-Kim test. From this hydrolysate of silk fibroin, amino acid sequences of eight peptides were identified by LC-MS/MS. Three peptides (GAGAGTGSSGFGPY, GAGAGSGAGSGAGAGSGAGAGY, and SGAGSGAGAGSGAGAGSGA) were synthesized to investigate whether they could improve memory. Passive avoidance test and Morris water maze test were performed, and all peptides showed memory-enhancing abilities on scopolamine-induced memory impairments in mice. In this study, we identified three novel peptides that could improve memory, and that silk fibroin hydrolysate was a mixture of various active peptides that could enhance memory.

cDNA Sequence and mRNA Expression of a Novel Serine Protease from the Firefly, Pyrocoelia rufa

  • Lee, Kwang-Sik;Kim, Seong-Ryul;Sohn, Hung-Dae;Jin, Byung-Rae
    • International Journal of Industrial Entomology and Biomaterials
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    • v.5 no.1
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    • pp.103-108
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    • 2002
  • We describe here the cDNA sequence and mRNA expression of a novel serine pretense from the firefly, Pyrocoelia rufa. The 771 bp cDNA encodes for 257 amino acid residues. The deduced protein of P. rufa serine pretense gene contains the catalytic triad and six-conserved cysteine residues. Alignment of the deduced protein of P. rufa serine pretense gene showed 47.4% protein sequence identity to known coleopteran insect Rhyzopertha dominica midgut trpsin-like enzyme. Northern blot analysis revealed that the P. rufa serine pretense is specifically expressed in the midgut of P. rufa larvae.

Design, Syntheses and Biological Evaluations of Nonpeptidic Caspase 3 Inhibitors

  • Kim, Eun-Sook;Yoo, Sung-Eun;Yi, Kyu-Yang;Lee, Sun-Kyung;Noh, Jae-Sung;Jung, Yong-Sam;Kim, Eun-Hee;Jeong, Nak-Chul
    • Bulletin of the Korean Chemical Society
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    • v.23 no.7
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    • pp.1003-1010
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    • 2002
  • Caspase 3, a member of cysteine protease family, is well known as a major apoptosis effector and is involved in cell death as a result of ischemic diseases such as stroke and myocardial infarction, therefore the inhibition of caspase 3 may protect those apoptotic cell damages. During the high-throughput screening of the compounds from the Korea Chemical Bank, berberine derivatives (A and B), an isoquinoline alkaloid, have been identified as potential inhibitors for caspase 3. Based on this finding we carried out molecular modeling study to identify the pharmacophoric elements of berberine structure which interact with a substrate-recognition binding site of caspase 3 and came up with several novel scaffolds. In this report, we will discuss the molecular modeling, syntheses and the enzyme inhibitory activities of these novel compounds.