• Title/Summary/Keyword: Poly(amino acid)

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Cloning and Functional Expression in Escherichia coli of the Polyhydroxyalkanoate Synthase (phaC) Gene from Alcaligenes sp. SH-69

  • Lee, Il;Nam, Sun-Woo;Rhee, Young-Ha;Kim, Jeong-Yoon
    • Journal of Microbiology and Biotechnology
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    • v.6 no.5
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    • pp.309-314
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    • 1996
  • Alcaligenes sp. SH-69 can synthesize poly(3-hydroxybutyrate-co-3-hydroxyvalerate) from a single carbon source such as glucose. To clone the phaC gene from Alcaligenes sp. SH-69, a polymerase chain reaction was performed using the oligomers synthesized based on the conserved regions of the phaC genes from other bacteria. A PCR product (550 bp) was partially sequenced and the deduced amino acid sequence was found to be homologous to that of the phaC gene from Alcaligenes eutrophus. Using the PCR fragment Southern blotting of Alcaligenes sp. SH-69 genomic DNA digested with several restriction enzymes was carried out. To prepare a partial genomic library, about 5-Kb genomic DNA fragments digested with EcoRI, which showed a positive signal in the Southern blotting, were eluted from an agarose gel, ligated with pUC19 cleaved with EcoRI, and transformed into Escherichia coli. The partial library was screened using the PCR fragment as a probe and a plasmid, named pPHA11, showing a strong hybridization signal was selected. Restriction mapping of the insert DNA in pPHA11 was performed. Cotransformation into E. coli of the plasmid pPHA11 and the plasmid pPHA21 which has phaA and phaB from A. eutrophus resulted in turbid E. coli colonies which are indicative of PHA accumulation. This result tells us that the Alcaligenes sp. SH-69 phaC gene in the pPHA11 is functionally active in E. coli and can synthesize PHA in the presence of the A. eutrophus phaA and phaB genes.

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Sequential Conjugation of 6-Aminohexanoic Acids and L-Arginines to Poly(amidoamine) Dendrimer to Modify Hydrophobicity and Flexibility of the Polymeric Gene Carrier

  • Yu, Gwang-Sig;Yu, Ha-Na;Choe, Yun-Hui;Son, Sang-Jae;Ha, Tai-Hwan;Choi, Joon-Sig
    • Bulletin of the Korean Chemical Society
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    • v.32 no.2
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    • pp.651-655
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    • 2011
  • We synthesized a novel cationic dendrimer consisting of a poly(amidoamine) dendrimer (PAMAM, generation 4) backbone with both L-arginine (Arg) at the termini and 6-aminohexanoic acid (Ahx) between the original core polymer and the peripheral Arg units. The sequential chemical modification of PAMAM G4 with Ahx and Arg resulted in higher transfection efficiency with much less cytotoxicity. PAMAM G4-Ahx-Arg formed stable polyplexes at weight ratios of 8:1 or higher (polymer: plasmid DNA), and the mean polyplex diameter was $180{\pm}20nm$. PAMAM G4-Ahx-Arg showed much higher transfection ability than PAMAM G4 or PAMAM G4-Ahx. Furthermore, PAMAM G4-Ahx-Arg was much less cytotoxic than PEI25KD and PAMAM G4-Arg. In addition to Arg grafting of the PAMAM dendrimer, which endows a higher transfection capability, the addition of Ahx spacer increased dendrimer hydrophobicity, introduced flexibility into the conjugated amino acids, and reduced cytotoxicity. Overall, it appears that the concomitant modification of PAMAM with Ahx and Arg could lead to new PAMAM conjugates with better performances.

Nucleotide Sequence Analyses of p10 Gene and its Promoter of Hyphantria cunea Nuclear Polyhedrosis Virus (Hyphantria cunea Nuclear Polyhedrosis Virus p10유전자와 프로모터의 염기서열 결정)

  • Park, Sun-A;Cha, Sung-Chul;Chang, Jae-Hyeok;Lee, Hyung-Hoan
    • The Journal of Korean Society of Virology
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    • v.26 no.1
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    • pp.131-137
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    • 1996
  • The sequences of p10 gene its promoter of Hyphantria cunea NPV were determined. According to the sequence analysis, the putative p10 gene ORF has 285 bp. The 5'-non-coding leader sequence of the p10 gene promoter contained the TATA box and the putative transcription initiation site TAAG motif. Poly (A) tail signals, AATAAA sequence was at site 65 base upstream from the 3' terminus. The deduced amino acid sequence of p10 protein was 95 with a predicted molecular weight of 10.26 kDa. In the p10 protein sequence, a hydrophobic region was present at the N-terminus of the protein, whereas the C-terminus was highly hydrophilic. The p10 protein of H. cunea NPV did not contain cysteine, histidine, trytophan, tryptophane, tyrosine, glutamine and asparagine residues.

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Characterization of a New Gene Resistant to Alkylating Agents and 3-Aminobenzamide When Knocked Out in Fission Yeast (분열형 효모에서 유전자 결실에 의해 알킬화제와 3-AMINOBENZAMIDE에 저항성을 나타내는 새로운 유전자의 특성 분석)

  • 박종군;차재영;황성진;박세근;김미영;백성민;최인순;이정섭
    • Journal of Life Science
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    • v.12 no.2
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    • pp.219-225
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    • 2002
  • The organization of eukayotic chromatin into specific conformation that are associated with transcription, replication, reapir and other nuclear processes are achieved via a series of DNA-protein interaction. These interactions are mediated by a range of DNA-binding domains such as SAP domain et at. By searching S. pombe genomic DNA database, we have found a gene named SAPuvs (SAP UV Sensitive) whose amino acid sequence is in part similar to SAP domain of Arabidopsis poly (ADP-ribose) polymerase and Ku7O. Knock-out cell of S. pombe SAPuvs gene was constructed using Ura4 as a selection marker. Survival analysis of knock-out cell indicated that treatment with UV significantly reduces the survival compared to wild type cell. Potentiation of MMS-induced cytotoxicity by 3AB post-treatment was observed in wild type cells, but not in knock-out cells. These data suggested that the protein encoded by SAPuvs gene is associated with chromatin reorganization during DNA repair.

Suppression of the Toll-like receptors 3 mediated pro-inflammatory gene expressions by progenitor cell differentiation and proliferation factor in chicken DF-1 cells

  • Hwang, Eunmi;Kim, Hyungkuen;Truong, Anh Duc;Kim, Sung-Jo;Song, Ki-Duk
    • Journal of Animal Science and Technology
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    • v.64 no.1
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    • pp.123-134
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    • 2022
  • Toll-like receptors (TLRs), as a part of innate immunity, plays an important role in detecting pathogenic molecular patterns (PAMPs) which are structural components or product of pathogens and initiate host defense systems or innate immunity. Precise negative feedback regulations of TLR signaling are important in maintaining homeostasis to prevent tissue damage by uncontrolled inflammation during innate immune responses. In this study, we identified and characterized the function of the pancreatic progenitor cell differentiation and proliferation factor (PPDPF) as a negative regulator for TLR signal-mediated inflammation in chicken. Bioinformatics analysis showed that the structure of chicken PPDPF evolutionarily conserved amino acid sequences with domains, i.e., SH3 binding sites and CDC-like kinase 2 (CLK2) binding sites, suggesting that relevant signaling pathways might contribute to suppression of inflammation. Our results showed that stimulation with polyinosinic:polycytidylic acids (Poly [I:C]), a synthetic agonist for TLR3 signaling, increased the mRNA expression of PPDPF in chicken fibroblasts DF-1 but not in chicken macrophage-like cells HD11. In addition, the expression of pro-inflammatory genes stimulated by Poly(I:C) were reduced in DF-1 cells which overexpress PPDPF. Future studies warrant to reveal the molecular mechanisms responsible for the anti-inflammatory capacity of PPDPF in chicken as well as a potential target for controlling viral resistance.

Cloning and Distribution of Facilitative Glucose Transporter 2 (SLC2A2) in Pigs

  • Zuo, Jianjun;Huang, Zhiyi;Zhi, Aimin;Zou, Shigeng;Zhou, Xiangyan;Dai, Fawen;Ye, Hui;Feng, Dingyuan
    • Asian-Australasian Journal of Animal Sciences
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    • v.23 no.9
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    • pp.1159-1165
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    • 2010
  • Glucose is the main energy source for mammalian cells and its absorption is co-mediated by two different families of glucose transporters, sodium/glucose co-transporters (SGLTs) and facilitative glucose transporters (GLUTs). Here, we report the cloning and tissue distribution of porcine GLUT2. The GLUT2 was cloned by RACE and its cDNA was 2,051 bp long (GenBank accession no. EF140874). An AAATAA consensus sequence at nucleotide positions 1936-1941 was located upstream of the poly $(A)^+$ tail. Open reading frame analysis suggested that porcine GLUT2 contained 524 amino acids, with molecular weight of 57 kDa. The amino acid sequence of porcine GLUT2 was 87% and 79.4% identical with human and mouse GLUT2, respectively. GLUT2 mRNA was detected at highest level in porcine liver, at moderate levels in the small intestine and kidney, and at low levels in the brain, lung, muscle and heart. In the small intestine, the highest level was in the jejunum. In conclusion, the mRNA expression of GLUT2 was not only differentially regulated by age, but also differentially distributed along the small intestine of piglets, which may be related to availability of different intestinal luminal substrate concentrations resulting from different food sources and digestibility.

Comparison of Colorless and Transparent Polyimide Films with Various Amine Monomers (다양한 아민 단량체를 이용한 무색 투명 폴리이미드 필름의 특성비교)

  • Kim, Youngmin;Chang, Jin-Hae
    • Applied Chemistry for Engineering
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    • v.23 no.3
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    • pp.266-270
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    • 2012
  • A series of polyimide was prepared by reacting 4,4'-(hexafluoroisopropylidene)diphthalic anhydride (6FDA) as the anhydride monomer and 2,2'-bis(trifluoromethyl)benzidine (TFB), 2,2'-bis(3-aminophenyl)hexafluoropropane (BAFP), 2,2'-bis(3- amino- 4-methylphenyl) hexafluoropropane (BAMF), bis(3-aminophenyl)sulfone (APS), p-xylyenediamine (p-XDA), or m-xylyenediamine (m-XDA) as the amine monomer in N,N-dimethylacetamide (DMAc). Colorless and transparent polyimide (PI) films were obtained by casting the poly(amic acid)s (PAAs) solution at various heat treatment temperatures. The thermal properties of the PI films were examined using differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), and thermomechanical analysis (TMA) and the mechanical properties were investigated using universal tensile machine (UTM), Their optical transparencies were also investigated using ultraviolet-visible (UV-vis.) spectrophotometry and colorimetry. The yellow index (YI) and coefficient of thermal expansion (CTE) values of all PIs were in the range 0.98~2.76 and 25.73~55.23 $ppm/^{\circ}C$, respectively.

Genetic Characterization of Encephalomyocarditis Virus Isolated from Aborted Swine Fetus in Korea

  • Song, Min-Suk;Joo, Young-Ho;Lee, Eun-Ho;Shin, Jin-Young;Kim, Chul-Jung;Shin, Kwang-Soon;Sung, Moon-Hee;Choi, Young-Ki
    • Journal of Microbiology and Biotechnology
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    • v.16 no.10
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    • pp.1570-1576
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    • 2006
  • An encephalomyocarditis virus (EMCV-CBNU) was isolated from an aborted swine fetus in October 2005. To investigate the genetic origin and virulence of the EMCV-CBNU strain, we determined the complete sequence of the virus and tested its virulence in mice. Genetic characterization revealed that the RNA genome was composed of 7,713 nucleotides with a single open reading frame (2,292 amino acids), coding 12 proteins. The EMCV-CBNU had the shortest poly(C) tract, consisting of 10 C's ($C_{10}$), compared with all the other EMCV strains reported in GenBank. Amino acid and phylogenetic analyses showed that EMCV-CBNU had the highest genetic identity with strain 2887A (99.7%), which was originally isolated from a fetus in a pig breeding farm that had a history of reproductive failure. Because rodents are the natural host of EMCV, we investigated the virulence of EMCV-CBNU in mice. Surprisingly, all mice inoculated with more than $1{\times}10^2\;TCID_{50}/0.1ml$ of EMCV-CBNU showed symptoms of hind limb paralysis and eventually died during 3 and 8 days postinoculation (DPI). Furthermore, when we inoculated the virus into pregnant mice, all dams and their fetuses died in 6 DPI. This is the first report on a full genomic analysis of swine EMCV in Korea, which exhibits high virulence in mice.

Isolation and Phylogenetic Analysis of Acyl-CoA-binding Protein Gene from Panax ginseng C.A. Meyer (인삼으로부터 Acyl-CoA-binding Protein 유전자의 동정 및 계통적 분석)

  • 인준교;류명현;최광태;최관삼;김세영;양덕춘
    • Korean Journal of Plant Tissue Culture
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    • v.28 no.4
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    • pp.201-204
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    • 2001
  • Acyl-CoA binding proteins (ACBP) are small highly-conserved cytosolic proteins that bind long-chain acyl-CoAs. A cDNA encoding ACBP was identified from cDNA library constructed from hairy root poly $A^{+}$ RNA in expressed sequence tags (EST) analysis. The cDNA clone was 453 bp long and carried an open reading frame of 264 bp (10 kDa). The ginseng ACBP amino acid sequence was compared with other reported plant ACBPs using the CLUSTALW. Ginseng ACBP is 89%, 81%, 80%, and 73% identical with ACBP from castor bean, lilly, Digitalis and Arabidopsis, respectively. However, ginseng ACBP is 5 amino acids smaller than Arabidopsis and rape seed ACBPs. Also there is no any known signal peptide sequence in ginseng ACBP.

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Cloning and Expression Characteristics of Pharbitis nil COP1 (PnCOP1) During the Floral Induction

  • Kim, Yun-Hee;Kim, Seong-Ryong;Heo, Yoon-Kang
    • Journal of Photoscience
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    • v.12 no.1
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    • pp.1-9
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    • 2005
  • The ubiquitin E3 ligase COP1 (Constitutive Photomorphogenesis 1) is a protein repressor of photomorphogenesis in Arabidopsisplants, and it found in various organisms, including animals. The COP1 protein regulates the stability of many of the light-signaling components that are involved in photomorphogenesis and in the developmental processes. To study the effect of COP1 on flowering in a short day plant, we have cloned a full-length of PnCOP1 (Pharbitis nil COP1) cDNA from Pharbitis nil Choisy cv. Violet, and we examined its transcript levels under various conditions. A full-length PnCOP1 cDNA consists of 2,280 bp nucleotidesthat contain 47 bp of 5'-UTR, 232 bp of 3'-UTR including the poly (A) tail, and 1,998 bp of the coding sequence. The deduced amino acid sequence contains 666 amino acids, giving it a theoretical molecular weight of 75 kD and a isolectric point of 6.2. The PnCOP1 contains three distinct domains, an N-terminal $Zn^2+$-binding RING-finger domain, a coiled-coil structure, and WD40 repeats at the C-terminal, implying that the protein plays a role in protein-protein interactions. The PnCOP1 transcript was detected in the cotyledon, hypocotyls and leaves, but not in root. The levels of the PnCOP1 transcript were reduced in leaves that were a farther distance away from the cotyledons. The expression level of the PnCOP1 gene was inhibited by light, while the expression was increased in the dark. During the floral inductive 16 hour-dark period for Pharbitis nil, the expression was increased and it reached its maximum at the 12th hour of the dark period. The levels of PnCOP1 mRNA were dramatically reduced upon light illumination. These results suggest that PnCOP1 may play an important function in the floral induction of Pharbitis nil.

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