• Applied Biological Chemistry
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• v.40 no.5
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• pp.380-387
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• 1997

Phytopathogenic Erwinia carotovora subsp. carotovora (Ecc) LY34 causes plant tissue maceration by secretion of pectinolytic enzymes such as pectate Iyase (PL) existed as multiple isoenzyme form. Genomic DNA from Ecc LY34 was digested with Sau3Al and ligated into the BamHI site of pBluescript ll $SK^+$. Among them, a clone hydrolyzing polypectate was selected and its DNA was digested with BamHI. Through the subsequent subcloning the resulting 3.1 kb fragment, corresponding to a peICI, was subcloned into pLYPA 100. The structural organization of a peICI gene encoding a 374 amino acid residues consists of an open reading frame (ORF) of 1,122 bp commencing with a ATG start codon and followed by a TAA stop codon. PeICI contained a typical prokaryotic signal peptide of 22-amino acid. Since the deduced amino acid sequences of PeICl protein was very similar to those of PelIII of Erwinia carotovora subsp. carotovora, and to those of Pel3 of Erwinia carotovora subsp. atroseptica, and to those of PeIC of Erwinia carotovora subsp. carotovora, it belong to the same family PLbc group. The 374-amino acld PeICI had a calculated Mr of 40,507 and pI of 7.60.

• BMB Reports
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• v.35 no.2
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• pp.213-219
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• 2002

Malonyl-CoA decarboxylase (E.C.4.1.1.9) catalyzes the conversion of malonyl-CoA to acetyl-CoA. Although the metabolic role of this enzyme has not been fully defined, it has been reported that its deficiency is associated with mild mental retardation, seizures, hypotonia, cadiomyopathy, developmental delay, vomiting, hypoglycemia, metabolic acidosis, and malonic aciduria. Here, we isolated a cDNA clone for malonyl CoA decarboxylase from a rat brain cDNA library, expressed it in E. coli, and characterized its biochemical properties. The full-length cDNA contained a single open-reading frame that encoded 491 amino acid residues with a calculated molecular weight of 54, 762 Da. Its deduced amino acid sequence revealed a 65.6% identity to that from the goose uropigial gland. The sequence of the first 38 amino acids represents a putative mitochondrial targeting sequence, and the last 3 amino acid sequences (SKL) represent peroxisomal targeting ones. The expression of malonyl CoA decarboxylase was observed over a wide range of tissues as a single transcript of 2.0 kb in size. The recombinant protein that was expressed in E. coli was used to characterize the biochemical properties, which showed a typical Michaelis-Menten substrate saturation pattern. The $K_m$ and $V_{max}$ were calculated to be $68\;{\mu}M$ and $42.6\;{\mu}mol/min/mg$, respectively.

• Journal of the Korean Institute of Telematics and Electronics C
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• v.36C no.9
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• pp.1-12
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• 1999

In this paper, we designed an embedded processor with 128Kbytes EPROM and 4Kbytes SRAM based on GMS30C2132 which RISC processor with DSP functions. And a new architecture of bus sharing to control the embedded memory and external memory unit i proposed aiming at one-cycle access between memories and CPU. For embedded 128Kbytes EPROM, we designed the new expansion interface for data size at data ordering with memory organization and the efficient interface for test. The embedded SRAM supports an extended stack area high speed DSP operation, instruction cache and variable data-length control which is accessed with 4K modulo addressing schemes. The proposed new architecture and circuits reduced the memory access cycle time from 40ns and improved operation speed 2-times for program benchmark test. The chip is occupied $108.68mm^2$ using $0.6{\mu}m$ CMOS technology.

• Korean Journal Plant Pathology
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• v.10 no.3
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• pp.228-234
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• 1994

Genomic RNA was extracted from Cy strain of odontoglossum ringspot tobamovirus (ORSV-Cy) isolated from infected leaves of tobacco cv. Samsun. Size of the genomic RNA was about 6.6 kb in length. The genomic RNA was fractionated using Sephadex G-50 column chromatography into 2 fractions. They were polyadenylated at their 3'-end using E. coli poly(A) polymerase. Polyadenylated viral RNA was recovered by oligo (dT) primer adapter containing NotI restriction site and Moloney murine leukemia virus SuperScript reverse transcriptase (RNase H-). Second-strand cDNA was synthesized by using E. coli DNA ligase, E. coli DNA polymerase I and E. coli RNase H. Recombinant plasmids containing cDNAs for ORSV-Cy RNA ranged from about 800 bp to 3,000 bp. Among the selected 238 recombinants, pORCY-124 clone was the largest one covering 3'-terminal half of the viral RNA. This clone contained two restriction sites for EcoRI and XbaI and one site for AccI, AvaI, BglII, BstXI, HindIII, PstI, and TthIII 1. respectively. The clone contained partial viral replicase, a full-length movement protein and a complete coat protein genes followed by a 3' untranslated region of 414 nucleotides based on restriction mapping and nucleotide sequencing analyses. Clones pORCY-028, -068, -072, -187 and -224 were overlapped with the pORCY-124. Clones pORCY-014 and -095 covered 5' half upstream from the middle region of the viral RNA, which was estimated based on restriction mapping and partial sequence analysis. Constructed cDNA library covered more than 90% of the viral genome.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.35 no.1
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• pp.1-12
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• 2013

Purpose: Vascular endothelial growth factor (VEGF)-C and its tyrosine kinase receptor, VEGF receptor (VEGFR)-3 are recently known to have lymphangiogenic activities in various tumor types. In this study, we determined whether the expression of lymphangiogenic factors correlate with nodal metastasis or survival in a nude mouse model of oral squamous cell carcinoma (OSCC). Methods: Three OSCC cells (KB, SCC4, SCC9) were xenografted into the right mandibular gland of athymic nude mice. The mice were followed for tumor development and growth, and the mice were sacrificed when they had lost more than 20% of their initial body weight, or the diameter of the induced tumor exceeds 20 mm. After necropsy, the murine tumors were examined histologically and radiologically (micro-positron emission tomography computed tomography) for regional or distant metastasis. We performed immunohistochemical assays with anti-VEGF-C, VEGFR-3, CD105, and D2-40 antibodies. Immunofluorescence double staining for LYVE-1/CD31 was also performed. To quantify the VEGF-C and VEGFR-3 level in the cancer tissue, Western blotting was performed. Finally, we determined the correlation between the degree of expression of VEGF-C/VEGFR-3 and the mean survival time. Results: OSCC tumor cells into the mandibular gland of the nude mice successfully resulted in the formation of recapitulating orthotopic tumor. Tumor cells of the induced tumor did not express VEGF-C. VEGF-C/VEGFR-3 expression was mainly distributed in the endothelial cells of the stromal area. There were no correlation between the degree of expression of VEGF-C/VEGFR-3 and the mean survival time of mice injected with different OSCC cell lines. Conclusion: An recapitulating orthotopic model of OSCC in nude mice was established, which copies the cervical nodal metastasis of human OSCC. Overexpression of lymphangiogenic factors seems to have no effect on survival of hosts in this in vivo experiment.

• Journal of The Korean Society of Grassland and Forage Science
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• v.17 no.3
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• pp.285-292
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• 1997

This study was conducted to obtain the transgenic Brnssica napus plants with tobacco Apase gene using the binary vector system of Agrobacteriurn fumefociens. The results obtained were summarized as follows: A repressible acid phosphatase gene of Saccharon~yces cerevisiae, pho105 was used for screening of tobacco Apase cDNA. In order to identify Apase gene in tobacco genome, Southern blot analysis was pcrformed and the Apase gcnc may be present as a single copy, or at most two or three copies, in tobacco genome. To isolate the tobacco Apase gene, tobacco cDNA library was constructed using purifed mRNA from -Pi treated tobacco root and the plaque forming unit of the library was 2.8 x $10^5$ pfu/m${\ell}$, therefore the library might cover all expressed mRNAs. Using pho5 as a probe. tobacco Apase cDNA was cloned, and restriction mapping and Southern blot analysis of cDNA insert were revealed that the 3.6 kb cDNA contained tobacco acid phosphatase cDNA. Plasmid pGA695 -tcAPl was constructed by subcloning tobacco Apase cDNA into the Hind site of pGA695 with 35s promoter which can be expressed constitutively in plants. The Brassica napus cotyledonary petioles were cocultivated with the ,4 grobacteriunz and transferred to the selection medium. The transformed and regenerated plants were transplanted to soil medium. Southern blot analysis was done on the transformed plants, and it was confirmed that a foregin gene was stably integrated into the genonies of B. nnpus plants.

• Proceedings of the Microbiological Society of Korea Conference
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• 1991.04a
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• pp.225-236
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• 1991

The genes encoding the thermostable $\alpha$-amylase and maltogenic amylase from Bacillus lichenciformis were cloned and expressed in E. coli. The recombinant plasmid pTA322 was found to contain a 3.1kb EcoRI genomic DNA fragment of the thermostable $\alpha$-amylase. The cloned $\alpha$-amylase was compared with the B. licheniformis native $\alpha$-amylase. Both $\alpha$-amylase have the same optimal temperature of $70^{\circ}C$ and are stable in the pH range of 6 and 9. The complete nucleotide sequences of the thermostable $\alpha$-amylase gene were determined. It was composed of one open reading rame of 1,536 bp. Start and stop codons are ATG and TAG. From the amino acid sequence deduced from the nucleotide sequence, the cloned thermostable $\alpha$-amylase is composed of 483 amino acid residues and its molecular weight is 55,200 daltons. The content of guanine and cytosine is $47.46mol\%$ and that of third base codon was $53_41mol\%$. The recombinant plasmid, pIJ322 encoding the maltogenic amylase contains a 3.5kb EcoRI-BamHI genomic DNA fragment. The optimal reaction temperature and pH of the maltogenci amylase were $50^{\circ}C$ and 7, respectively. The maltogenic amylase was capable of hydrolysing pullulan, starch and cyclodextrin to produce maltose from starch and panose from pullulan. The maltogenic amylase also showed the transferring activity. The maltogenic amylase gene is composed of one open reading frame of 1,734bp. Start and stop codons are ATG and ATG. At 2bp upstream from start codon, the nucleotide sequence AAAGGGGGAA seems to be the ribosome-binding site(RBS, Shine-Dalgarno sequence). A putative promoter(-35 and-10 regions) was found to be GTTAACA and TGATAAT. From deduced amino acid sequence from the nucleotide srquence, this enzyme was comosed of 578 amino acid residues and its molecular weight was 77,233 daltons. The content of guanine and cytosine was $48.1mol\%$. The new recombinant plasmid, pTMA322 constructed by inserting the thermostable $\alpha$-amylase gene in the EcoRI site of pIJ322 to produce both the thermostable $\alpha$-amylase and the maltogenic amylase were expressed in the E. coli. The two enzymes expressed from E. coli containing pTMA322 was reacted with the $15\%$ starch slurry at $40^{\circ}C$ for 24hours. The distribution of the branched oligosaccharides produced by the single-step process was of the ratio 50 : 50 between small oligosaccharide up DP3 and large oligosaccharide above DP3.

• 한국생물공학회:학술대회논문집
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• 2005.04a
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• pp.402-406
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• 2005

Lipomyces starkeyi produces a novel glucanhydrolase containing endo-dextranase and ${\alpha}-amylase$ activities. A cDNA from L. starkeyi encoding a dextranase was isolated and characterized. The 2,052 kb cDNA fragment (lsd1) carrying dextranase gene showed one open reading frame (ORF) composed of 1,824 bp flanked by a 41 bp 5'-UTR and a 184 bp 3'-UTR including a poly(A) tail of 27 bp. The ORF encodes for a 608 amino acid with a predicted molecular mass of 67.6 kDa. There was 77% deduced amino acid sequence identity between the LSD1 dextranase and the dextranase from Penicillium minioluteum. The primary structure of the dextranase from L. starkeyi has distant similarity with enzymes belonging to glycosyl hydrolase family 49. The lsd1 protein was expressed in the Saccharmyces cerevisiae under control of GAL1 promoter and active dextranase was produced.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.1
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• pp.13-16
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• 2001

Bovine ${\beta}1$-adrenergic receptor (AR) cDNA was cloned using degenerative primers. Bovine ${\beta}1$-AR coded for 467 amino acids and the comparison of the deduced amino acid sequence with that of sheep showed 93.4% identity. Northern blot analysis indicated that transcript size for the bovine ${\beta}1$-AR was 3.6 kb in the adipose tissue. The expression level of three $\beta$-ARs (1, 2, and 3) in bovine abdominal, subcutaneous, and perirenal adipose tissues were examined using reverse transcription-polymerase chain reaction (RT-PCR), and the levels of ${\beta}1$- and ${\beta}3$-AR mRNA were found to be lower in the subcutaneous adipose tissue than in the abdominal and perirenal adipose tissues. These results suggest that the expression of $\beta$-ARs mRNA are differentially regulated among the adipose tissues.

• Journal of the Korean Magnetic Resonance Society
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• v.11 no.2
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• pp.73-84
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• 2007

MTH1821 (UniProtKB/TrEMBL ID O27849) is a 96-residue hypothetical protein from the open reading frame of Methanobacterium thermoautotrophicum H one of the target organisms of structural genomics pilot project. Proteins which contain conserved sequence compared with MTH1821 have not been discovered yet and the functional and structural information for MTH1821 is not available. Here, we present the sequence-specific backbone resonance using multidimensional heteronuc1ear NMR spectroscopy and propose the secondary structure using GetSBY software. The backbone resonances of N, HN, $C_{\alpha}$, $C_{\beta}$, CO and $H_{\alpha}$ which are necessary for a prediction of secondary structure by GetSBY were assigned about 98% (557/568). The secondary structure of MTH1821 confirmed that it is comprised of four strand regions and two helical regions. This report will provide a valuable resource for the calculation solution structure of MTH1821 and for the other hypothetical protein that is targeted for structural-based functional discovery.