• Applied Biological Chemistry
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• v.38 no.1
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• pp.55-62
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• 1995

To understand the molecular structure of Korean garlic viruses, cDNA cloning of virus genomic RNA was attempted. Virus particles were isolated from virus-infected garlic leaves and a cDNA library was constructed from garlic virus RNA. One of these clones, S81, selected by random sequencing has been identified as a member of potexvirus group other than potyvirus and carlavirus. The clone is 873 bp long contains most of the coat protein (CP) coding region and 3'-noncoding region including poly(A) tail. A putative polyadenylation signal sequence (AAUAAA) and the hexanucleotide motif (ACUUAA), a replicational cis-acting element conserved in the 3'-noncoding region of potexvirus RNAs are noticed. The clone S81 shows about 30-40% identity in both nucleotide and amino acid sequences with CPs of potexviruses. The genome size of the virus was analysed to be 7.46 knt by Northern blot analysis, which was longer than those of other potexviruses. The open reading frame encoding CP was expressed as a fusion protein (S81CP) in Escherichia coli and the recombinant protein was purified by immobilized metal binding affinity chromatography. Polyclonal antibody was raised against S81CP in rabbit to examine the occurrence of garlic potexvirus in Korean garlic plants by immunoblot analysis. Two virus protein bands of Mr 27,000 and 29,000 from garlic leaf extract of various cultivars reacted with the antibody. It was shown that Mr 27,000 band might not be a degradation product of Mr 29,000 band, suggesting that two types of potexvirus different in size of coat protein could exist in Korean garlic plants.

• Journal of Veterinary Clinics
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• v.26 no.2
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• pp.123-129
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• 2009

Patent ductus arteriosus(PDA) is an abnormal shunt between the descending aorta and pulmonary artery through the incompletely closed ductus arteriosus and is the most common congenital heart defect in dogs. Recent human genetic studies found that a the gene mutation in transcription factor AP-2 beta(TFAP2B) was responsible for syndromic cases of PDA. Mutations in the TFAP2B gene are associated with certain congenital cardiac defects in humans that include PDA. In this study, we isolated the entire coding exons of canine TFAP2B gene for genetic screening in dogs with PDA. Analysis of the deduced amino acid sequence suggested that the canine TFAP2B are phylogenetically closer to the human TFAP2B(100% identity in amino acid sequence) than mouse and rat. In cTFAP2B gene screening, one single c.936+203G>A base change was found in affected Maltese dogs with PDA. However, further screening found the same base change in one unaffected control dog, suggesting this base change might be polymorphism. No other base changes were found in other dog breeds enrolled in this study. Because the base change was located in the intronic region and found in an unaffected control dog, TFAP2B might not be responsible for familial PDA in Malteses and sporadic cases of other dog breeds, although the gene promoter region should be investigated before reaching to this conclusion. A future study that may take this study further would be to collect more samples and to screen TFAP2B in various breeds of dogs with PDA and other various congenital heart defects.

• Microbiology and Biotechnology Letters
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• v.38 no.2
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• pp.151-157
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• 2010

Barley malt produces two different $\alpha$-amylase isozymes (AMY1 and AMY2), which share up to 80% of amino acid sequence identity with each other. However, their enzymatic properties differ remarkably. In this study, five chimeric enzymes between AMY1 and 2 were constructed by staggered extension process (StEP) technique, and their enzymatic properties were characterized. According to the results, chimeric AMY-D2, D8, and E12 showed the mixed or intermediate types of calcium-dependent activity between AMY1 and 2. Meanwhile, only AMY-E10 chimera could be significantly inhibited by barley $\alpha$-amylase/subtilisin inhibitor (BASI) protein. Chimera AMY-C6 showed the same calcium-dependency as AMY1, while AMY-E10 was closely similar to AMY2. As a result, it can be proposed that some amino acid residues in the region II, III, and IV of barley $\alpha$-amylases can play very important roles in the interaction with BASI, and those in III, V, VI, and VII may partly affect on the calcium-dependent activity.

• Journal of Life Science
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• v.14 no.4
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• pp.636-643
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• 2004

D-Xylose isomerase produced by Lactococcus sp. JK-8, isolated from kimchi, was purified 17-fold of homogeneity, and its physicochemical properties were determined. Although the N-terminal amino acid sequence of D-xylose isomerase was analysed to Ala-Tyr-Phe-Asn-Asp-Ile-Ala-Pro-Ile-Lys, it was not similar to that of Lactobacillus enzyme. The molecular weight of the purified enzyme was estimated to be 180 kDa by gel filtration, 45 kDa by SDS-PAGE and the enzyme was homotetramer. The optimum pH of the enzyme was around 7 and stable between pH 6 and 8. The optimum reaction temperature was 7$0^{\circ}C$ and stable up to 7$0^{\circ}C$ in the presence of 1 mM $Mn^{2+}$. Like other D-xylose isomerases, this enzyme required divalent cation, such as $Mg^{2+}$, $Co^{2+}$, or $Mn^{2+}$ for the activity and thermostability. $Mn^{2+}$was the best activator. Substrate specificity studies showed that this enzyme was highly active on D-xylose. The enzyme had an isoelectric point of 4.8, and fm values for D-xylose was 5.9 mM.

• Korean Journal of Microbiology
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• v.41 no.2
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• pp.99-104
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• 2005

Corynebacterial clones which exert regulatory effects on the expression of the glyoxylate bypass genes were isolated using a reporter plasmid carrying the enteric lacZ fused to the aceB promoter of Corynebacterium glutamicum. Some clones carried common fragments as turned out by DNA mapping technique. Subcloning analysis followed by the measurement of $\beta-galactosidase$ activity in Escherichia coli identified the region responsible for the aceB-repressing activity. Sequence analysis of the DNA fragment identified two independent ORFs of ORF1 and ORF2. Among them, ORF2 was turned out to be responsible for the aceB-repressing activity. ORF1 encoded a 23,216 Da protein composed of 206 amino acids. Sequence similarity search indicated that the ORF may encode a ECF-type $\sigma$ factor and designated sigH. To identify the function of sigH, C. glutamicum sigH mutant was constructed by gene disruption technique and the sigH mutant showed growth retardation as compared to the wild type strain. In addition, the mutant strain showed sensitivity to oxidative-stress generating agent plumbagin. This result imply that sigH is probably involved in the stress response occurring during normal cell growth.

• Journal of Life Science
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• v.21 no.12
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• pp.1666-1677
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• 2011

The regulation of gene expression plays an important role in cell cycle controls. In this study, a novel gene, the $mas1^+$($\underline{mi}$tosis $\underline{as}$sociated protein) gene, a homolog of human CIP29/Hcc1, was isolated and characterized from fission yeast Schizosaccharomyces pombe (S. pombe) using a gene-specific polymerase chain reaction. The isolated gene contained a complete open reading frame capable of encoding 245 amino acid residues with a typical promoter, as judged by nucleotide sequence analysis. It was also found that a PCB ($\underline{p}$ombe cell $\underline{c}$ycle $\underline{b}$ox) is located in the promoter region, which controls M-$G_1$ specific transcription in S. pombe. The quantitative analysis of the $mas1^+$ transcript against $adh1^+$ showed that the pattern of expression is similar to that of the septation index. Cytokinesis of mas1 mutant was greatly delayed at $25^{\circ}C$ and $36^{\circ}C$, and a large number of multi-septate cells were produced. The mas1 mutant had 2C, 4C and 6C DNA contents, as determined by FACS analysis. In addition, the number of multi-septate cells significantly increased. When cells were cultured in nitrogen starvation medium to increase proliferation, the abnormal phenotypes of mas1 mutant dramatically increased. These phenotypes could be rescued by an overexpression of the $mas1^+$ gene. The mas1 protein localized in the nuclei of S. pombe and human HeLa cells, as evidenced by Mas1-EGFP signals. The abnormal growth pattern and the morphology of mas1 mutant were complemented by a plasmid carrying human CIP29/Hcc-1cDNA. In addition, CIP29 /Hcc-1 transcript level increased in active cell proliferation stages in the developing mouse embryos. These results indicate that the $mas1^+$ ishomologous to the human CIP29/Hcc1 gene and is involved in cytokinesis and cell shape control.

• Clinical and Experimental Pediatrics
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• v.48 no.4
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• pp.425-432
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• 2005

Purpose : Polyglutamine diseases are a group of diseases caused by the expansion of a polyglutamine tract in the protein. The present study was performed to verify if polyglutamine disease transgenic Drosophila models show similar dysfunctions as are seen in human patients. Methods : Polyglutamine disease transgenic Drosophila were tested for their climbing ability. And using genetic methods, the effects of anti-apoptotic gene bcl-2 and chemical chaperones on neurodegeneration were observed. Also, spinocerebellar ataxia 2 (SCA2) transgenic Drosophila lines were generated for future studies. Results : Expanded forms of spinocerebellar ataxia 3 (SCA3) transgenic protein causes characteristic locomotor dysfunction when expressed in the nervous system of Drosophila but the anti-apoptotic gene bcl-2 shows no evidence of ameliorating the deleterious effect of the expanded protein. However, Glycerol, a chemical chaperone, seemed to reduce the toxicity, at least in the eyes of the transgenic flies. The level SCA2 expression is too weak in the transgenic SCA2 Drosophila for evaluation. Conclusion : SCA3 transgenic Drosophila show ataxic behavior as observed in human patients. Chemical chaperones such as glycerol may prove beneficial in this class of genetic disease, which has no current method of cure.

• Korean journal of applied entomology
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• v.45 no.1 s.142
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• pp.15-24
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• 2006

Inhibitor kB (IkB)-like gene has been found in the genome of Cotesia plutellae bracovirus (CpBV), which is the obligatory symbiont of an endoparsitoid wasp, C. plutellae. The open reading frame of CpBV-IkB was 417 bp and encoded 138 amino acids. Four ankyrin repeat domains were found in CpBV-IkB, which shared high homology with other known polydnavirus IkBs. Considering a presumptive cellular IkB based on Drosophila Cactus, CpBV-IkB exhibited a truncated structure with deletion of signal-receiving domains, which suggested its irreversible inhibitory role in NFkB signal transduction pathway of the parasitized host in response to the wasp parasitization. CpBV-IkB was expressed only in the parasitized diamondback moth, Plutella flostella. Its expression was estimated by quantitative RT-PCR during parasitization period, showing a constitutive expression pattern from the first day of parasitization. An indirect functional analysis of CpBV-IkB was conducted and suggested a hypothesis of host antivirus inhibition.

• Korean Journal of Microbiology
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• v.50 no.3
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• pp.210-215
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• 2014

Pseudomonas nitroreducens TX1 was isolated from a rice field drainage in Taiwan. The bacterium is of special interest because of its capability to use a group of nonionic surfactants such as alkylphenol polyethoxylates even at high concentrations as a sole carbon source. In this study, a small cryptic circular plasmid, pTX1, was characterized from P. nitroreducens TX1. It is 2,286 bp in length with a GC content of 63.3% and harbors three open reading frames, $Rep_{pTX1}$ and functionally unidentified ORF1 and ORF2. The predicted $rep_{pTX1}$ gene product is homologous to Rep proteins of plasmids belonging to the pC194/pUB110 family, which is predominantly found in Gram-positive bacteria and is known to replicate by the rolling-circle mechanism. The copy number of pTX1 was estimated to be about 150 in each cell. Based on the genetic fingerprints and comparison with other plasmids, it is concluded that pTX1 replicates by a rolling circle mechanism which is rarely found for Pseudomonas plasmids.

• Journal of Plant Biotechnology
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• v.32 no.3
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• pp.167-173
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• 2005

Three different cDNAS for cinnamate 4-hydroxylase (C4H) which are involved in the second step of the general phenylpropanoid pathway were isolated and designated as pc4h1 (1,755 bp), pc4h2 (1,655 bp), and pc4h3 (1,316 bp), respectively. The nucleotide sequence analysis revealed that both pc4h1 and pc4h2 clones encode polypeptides of 505 amino acids frame but pc4h3 clone was truncated at the 5'-end of coding region. The alignment of the deduced amino acid sequences showed that PC4H1 and PC4H2 are highly homologous (95.8% identical) with each other and contain three conserved domains which are typical in cytochrome P450 monooxygenase: proline-rich region, threonine-containing binding pocket for the oxygen molecule, and heme binding region. In addition, result of the phylogenic tree analysis revealed that both pepper C4Hs belong to Class 1. pc4h2 transcription was strongly induced in wounded fruit (400%) and root (200%) relative to its very low basal level but not in leaf or stem tissue. In case of pc4h1, the basal level of transcription was higher than pc4h2 but induction by wounding was lower in fruit and root while leaf and stem tissues did not respond to wounding. The basal level of pc4h3 transcripts was not, if any, detectable and response to wounding was not observed.