• Korean Journal of Microbiology
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• v.27 no.4
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• pp.317-322
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• 1989

A gene can be selectively overexpressed in E. coli by utilizing the phage T7 RNA polymerase's stringent recognition and active transcription of the T7 promoter. The T7 expression system was constructed such that the T7 RNA polymerase gene is under the control of lacUV5 promoter in one plasmid, and that the target gene, the promoterless chloramphenicol acetyltransferase (CAT) gene with E. coli ribosome binding site is under the control of T7 promoter in the other plasmid. Only the E. coli cells containing both plasmids show high resistance to chloramphenicol. When the copy number of the runaway plasmid containing the polymerase gene was varied by a temperature shift, amounts of the CAT protein synthesized upon induction was correspondingly changed as shown in SDS gel electrophoresis.

• Archives of Pharmacal Research
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• v.21 no.4
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• pp.470-474
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• 1998

A kanamycin producer, Streptomyces kanamyceticus IFO 13414 is highly resistant to kanamycin. Cloning of the kanamycin resistance genes in S. lividans 1326 with pIJ702 gave several kanamycin resistant transformants. Two transformants, S. lividans SNUS 90041 and S. lividan. SNUS 91051 showed similar resistance patterns to various aminoglycoside antibiotics. Gene mapping experiments revealed that plasmids pSJ5030 and pSJ2131 isolated from the transformants have common resistant gene fragments. Subcloning of pSJ5030 gave a 1.8 Kb gene fragment which showed resistance to kanamycin. Cell free extracts of S. lividans SNUS 90041, S. lividans SNUS 91051 and subclone a S. lividans SNUS 91064 showed kanamycin acetyltransferase activity. The detailed gene map is included.

• Journal of Microbiology and Biotechnology
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• v.17 no.4
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• pp.681-684
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• 2007

An immunoassay method was developed to quantitatively detect phosphinothricin-N-acetyltransferase (PAT) encoded by the Bialaphos resistance (bar) gene in genetically modified (GM) pepper. The histidine-tagged PAT was overexpressed in Escherichia coli M15 (pQE3l-bar) and efficiently purified by $Ni^{2+}$ affinity chromatography. A developed sandwich enzyme-linked immunosorbent assay (S-ELISA) method (detection limit: $0.01{\mu}g/ml$) was 100-fold more sensitive than a competitive indirect ELISA (CI-ELISA) method or Western blot analysis in detecting the recombinant PAT. In real sample tests, PAT in genetically modified herbicide-tolerant (GMHT) peppers was successfully quantified [$4.9{\pm}0.4{\mu}g/g$ of sample (n=6)] by the S-ELISA method. The S-ELISA method developed here could be applied to other GMHT crops and vegetables producing PAT.

• Korean Journal of Biological Psychiatry
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• v.17 no.4
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• pp.218-225
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• 2010

Objectives : The potential association between choline acetyltransferase(CHAT) polymorphism and the risk of mild cognitive impairment(MCI) has not been investigated in Korea. We examined the main effect of CHAT polymorphism and its interaction with apolipoprotein E(APOE) polymorphism in the development of MCI in elderly Korean sample. Methods : We analyzed CHAT 2384G > A polymorphism and APOE polymorphism among 149 MCI subjects with MCI and 298 normal controls. We tested the association between MCI and CHAT A allele status using a logistic regression model. In addition, we employed generalized multifactor dimensionality reduction(GMDR) to investigate the interaction between CHAT and APOE with regard to the risk of MCI. Results : The CHAT A allele was associated with AD risk(OR = 1.59, 95% CI = 1.02-2.48, p = 0.042). No significant gene-gene interaction between CHAT and APOE was found in GMDR method(testing balanced accuracy = 0.540, p = 0.055). Conclusion : The CHAT A allele was associated with MCI risk in the Korean elderly. Its interaction with the APOE ${\varepsilon}4$ allele was not significant with regard to the development of MCI.

• Korean Journal of Weed Science
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• v.17 no.4
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• pp.390-399
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• 1997

This experiment was conducted to introduce PAT (phosphinothricin acetyltransferase, non-selective herbicide bialaphos resistant gene) gene into potato (Solanum tuberosum. cv. Desiree). Optimal shoot regeneration from leaf discs and stem segments was obtained in MS medium supplemented with 0.1 mg/L IBA and 0.5 mg/L BA, and the frequency of shoot regeneration was 54% in left discs and 46% in stem segments. In this condition, leaf discs and stem segments of potato were co-cultivated with A. tumefaciens MP90 which contained binary vector with GUS: :NPTII gene and PAT gene. Transgenic shoots were regenerated from leaf and stem-derived calli on selection medium with 100mg/L kanamycin. The 100${\mu}M$ acetosyringone treatment during the co-cultivation highly enhanced(4 times than the control) the shoot regeneration on selection medium. When the putative transgenic plants were transferred to medium with 10mg/L basta, all of them were survived. After PCR. GUS test, and Southern blot analysis of the survived plant, we confirmed that the gene was stably integrated into the potato genome and expressed. After the transgenic plants were transplanted in soil, and the transgenic plants were sprayed with the herbicide basta (300ml/10a), the transgenic plants remained green but control plants were died.

• Journal of Microbiology
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• v.40 no.1
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• pp.43-50
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• 2002

A cluster of genes encoding the pyruvate dehydrogenase complex (PDC) of Streptomyces seoulensis, a Gram-positive bacterium, was cloned and sequenced. The genes of S. seoulensis consist of four open reading frames. The first gene, lpd, which encodes a lipoamide dehydrogenase, is followed by pdhB encoding a dihydrolipoamide acetyltransferase (E2p), pdhR, a regulatory gene, and pdhA encoding a pyruvate dehydrogenase component (Elp). Elp had an unusual homodimeric subunit, which has been known only in Gram-negative bacteria S. seoulensis E2p contains two lipoyl domains like those of humans and Streptomyces faecalis. The pdhR gene appears to be clustered with the structural genes of S. seoulensis PDC. The PdhR-overexpressed S. seoulensis howed growth retardation and the decrease of Elp, indicating that PdhR regulates the function of PDC by repressing the expression of Elp. A strain of Streptomyces licidans overexpressing S. seoulensis PdhR showed a significant decreasein the level of actinorhodin, implying a regulatory role for Streptomyces PDC in antibiotic biosynthesis.

• BMB Reports
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• v.31 no.5
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• pp.484-491
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• 1998

Acetylation of lysine residues within the aminoterminal domains of the core histones plays a critical role in chromatin assemhly as well as in regulation of gene expression. To study the biochemical function of histone acetylation, we have cloned a cDNA encoding the catalytic subunit of human histone acetyltransferase, Hat1. Analysis of the predicted amino acid sequence of human Hat1 revealed an open reading frame of 419 amino acids with a calculated molecular mass of 49.5 kDa and an isoelectric point of 5.5. The amino acid sequence of human Hat1 is homologous to those of known and putative Hat1 proteins from various species throughout the entire open reading frame. The recombinant human Hat1 protein expressed in bacteria possesses histone H4 acetyltransferase activity in vitro. Both RbAp46 and RbAp48, which participate in various processes of histone metabolism, enhance the histone acetyltransferase activity of the recombinant human Hat1, indicating that they are both able to functionally interact with the human Hat1 in vitro.

• Korean Journal of Microbiology
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• v.40 no.2
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• pp.87-93
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• 2004

Dynamic modification of cytoplasmic and nuclear proteins by O-linked N-acetylglucosamine (O-GlcNAc) on Ser and Thr residues is ubiquitous in higher eukaryotes. And this modification may serve as a signaling mod-ification analogous to protein phosphorylation. Addition and cleavage of O-GlcNAc are catalyzed by O-linked GlcNAc transferase (OGT) and O-linked N-acety1glucosaminidase (O-GlcNAcase), respectively. Two types of human O-GlcNAcase gene were cloned and expressed as three fusion proteins in Escherichia coli. O-GlcNA-case activity showed in the order of thioredoxin fusion> $6{\times}His$ tag> GST fusion. O-GlcNAcase had enzy-matic activity against only ${\rho}$NP-GlcNAc of seven tested substrate analogs. Blast search revealed that O-GlcNAcase has two conserved domains, amino terminal hyaluronidase-like domain and carboxy terminal N-acetyltransferase domain. Extensive deletion studies were done to define catalytically important domains. The deletions of hyaluronidase-like domain and N-acetyltransferase domain abolished enzyme activity. But, N-ter-minal 55 amino acid deletion and C-terminal truncation showed lower activity. Based on deletion analysis, we suggest that hyaluronidase-like domain is essential for enzyme activity and carboxy terminal N-acetyltrans-ferase domain may be modulatory function.

• Korean Journal of Microbiology
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• v.36 no.4
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• pp.273-278
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• 2000

The aacCl, aacC2, aacC3, and aacC4 genes, which encode aminoglycoside acetyltransferase AAC(3)-I, AA(3)-II, AAC(3)-III, and AAC(3)-IV, respectively, and aerolysin genes in water samples from Juam lake were surveyed by polymerase chain reaction. Surface water was collected from January 1996 to December 1998, and then bacterial DNA was extracted from the water. Twelve samples were tested by PCR to servey the genes for aminoglycoside acetyltransferase and aerolysin in the lake water. The aacC2 gene was detected in 9 of 12 DNA samples. Among 9 samples showing aacC2 positive, 7 samples were associated with Tn3 sequence. However, none of the twelve samples amplified the expected DNA fragment for aacC1, aacC3, and aacC4 genes. PCR primer to detect the aerolysin gene was designed using the conserved region of the genes for aerolysin and hemolysin of Aeromonas spp. This primer set successfully amplified the expected 414 bp PCR product with the DNA samples from the lake water. The aerolysin gene was detected in 7 of 12 DNA samples. When Southern hybridization of the gel with probe was performed, the aerolysin gene was detected in 10 of 12 DNA samples. However, the seasonal fluctuation of these genes was not found.

• Korean Journal of Microbiology
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• v.27 no.3
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• pp.194-200
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• 1989

The nucleotide sequence of inducible chloramphenicol acetyl-transferase(CAT) gene isolated from a small plasmid pSBK203 of Staphylococcus aureus was determined. The base sequence shows that structural gene of pSBK203-CAT encodes a protein of 213 amino acids and has a leader region which encodes a short polypeptide of 9 amino-acids in its upstream. vertical bar /sup 35/S vertical bar-Methionine labelled CAT gene product in minicell showed almost same mobility with pC194-CAT of which molecular weight is 24Kdal on polyacrylamide gel electrophoresis. Predicted amino acid sequence of pSBK203-CAT has revealed a high degree of homology with the CATs of pC194 and pC221 than those of cat-86, Tn9 and proteus mirabilis PM13.