• Journal of Microbiology
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• v.35 no.4
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• pp.327-333
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• 1997

Although extensively characterized in human cells, no heterogeneous nuclear ribonucleoprotein(hnRNP) has been found in the fission yeast Schizosaccharomyces pombe which is amenable to genetic studies and more similar to mammals than Saccharomyces cerevisiae is in terms of RNA processing. As a first step to characterize hnRNPs from S. pombe, attempt was made to find human hnRNP A1 homologs from S. pombe. The RNA molecule (A1 winner) containing the consensus high-affinity hnRNP A1 binding site (UAGGGA/U) was synthesized in vitro and used in an ultraviolet(UV) light-induced protein-RNA cross-linking assay. A number of S, pombe proteins bound to the A1 winner RNA. An approximately 50-kDa protein(p50) cross-linked more efficiently to the A1 winner RNA than other proteins. The p50 protein did not cross-link to a nonspecific RNA, but rather to the A1-5’ SS RNA in which the consensus 5’ splice junction sites of S. pombe introns were abolished. This suggests that the p50 protein, however, did not bind to the single-stranded DNA to shich the human hnRNP A1 could bind and be eluted with 0.5M NaCl. Further analysis should reveal more features of this RNA-binding protein.

• Journal of Microbiology
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• v.45 no.6
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• pp.534-540
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• 2007

The Streptomyces coelicolor A3(2) genome contains six operons (rrnA to F) for ribosomal RNA synthesis. Transcription from rrnD occurs from four promoters (p1 to p4). We found that transcripts from the p1 and p3 promoters were most abundant in vivo in the early exponential phase. However, at later phases of exponential and stationary growth, transcripts from the p1 promoter decreased drastically, with the p3 and p4 transcripts constituting the major forms. Partially purified RNA polymerase supported transcription from the p3 and p4 promoters, whereas pure reconstituted RNA polymerase with core enzyme (E) and the major vegetative sigma factor ${\sigma}^{HrdB}$ ($E{\cdot}{\sigma}^{HrdB}$) did not. In order to assess any potential requirement for additional factor(s) that allow transcription from the p3 and p4 promoters, we fractionated a partially purified RNA polymerase preparation by denaturing gel filtration chromatography. We found that transcription from the p3 and p4 promoters required factor(s) of about 30-35 kDa in addition to RNAP holoenzyme ($E{\cdot}{\sigma}^{HrdB}$). Therefore, transcription from the p3 and p4 promoters, which contain a consensus -10 region but no -35 for ${\sigma}^{HrdB}$ recognition, are likely to be regulated by transcription factor(s) that modulate RNA polymerase holoenzyme activity in S. coelicolor.

• Journal of Microbiology and Biotechnology
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• v.8 no.5
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• pp.449-454
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• 1998

A thennostable chitosanase was purified from Bacillus sp. KFB-C108, by fractionation of 30 to 70% saturation with ammonium sulfate, DEAE-Toyopearl chromatography, Butyl-Toyopearl chromatography, and TSK-Gel HW-55F gel filtration. The purified enzyme showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the molecular weight was estimated to be 48 kDa. The enzyme degraded soluble chitosan and colloidal chitosan, but did not degrade glycol chitosan, chitin, and the other compounds investigated. There was no effect on the chitosanase activity by treatment with chelating agents, alkylating agents, and various metals investigated, and only cobalt ions inhibited the activity. Optimum temperature and pH were $55^{\circ}C$ and 6.5, respectively. The enzyme was stable after heat treatment at $80^{\circ}C$ for 10 min or $70^{\circ}C$ for 30 min and fairly stable in several organic solvents as well. Chitosan was hydrolyzed to $(GlcN)_4$as a major product by incubation with the enzyme.

• Molecules and Cells
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• v.24 no.3
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• pp.437-440
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• 2007

$OGL-20P^T$-358 is a novel 66 amino acid residue protein from the hyperthermophile Thermococcus thioreducens sp. nov., strain $OGL-20P^T$, which was collected from the wall of the hydrothermal black smoker in the Rainbow Vent along the mid-Atlantic ridge. This protein, which has no detectable sequence homology with proteins or domains of known function, has a calculated pI of 4.76 and a molecular mass of 8.2 kDa. We report here the backbone $^1H$, $^{15}N$, and $^{13}C$ resonance assignments of $OGL-20P^T$-358. Assignments are 97.5% (316/324) complete. Chemical shift index was used to determine the secondary structure of the protein, which appears to consist of primarily ${\alpha}$-helical regions. This work is the foundation for future studies to determine the three-dimensional solution structure of the protein.

• Molecules and Cells
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• v.20 no.1
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• pp.43-50
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• 2005

Glutaredoxin (Grx) is a small, heat-stable redox protein acting as a multi-functional glutathione (GSH)-dependent disulfide oxidoreductase. We have cloned the monothiol Grx5 gene from the genomic DNA of the fission yeast Schizosaccharomyces pombe. It has 1,904 bp, with one intron, and encodes a putative protein of 146 amino acids with a molecular mass of 16.5 kDa. Recombinant Grx5 produced functional Grx in S. pombe cells. NO-generating sodium nitroprusside (SNP, 1.0 and 2.0 mM) and potassium chloride (KCl, 0.2 and 0.5 M) increased the synthesis of ${\beta}$-galactosidase from a Grx5-lacZ fusion gene, and transcription of Grx5 was also enhanced by SNP and KCl. Synthesis of ${\beta}$-galactosidase from the Grx5-lacZ fusion was lower in Pap1-negative TP108-3C cells than in wild type KP1 cells, and when Pap1 was overproduced in KP1 cells, the level of ${\beta}$-galactosidase increased. We also found that Pap1 is involved in the induction of Grx5 by SNP and KCl. S. pombe Grx5 may play a crucial role in responses to nitrosative and osmotic stresses.

• Journal of Microbiology and Biotechnology
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• v.18 no.10
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• pp.1672-1677
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• 2008

IscR (iron-sulfur cluster regulator) has been reported to be a repressor of the iscRSUA operon, and in vitro transcription reactions have revealed that IscR has a repressive effect on the iscR promoter in the case of [$Fe_{2}S_{2}$] cluster loading. In the present study, the iscR gene from A. ferrooxidans ATCC 23270 was cloned and successfully expressed in Escherichia coli, and then purified by one-step affinity chromatography to homogeneity. The molecular mass of the IscR was 18 kDa by SDS-PAGE. The optical and EPR spectra results for the recombinant IscR confirmed that an iron-sulfur cluster was correctly inserted into the active site of the protein. However, no [$Fe_{2}S_{2}$] cluster was assembled in apoIscR with ferrous iron and sulfide in vitro. Therefore, the [$Fe_{2}S_{2}$] cluster assembly in IscR in vivo would appear to require scaffold proteins and follow the Isc "AUS" pathway.

• Food Science and Biotechnology
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• v.17 no.3
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• pp.489-494
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• 2008

In order to develop a potent antidementia $\beta$-secretase inhibitor from phytochemicals, $\beta$-secretase inhibitory activities of extracts from many medicinal plants and herbs were determined. Water extracts from Rubus coreanus showed the highest $\beta$-secretase inhibitory activity of 84.5%. After purification of the $\beta$-secretase inhibitor from R. coreanus using systematic solvent extraction, ultrafiltration, Sephadex G-10 column chromatography, and reverse-phase high performance liquid chromatography (HPLC), a purified $\beta$-secretase inhibitor with $IC_{50}$ inhibitory activity of $6.3{\times}10^3\;ng/mL$ ($1.56{\times}10^{-6}\;M)$ was obtained with a 0.08% solid yield. The molecular mass of the purified $\beta$-secretase inhibitor was estimated to be 576 Da by liquid chromatography-mass spectrometry (LC-MS) and $\beta$-secretase inhibitor also is a new tetrapeptide with the sequence Gly-Trp-Trp-Glu. The purified $\beta$-secretase inhibitory peptide inhibited $\beta$-secretase non-competitively and also show less inhibition on trypsin, however no inhibition on other proteases such as $\alpha$-secretase, chymotrypsin, and elastase.

• Journal of Microbiology and Biotechnology
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• v.26 no.1
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• pp.20-27
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• 2016

Lactobacillus kefiranofaciens ZW3 was obtained from kefir grains, which have high lactose hydrolytic activity. In this study, a heterodimeric LacLM-type β-galactosidase gene (lacLM) from ZW3 was isolated, which was composed of two overlapping genes, lacL (1,884 bp) and lacM (960 bp) encoding large and small subunits with calculated molecular masses of 73,620 and 35,682 Da, respectively. LacLM, LacL, and LacM were expressed in Escherichia coli BL21(DE3) and these recombinant proteins were purified and characterized. The results showed that, compared with the recombinant holoenzyme, the recombinant large subunit exhibits obviously lower thermostability and hydrolytic activity. Moreover, the optimal temperature and pH of the holoenzyme and large subunit are 60℃ and 7.0, and 50℃ and 8.0, respectively. However, the recombinant small subunit alone has no activity. Interestingly, the activity and thermostability of the large subunit were greatly improved after mixing it with the recombinant small subunit. Therefore, the results suggest that the small subunit might play an important role in maintaining the stability of the structure of the catalytic center located in the large subunit.

• Biomedical Science Letters
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• v.18 no.3
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• pp.310-312
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• 2012

Cytochrome $b_5$ is involved in the reduction of methemoglobin back to hemoglobin, thereby maintaining normal function of the blood to carry oxygen around. Congenital abnormal condition of this enzyme causes a rare disease called methemoglobinemia. At least 4 different retropseudogenes are reported so far for cytochrome $b_5$. However, type III pseudogene has attracted most attention because it contains open reading frame in its structure. Although there is no evidence yet if this pseudogene is actually expressed in the cell or the blood the possibility of its expression needs to be elucidated. We have isolated type III pseudogene by polymerase chain reaction and cloned into pGEX-4T-1 expression vector followed by SDS-PAGE. Protein was expressed and the size of the expressed protein was 28 kDa as expected in its genetic code. This result also shows that the protein is not harmful for the viability of the microorganism. This study may contribute to the genetic diagnosis of cardiac diseases, possibly caused by cytochrome $b_5$.

• The Journal of Industrial Distribution & Business
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• v.11 no.2
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• pp.17-23
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• 2020

Purpose: The purpose of this study is to understand the effect of the unique product recommendation system on customer satisfaction. Research design, data and methodology: The survey method used the self-recording way in which the respondents selected for the study and distributed 300 questionnaires, and with due personal care, researchers collected all the distributed questionnaires. Results: The result implies that the characteristics of the product recommendation system should be more secure and developed. Conclusions: The aspects of the product recommendation system were selected as factors of price fairness, accuracy, and quality through previous studies, and the empirical analysis of the effect of the characteristics of the product recommendation system on customer satisfaction was summarized as follows. Among the attributes of the product recommendation system, the attributes of price fairness, accuracy, and quality affect customer satisfaction. Among them, the beta value of quality was the highest, and the effect of quality was the largest among the three factors. Based on the results of the study, the implications for the characteristics of the product recommendation system are summarized as follows. The aspects of the product recommendation system have a positive effect on customer satisfaction, so it is necessary to fill the needs of consumers based on the survey focused on quality